WO2024228964A1

WO2024228964A1 - Treatments comprising an mtor inhibitor nanoparticle composition

Info

Publication number: WO2024228964A1
Application number: PCT/US2024/026882
Authority: WO
Inventors: Erlinda M. Gordon; Neil P. Desai; Anita N. SCHMID
Original assignee: Aadi Bioscience Inc
Current assignee: Whitehawk Therapeutics Inc
Priority date: 2023-04-30
Filing date: 2024-04-29
Publication date: 2024-11-07
Anticipated expiration: 2025-10-30

Abstract

The present application, in certain aspects, pertains to methods and compositions for the treatment of cancers, such as undifferentiated pleomorphic sarcoma or leiomyosarcoma, using : (a) a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin; and, optionally, (b) an anti-PD-1 antibody.

Description

TREATMENTS COMPRISING AN MTOR INHIBITOR NANOPARTICLE

COMPOSITION

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the priority benefit of U.S. Provisional Patent Application Serial. No. 63/463,052, filed on April 30, 2023, the entire contents of which are incorporated herein by reference for all purposes.

FIELD OF THE INVENTION

[0002] The present application, in certain aspects, pertains to methods and compositions for the treatment of undifferentiated pleomorphic sarcoma or leiomyosarcoma, using a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin.

BACKGROUND OF THE INVENTION

[0003] Advanced solid malignancies are most often fatal and innovative treatments are urgently needed. The mammalian target of rapamycin (mTOR) is a conserved serine/threonine kinase that serves as a central hub of signaling in the cell to integrate intracellular and extracellular signals and to regulate cellular growth and homeostasis. Activation of the mTOR pathway is associated with cell proliferation and survival, while inhibition of mTOR signaling leads to inflammation and cell death. Dysregulation of the mTOR signaling pathway has been implicated in an increasing number of human diseases, including cancer such as advanced solid malignancies and autoimmune disorders. Consequently, mTOR inhibitors have found wide applications in treating diverse pathological conditions such as solid tumors, hematological malignancies, organ transplantation, restenosis, and rheumatoid arthritis.

[0004] Sirolimus (INN/USAN), also known as rapamycin, is an immunosuppressant drug used to prevent rejection in organ transplantation; it is especially useful in kidney transplants. Sirolimus-eluting stents were approved in the United States to treat coronary restenosis. Additionally, sirolimus has been demonstrated as an effective inhibitor of tumor growth in various cell lines and animal models. Other limus drugs, such as analogs of sirolimus, have been designed to improve the pharmacokinetic and pharmacodynamic properties of sirolimus. For example, Temsirolimus was approved in the United States and Europe for the treatment of renal cell carcinoma. Everolimus was approved in the U. S. for treatment of advanced breast cancer, pancreatic neuroendocrine tumors, advanced renal cell carcinoma, and subependymal giant cell astrocytoma (SEGA) associated with Tuberous Sclerosis. The mode of action of sirolimus is to bind the cytosolic protein FK-binding protein 12 (FKBP12), and the sirolimus-FKBP12 complex in turn inhibits the mTOR pathway by directly binding to the mTOR Complex 1 (mTORCl).

[0005] Albumin-based nanoparticle compositions have been developed as a drug delivery system for delivering substantially water insoluble drugs. See, for example, U. S. Pat.

Nos.5, 916, 596; 6,506,405; 6,749,868, and 6,537,579, 7,820,788, and 7,923,536. Abraxane®, an albumin stabilized nanoparticle formulation of paclitaxel, was approved in the United States in 2005 and subsequently in various other countries for treating metastatic breast cancer. It was recently approved for treating non-small cell lung cancer in the United States, and has also shown therapeutic efficacy in various clinical trials for treating difficult-to-treat cancers such as bladder cancer and melanoma. Albumin derived from human blood has been used for the manufacture of Abraxane® as well as various other albumin-based nanoparticle compositions. Albumin-based nanoparticle composition comprising sirolimus, e.g, «a6-sirolimus or Fyarrao®, are known, e.g, US. Pat. No. 8,911,786 and US Pat. No. 11,497,737.

[0006] However, there remains a continuing need in the art for advanced treatments of certain cancers.

BRIEF SUMMARY OF THE INVENTION

[0007] The present application, in certain aspects, provides a method of treating undifferentiated pleomorphic sarcoma in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and optionally (b) an effective amount of a second therapeutic agent (such as an anti-PD-1 antibody).

[0008] In some embodiments, the undifferentiated pleomorphic sarcoma has phosphatase and tensin homolog (PTEN) loss. In some embodiments, the PTEN loss is a loss-of-function mutation or epigenetic silencing. In some embodiments, the individual is selected for the treatment on the basis of having the PTEN loss. In some embodiments, the method further comprises selecting the individual on the basis of having the PTEN loss. In some embodiments, the undifferentiated pleomorphic sarcoma has a tuberous sclerosis complex 2 (TSC2) mutation. In some embodiments, the TSC2 mutation is a missense mutation, nonsense mutation, deletion, splicing site mutation, insertion, substation, rearrangement, or frameshift, or a combination thereof. In some embodiments, the individual is selected for the treatment on the basis of the TSC2 mutation. In some embodiments, the method further comprises selecting the individual on the basis of having the TSC2 mutation.

[0009] In other aspects, the present application provides a method of treating leiomyosarcoma in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody).

[0010] In some embodiments, the leiomyosarcoma is estrogen receptor-positive leiomyosarcoma. In some embodiments, the individual is selected for the treatment on the basis of having the estrogen receptor-positive leiomyosarcoma. In some embodiments, the method further comprises selecting the individual on the basis of having the estrogen receptor-positive leiomyosarcoma.

[0011] In some embodiments, the undifferentiated pleomorphic sarcoma or the leiomyosarcoma is locally advanced, advanced, malignant, advanced malignant, or metastatic. In some embodiments, the undifferentiated pleomorphic sarcoma or the leiomyosarcoma is relapsed or refractory to a prior treatment. In some embodiments, the prior treatment comprises a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin.

[0012] In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is from about 10 mg/m² to about 150 mg/m² In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 100 mg/m². In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 75 mg/m². In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 56 mg/m². In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 45 mg/m². In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 30 mg/m². [0013] In some embodiments, the mTOR inhibitor nanoparticle composition is administered weekly. In some embodiments, the mTOR inhibitor nanoparticle composition is administered 2 out of every 3 weeks. In some embodiments, the mTOR inhibitor nanoparticle composition is administered on days 8 and 15 of a 21 -day cycle.

[0014] In some embodiments, the mTOR inhibitor nanoparticle composition and the anti- PD-1 antibody are administered concurrently to the individual. In some embodiments, the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered sequentially to the individual. In some embodiments, the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered simultaneously to the individual.

[0015] In some embodiments, the second therapeutic agent (e.g., anti-PD-1 antibody) is administered at an amount of about 1 mg/kg to about 10 mg/kg.

[0016] In some embodiments, the second therapeutic agent (e.g., anti-PD-1 antibody) is administered every three weeks. In some embodiments, the second therapeutic agent (e.g., anti- PD-1) antibody is administered on day 1 of a 21 -day cycle.

[0017] In some embodiments, the second therapeutic agent (e.g., anti-PD-1 antibody) is administered for at least one cycle prior to administration of the mTOR inhibitor nanoparticle composition.

[0018] In some embodiments, the mTOR inhibitor is a limus drug. In some embodiments, the limus drug is sirolimus.

[0019] In some embodiments, the average diameter of the nanoparticles in the composition is no greater than about 150 nm. In some embodiments, the average diameter of the nanoparticles in the composition is no greater than about 120 nm.

[0020] In some embodiments, the weight ratio of the albumin to the mTOR inhibitor in the nanoparticle composition is no greater than about 9:1.

[0021] In some embodiments, the nanoparticles comprise the mTOR inhibitor associated with the albumin. In some embodiments, the nanoparticles comprise the mTOR inhibitor coated with the albumin.

[0022] In some embodiments, the mTOR inhibitor nanoparticle composition is administered intravenously, intraarterially, intraperitoneally, intravesicularly, subcutaneously, intrathecally, intrapulmonarily, intramuscularly, intratracheally, intraocularly, transdermally, orally, or by inhalation. In some embodiments, the mTOR inhibitor nanoparticle composition is administered intravenously.

[0023] In some embodiments, the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab. In some embodiments, the anti-PD-1 antibody is nivolumab.

[0024] In some embodiments, the individual is human.

[0025] The disclosures of all publications, patents, patent applications and published patent applications referred to herein are hereby incorporated herein by reference in their entirety.

DETAILED DESCRIPTION OF THE INVENTION

[0026] The present application provides, in certain aspects, treatments for cancer, such as undifferentiated pleomorphic sarcoma (including undifferentiated pleomorphic sarcoma having a PTEN loss and/or a TSC2 mutation) or leiomyosarcoma (including estrogen receptor-positive leiomyosarcoma), comprising (a) a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin (e.g, nab- sirolimus); and optionally (b) a second therapeutic agent (e.g., an anti-PD-1 antibody (e.g, nivolumab)). In some embodiments, the method is for the treatment of undifferentiated pleomorphic sarcoma having a PTEN loss and a TSC2 mutation in an individual in need thereof. In some embodiments, the method is for the treatment of estrogen receptor-positive leiomyosarcoma in an individual in need thereof.

[0027] Certain subject matter the present application is based, at least in part, on the finding that individuals with undifferentiated pleomorphic sarcoma (including undifferentiated pleomorphic sarcoma having a PTEN loss and/or a TSC2 mutation) or leiomyosarcoma (including estrogen receptor-positive leiomyosarcoma) unexpectedly showed improved response to treatment with «a/>-sirolimus and an anti-PD-1 antibody, nivolumab, as compared to individuals receiving the combination treatment and having other cancer subtypes.

[0028] Thus, provided herein, in certain aspects, is a method of treating undifferentiated pleomorphic sarcoma having a PTEN loss and/ or a TSC2 mutation (e.g, a PTEN loss and a TSC2 mutation) in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin, e.g, /706-sirolimus: and optionally (b) a second therapeutic agent (e.g., an effective amount of an anti-PD-1 antibody, e.g, nivolumab).

[0029] In other aspects, provided herein is a method of treating estrogen receptor-positive leiomyosarcoma in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin, e.g, /706-sirolimus: and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody, e.g, nivolumab).

I. Definitions

[0030] As used herein "nab" stands for nanoparticle albumin-bound, and “«a6-sirolimus” is an albumin stabilized nanoparticle formulation of sirolimus. «a6-sirolimus is also known as nab- rapamycin, which has been previously described. See, for example, U.S. Patent Nos. 8,911,786 and 11,497,737, each of which is incorporated herein by reference in their entirety.

[0031] As used herein, “treatment” or “treating” is an approach for obtaining beneficial or desired results including clinical results. For purposes of this invention, beneficial or desired clinical results include, but are not limited to, one or more of the following: alleviating one or more symptoms resulting from the disease, diminishing the extent of the disease, stabilizing the disease (e.g, preventing or delaying the worsening of the disease), preventing or delaying the spread (e.g, metastasis) of the disease, preventing or delaying the recurrence of the disease, reducing recurrence rate of the disease, delay or slowing the progression of the disease, ameliorating the disease state, providing a remission (partial or total) of the disease, decreasing the dose of one or more other medications required to treat the disease, delaying the progression of the disease, increasing the quality of life, and/or prolonging survival. In some embodiments, the treatment reduces the severity of one or more symptoms associated with cancer by at least about any of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or 100% compared to the corresponding symptom in the same subject prior to treatment or compared to the corresponding symptom in other subjects not receiving the treatment. Also encompassed by "treatment" is a reduction of pathological consequence of cancer. The methods of the invention contemplate any one or more of these aspects of treatment. [0032] The terms “recurrence,” “relapse” or “relapsed” refers to the return of a cancer or disease after clinical assessment of the disappearance of disease. A diagnosis of distant metastasis or local recurrence can be considered a relapse.

[0033] The term “refractory” or “resistant” refers to a cancer or disease that has not responded to treatment.

[0034] As used herein, “delaying” the development of cancer means to defer, hinder, slow, retard, stabilize, and/or postpone development of the disease. This delay can be of varying lengths of time, depending on the history of the disease and/or individual being treated. As is evident to one skilled in the art, a sufficient or significant delay can, in effect, encompass prevention, in that the individual does not develop the disease. A method that “delays” development of cancer is a method that reduces probability of disease development in a given time frame and/or reduces the extent of the disease in a given time frame, when compared to not using the method. Such comparisons are typically based on clinical studies, using a statistically significant number of subjects. Cancer development can be detectable using standard methods, including, but not limited to, computerized axial tomography (CAT scan), Magnetic Resonance Imaging (MRI), ultrasound, clotting tests, arteriography, biopsy, urine cytology, and cystoscopy. Development may also refer to cancer progression that may be initially undetectable and includes occurrence, recurrence, and onset.

[0035] The term “effective amount” used herein refers to an amount of a compound or composition sufficient to treat a specified disorder, condition or disease such as ameliorate, palliate, lessen, and/or delay one or more of its symptoms. In reference to cancer, an effective amount comprises an amount sufficient to cause a tumor to shrink and/or to decrease the growth rate of the tumor (such as to suppress tumor growth) or to prevent or delay other unwanted cell proliferation in cancer. In some embodiments, an effective amount is an amount sufficient to delay development of cancer. In some embodiments, an effective amount is an amount sufficient to prevent or delay recurrence. In some embodiments, an effective amount is an amount sufficient to reduce recurrence rate in the individual. An effective amount can be administered in one or more administrations. The effective amount of the drug or composition may: (i) reduce the number of cancer cells; (ii) reduce tumor size; (iii) inhibit, retard, slow to some extent and preferably stop cancer cell infiltration into peripheral organs; (iv) inhibit (z.e., slow to some extent and preferably stop) tumor metastasis; (v) inhibit tumor growth; (vi) prevent or delay occurrence and/or recurrence of tumor; (vii) reduce recurrence rate of tumor, and/or (viii) relieve to some extent one or more of the symptoms associated with the cancer.

[0036] As is understood in the art, an “effective amount” may be in one or more doses, i.e., a single dose or multiple doses may be required to achieve the desired treatment endpoint. An effective amount may be considered in the context of administering one or more therapeutic agents, and a nanoparticle composition (e.g, a composition including sirolimus and an albumin) may be considered to be given in an effective amount if, in conjunction with one or more other agents, a desirable or beneficial result may be or is achieved. The components (e.g, the first and second therapies) in a combination therapy of the invention may be administered sequentially, simultaneously, or concurrently using the same or different routes of administration for each component. Thus, an effective amount of a combination therapy includes an amount of the first therapy and an amount of the second therapy that when administered sequentially, simultaneously, or concurrently produces a desired outcome.

[0037] ‘In conjunction with” or “in combination with” refers to administration of one treatment modality in addition to another treatment modality, such as administration of a nanoparticle composition described herein in addition to administration of the other agent to the same individual under the same treatment plan. As such, "in conjunction with" or “in combination with” refers to administration of one treatment modality before, during or after delivery of the other treatment modality to the individual.

[0038] The term “simultaneous administration,” as used herein, means that a first therapy and second therapy in a combination therapy are administered with a time separation of no more than about 15 minutes, such as no more than about any of 10, 5, or 1 minutes. When the first and second therapies are administered simultaneously, the first and second therapies may be contained in the same composition (e.g, a composition comprising both a first and second therapy) or in separate compositions (e.g., a first therapy is contained in one composition and a second therapy is contained in another composition).

[0039] As used herein, the term “sequential administration” means that the first therapy and second therapy in a combination therapy are administered with a time separation of more than about 15 minutes, such as more than about any of 20, 30, 40, 50, 60, or more minutes. Either the first therapy or the second therapy may be administered first. The first and second therapies are contained in separate compositions, which may be contained in the same or different packages or kits.

[0040] As used herein, the term “concurrent administration” means that the administration of the first therapy and that of a second therapy in a combination therapy overlap with each other.

[0041] The term “antibody” includes full-length antibodies and antigen-binding fragments thereof. A full-length antibody comprises two heavy chains and two light chains. The variable regions of the light and heavy chains are responsible for antigen binding. The variable region in both chains generally contain three highly variable loops called the complementarity determining regions (CDRs) (light chain (LC) CDRs including LC-CDR1, LC-CDR2, and LC- CDR3, heavy chain (HC) CDRs including HC-CDR1, HC-CDR2, and HC-CDR3). CDR boundaries for the antibodies and antigen-binding fragments disclosed herein may be defined or identified by the conventions of Kabat, Chothia, or Al-Lazikani (Al-Lazikani 1997; Chothia 1985; Chothia 1987; Chothia 1989; Kabat 1987; Kabat 1991). The three CDRs of the heavy or light chains are interposed between flanking stretches known as framework regions (FRs), which are more highly conserved than the CDRs and form a scaffold to support the hypervariable loops. The constant regions of the heavy and light chains are not involved in antigen binding, but exhibit various effector functions. Antibodies are assigned to classes based on the amino acid sequence of the constant region of their heavy chain. The five major classes or isotypes of antibodies are IgA, IgD, IgE, IgG, and IgM, which are characterized by the presence of a, 6, e, y, and p heavy chains, respectively. Several of the major antibody classes are divided into subclasses such as IgGl (y 1 heavy chain), lgG2 (y2 heavy chain), lgG3 (y3 heavy chain), lgG4 (y4 heavy chain), IgAl (al heavy chain), or lgA2 (a2 heavy chain).

[0042] The term “antigen-binding fragment” as used herein refers to an antibody fragment including, for example, a diabody, a Fab, a Fab', a F(ab')2, an Fv fragment, a disulfide stabilized Fv fragment (dsFv), a (dsFv)2, a bispecific dsFv (dsFv-dsFv¹), a disulfide stabilized diabody (ds diabody), a single-chain antibody molecule (scFv), an scFv dimer (bivalent diabody), a multispecific antibody formed from a portion of an antibody comprising one or more CDRs, a camelized single domain antibody, a nanobody, a domain antibody, a bivalent domain antibody, or any other antibody fragment that binds to an antigen but does not comprise a complete antibody structure. An antigen-binding fragment is capable of binding to the same antigen to which the parent antibody or a parent antibody fragment (e.g, a parent scFv) binds. In some embodiments, an antigen-binding fragment may comprise one or more CDRs from a particular human antibody grafted to a framework region from one or more different human antibodies.

[0043] As use herein, the term “specifically binds” or “is specific for” refers to measurable and reproducible interactions, such as binding between a target and an antibody or antibody moiety that is determinative of the presence of the target in the presence of a heterogeneous population of molecules, including biological molecules. For example, an antibody or antibody moiety that specifically binds to a target (which can be an epitope) is an antibody or antibody moiety that binds this target with greater affinity, avidity, more readily, and/or with greater duration than its bindings to other targets. In some embodiments, an antibody or antibody moiety that specifically binds to an antigen reacts with one or more antigenic determinants of the antigen (for example PD-1 or a portion thereol) with a binding affinity that is at least about 10 times its binding affinity for other targets.

[0044] As used herein, by “pharmaceutically acceptable” or “pharmacologically compatible” is meant a material that is not biologically or otherwise undesirable, e.g., the material may be incorporated into a pharmaceutical composition administered to a patient without causing any significant undesirable biological effects or interacting in a deleterious manner with any of the other components of the composition in which it is contained. Pharmaceutically acceptable carriers or excipients have preferably met the required standards of toxicological and manufacturing testing and/or are included on the Inactive Ingredient Guide prepared by the U. S. Food and Drug administration.

[0045] As used herein, the term “individual” refers to a mammal and includes, but is not limited to, human, bovine, horse, feline, canine, rodent, rat, mouse, dog, or primate. In some embodiments, the individual is a human individual.

[0046] The terms “comprising,” “having,” “containing,” and “including,” and other similar forms, and grammatical equivalents thereof, as used herein, are intended to be equivalent in meaning and to be open ended in that an item or items following any one of these words is not meant to be an exhaustive listing of such item or items, or meant to be limited to only the listed item or items. For example, an article “comprising” components A, B, and C can consist of (z.e., contain only) components A, B, and C, or can contain not only components A, B, and C but also one or more other components. As such, it is intended and understood that “comprises” and similar forms thereof, and grammatical equivalents thereof, include disclosure of embodiments of “consisting essentially of’ or “consisting of.”

[0047] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit, unless the context clearly dictate otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0048] Reference to “about” a value or parameter herein includes (and describes) variations that are directed to that value or parameter per se. For example, description referring to “about X” includes description of “X.”

[0049] As used herein, including in the appended claims, the singular forms “a,” “or,” and “the” include plural referents unless the context clearly dictates otherwise.

[0050] Those skilled in the art will recognize that several embodiments are possible within the scope and spirit of the present disclosure. The following description illustrates the disclosure and, of course, should not be construed in any way as limiting the scope of the inventions described herein.

II. Methods of treatment

[0051] Provided herein are methods for the treatment of a cancer, including undifferentiated pleomorphic sarcoma (including undifferentiated pleomorphic sarcoma having a PTEN loss and/or a TSC2 mutation) or leiomyosarcoma (including estrogen receptor-positive leiomyosarcoma), in an individual in need thereof, the treatments comprising administering to the individual (a) a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereol) and an albumin; and optionally (b) a second therapeutic agent (e.g., an anti-PD-1 antibody).

[0052] In some embodiments, provided is a method of treating undifferentiated pleomorphic sarcoma having a PTEN loss and/or a TSC2 mutation in an individual in need thereof, the treatments comprising administering to the individual (a) a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin (e.g, m6-sirolimus); and optionally (b) a second therapeutic agent (e.g., an anti-PD-1 antibody (e.g, nivolumab)). In some embodiments, the PTEN loss is a loss-of-function mutation or epigenetic silencing. In some embodiments, the individual is selected for the treatment on the basis of having the PTEN loss. In some embodiments, the TSC2 mutation is a missense mutation, nonsense mutation, deletion, splicing site mutation, insertion, substation, rearrangement, or frameshift, or a combination thereof. In some embodiments, the individual is selected for the treatment on the basis of the TSC2 mutation. In some embodiments, the method further comprises selecting the individual on the basis of having the PTEN loss and/ or the TSC2 mutation.

[0053] In some embodiments, provided is a method of treating estrogen receptor-positive leiomyosarcoma in an individual in need thereof, the treatments comprising administering to the individual (a) a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin (e.g, na6-sirolimus); and optionally (b) a second therapeutic agent (e.g., an anti-PD-1 antibody (e.g, nivolumab)).

[0054] In some embodiments, the cancer, such as the undifferentiated pleomorphic sarcoma or the leiomyosarcoma, is locally advanced, advanced, malignant, advanced malignant, or metastatic. In some embodiments, the cancer, such as the undifferentiated pleomorphic sarcoma or the leiomyosarcoma, is relapsed, refractory, or resistant to a prior treatment. In some embodiments, the prior treatment comprises a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin, e.g, m/j-sirolimus. In some embodiments, prior treatment comprises a non-nanoparticle formulation of an mTOR inhibitor, such as sirolimus.

[0055] The methods provided herein are applicable to all stages of cancer, such as the undifferentiated pleomorphic sarcoma or leiomyosarcoma, including stages, I, II, III, and IV, according to the American Joint Committee on Cancer (AJCC) staging groups. In some embodiments, the cancer, such as the undifferentiated pleomorphic sarcoma or leiomyosarcoma, is an early stage cancer, non-metastatic cancer, primary cancer, advanced cancer, locally advanced cancer, metastatic cancer, cancer in remission, cancer in an adjuvant setting, or cancer in a neoadjuvant setting. In some embodiments, the solid tumor is localized resectable, localized unresectable, or unresectable. In some embodiments, the cancer, such as the undifferentiated pleomorphic sarcoma or leiomyosarcoma, is localized resectable or borderline resectable. A. Dosing and Methods of Administration

[0056] The dose of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) administered to an individual (e.g, a human) may vary with the particular composition, the method of administration, and the particular stage of tumor being treated. The amount should be sufficient to produce a desirable response, such as a therapeutic or prophylactic response against the tumor. In some embodiments, the amount of mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in the composition is below the level that induces a toxicological effect (e.g, an effect above a clinically acceptable level of toxicity) or is at a level where a potential side effect can be controlled or tolerated when the mTOR inhibitor nanoparticle composition is administered to the individual.

[0057] In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered to the individual simultaneously with the second therapeutic agent such as an anti-PD-1 antibody. For example, the mTOR inhibitor nanoparticle compositions and the anti-PD-1 antibody are administered with a time separation of no more than about 15 minutes, such as no more than about any of 10, 5, or 1 minutes. In one example, wherein the compounds are in solution, simultaneous administration can be achieved by administering a solution containing the combination of compounds. In another example, simultaneous administration of separate solutions, one of which contains the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the other of which contains the anti-PD-1 antibody, can be employed. In one example, simultaneous administration can be achieved by administering a composition containing the combination of compounds. In another example, simultaneous administration can be achieved by administering two separate compositions, one comprising the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the other comprising the anti-PD-1 antibody. In some embodiments, simultaneous administration of the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in the nanoparticle composition and the anti-PD-1 antibody can be combined with supplemental doses of the mTOR inhibitor and/or the anti-PD-1 antibody.

[0058] In other embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent such as anti-PD-1 antibody are not administered simultaneously. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered before the anti-PD-1 antibody. In other embodiments, the anti-PD-1 antibody is administered before the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition). The time difference in non-simultaneous administrations can be greater than 1 minute, five minutes, 10 minutes, 15 minutes, 30 minutes, 45 minutes, 60 minutes, two hours, three hours, six hours, nine hours, 12 hours, 24 hours, 36 hours, or 48 hours. In other embodiments, the first administered compound is provided time to take effect on the patient before the second administered compound is administered. In some embodiments, the difference in time does not extend beyond the time for the first administered compound to complete its effect in the patient, or beyond the time the first administered compound is completely or substantially eliminated or deactivated in the patient.

[0059] In some embodiments, the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent such as anti-PD-1 antibody are concurrent, /.£., the administration period of the mTOR inhibitor nanoparticle composition and that of the anti-PD-1 antibody overlap with each other. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered for at least one cycle (for example, at least any of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 15, 16, 17, 18, 19, or 20 cycles) prior to the administration of the anti-PD-1 antibody. In some embodiments, the anti-PD-1 antibody is administered for at least any of one, two, three, or four weeks. In some embodiments, the administrations of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody are initiated at about the same time (for example, within any one of 1, 2, 3, 4, 5, 6, or 7 days). In some embodiments, the administrations of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD- 1 antibody are terminated at about the same time (for example, within any one of 1, 2, 3, 4, 5, 6, or 7 days). In some embodiments, the administration of the anti-PD-1 antibody continues (for example for about any one of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months) after the termination of the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition). In some embodiments, the administration of the anti-PD-1 antibody is initiated after (for example after about any one of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months) the initiation of the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition). In some embodiments, the administrations of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody are initiated and terminated at about the same time. In some embodiments, the administrations of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody are initiated at about the same time and the administration of the anti-PD-1 antibody continues (for example for about any one of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months) after the termination of the administration of the mTOR inhibitor nanoparticle composition. In some embodiments, the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody stop at about the same time and the administration of the anti-PD-1 antibody is initiated after (for example after about any one of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months) the initiation of the administration of the mTOR inhibitor nanoparticle composition.

[0060] In some embodiments, the second therapeutic agent such as anti-PD-1 antibody is administered on day 1 of a 21 -day cycle, and the mTOR inhibitor nanoparticle composition is administered on days 8 and 15 of the 21 -day cycle. In some embodiments, the first cycle only includes administration of the anti-PD-1 antibody, e.g, on day 1. In some embodiments, the administration of the composition comprising an mTOR inhibitor and an albumin and an anti- PD-1 antibody continues for at least any of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 15, 16, 17, 18, 19, or 20 cycles.

[0061] In some embodiments, the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent such as anti-PD-1 antibody are non-concurrent. For example, in some embodiments, the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is terminated before the anti-PD-1 antibody is administered. In some embodiments, the administration of the anti-PD-1 antibody is terminated before the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered. The time period between these two non-concurrent administrations can range from about two to eight weeks, such as about four weeks.

[0062] The dosing frequency of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent such as anti-PD-1 antibody may be adjusted over the course of the treatment, based on the judgment of the administering physician. When administered separately, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent such as anti-PD-1 antibody can be administered at different dosing frequency or intervals. For example, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) can be administered weekly, while the anti-PD-1 antibody can be administered more or less frequently. In some embodiments, a sustained continuous release formulation of the nanoparticle and/or anti-PD-1 antibody may be used. Various formulations and devices for achieving sustained release are known in the art. A combination of the administration configurations described herein can also be used.

[0063] The mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent such as anti-PD-1 antibody can be administered using the same route of administration or different routes of administration. In some embodiments (for both simultaneous and sequential administrations), the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereol) in the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered at a predetermined ratio.

[0064] The doses required for the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereol) in the mTOR inhibitor nanoparticle composition and/or the second therapeutic agent such as anti-PD-1 antibody may (but not necessarily) be the same or lower than what is normally required when each agent is administered alone. Thus, in some embodiments, a subtherapeutic amount of the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in the mTOR inhibitor nanoparticle composition and/or the anti-PD-1 antibody is administered. “Subtherapeutic amount” or “subtherapeutic level” refer to an amount that is less than the therapeutic amount, that is, less than the amount normally used when the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and/or the anti-PD-1 antibody are administered alone. The reduction may be reflected in terms of the amount administered at a given administration and/or the amount administered over a given period of time (reduced frequency).

[0065] In some embodiments, enough second therapeutic agent (e.g., anti-PD-1 antibody) is administered so as to allow reduction of the normal dose of the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in the mTOR inhibitor nanoparticle composition required to affect the same degree of treatment by at least about any of 5%, 10%, 20%, 30%, 50%, 60%, 70%, 80%, 90%, or more. In some embodiments, enough mTOR inhibitor (such as a limus drug, e.g., sirolimus or a derivative thereof) in the mTOR inhibitor nanoparticle composition is administered so as to allow reduction of the normal dose of the second therapeutic agent (e.g. anti-PD-1 antibody) required to affect the same degree of treatment by at least about any of 5%, 10%, 20%, 30%, 50%, 60%, 70%, 80%, 90%, or more.

[0066] In some embodiments, the dose of both the mTOR inhibitor (such as a limus drug, e.g., sirolimus or a derivative thereof) in the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are reduced as compared to the corresponding normal dose of each when administered alone. In some embodiments, both the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in the mTOR inhibitor nanoparticle composition and the anti- PD-1 antibody are administered at a subtherapeutic, i.e., reduced, level. In some embodiments, the dose of the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in the mTOR inhibitor nanoparticle composition and/or the anti-PD-1 antibody is substantially less than the established maximum toxic dose (MTD). For example, the dose of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and/or the anti- PD-1 antibody is less than about 50%, 40%, 30%, 20%, or 10% of the MTD.

[0067] A combination of the administration configurations described herein can be used. The combination therapy methods described herein may be performed alone or in conjunction with another therapy, such as surgery, radiation, gene therapy, immunotherapy, bone marrow transplantation, stem cell transplantation, hormone therapy, targeted therapy, cryotherapy, ultrasound therapy, photodynamic therapy, and/or chemotherapy and the like. Additionally, a person having a greater risk of developing the solid tumor may receive treatments to inhibit and/or delay the development of the disease.

[0068] As will be understood by those of ordinary skill in the art, in some embodiments, the appropriate doses of second agents will be approximately those already employed in clinical therapies wherein the anti-PD-1 antibody is administered alone or in combination with other chemotherapeutic agents. Variation in dosage will likely occur depending on the condition being treated. As described above, in some embodiments, the second chemotherapeutic agent may be administered at a reduced level.

[0069] In some embodiments, the amounts of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody are below the levels that induce a toxicological effect (i.e., an effect above a clinically acceptable level of toxicity) or are at a level where a potential side effect can be controlled or tolerated when the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered to the individual.

[0070] In some embodiments, the amount of the mTOR inhibitor nanoparticle composition

(such as sirolimus/albumin nanoparticle composition) is close to a maximum tolerated dose (MTD) of the composition following the same dosing regimen when administered with the anti- PD-1 antibody. In some embodiments, the amount of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is more than about any of 80%, 90%, 95%, or 98% of the MTD when administered with the anti-PD-1 antibody.

[0071] In some embodiments, the amount of an mTOR inhibitor (such as a limus drug, e.g, sirolimus) in the mTOR inhibitor nanoparticle composition is about any of 25 mg/m², 30 mg/m², 45 mg/m², 50 mg/m², 56 mg/m², 60 mg/m², 75 mg/m², 80 mg/m², 90 mg/m², 100 mg/m², 120 mg/m², 160 mg/m², 175 mg/m², 180 mg/m², 200 mg/m², 210 mg/m², 220 mg/m², 250 mg/m², 260 mg/m², 300 mg/m², 350 mg/m², 400 mg/m², 500 mg/m², 540 mg/m², 750 mg/m², 1000 mg/m², or 1080 mg/m² mTOR inhibitor. In some embodiments, the mTOR inhibitor nanoparticle composition includes less than about any of 350 mg/m², 300 mg/m², 250 mg/m², 200 mg/m², 150 mg/m², 120 mg/m², 100 mg/m², 90 mg/m², 50 mg/m², or 30 mg/m² mTOR inhibitor (such as a limus drug, e.g, sirolimus). In some embodiments, the amount of the mTOR inhibitor (such as a limus drug, e.g, sirolimus) per administration is less than about any of 25 mg/m², 22 mg/m², 20 mg/m², 18 mg/m², 15 mg/m², 14 mg/m², 13 mg/m², 12 mg/m², 11 mg/m², 10 mg/m², 9 mg/m², 8 mg/m², 7 mg/m², 6 mg/m², 5 mg/m², 4 mg/m², 3 mg/m², 2 mg/m², or 1 mg/m². In some embodiments, the effective amount of mTOR inhibitor (such as a limus drug, e.g, sirolimus) in the mTOR inhibitor nanoparticle composition is included in any of the following ranges: about 1 to about 5 mg/m², about 5 to aboutlO mg/m², about 10 to about 25 mg/m², about 25 to about 50 mg/m², about 50 to about 75 mg/m², about 75 to about 100 mg/m², about 100 to about 125 mg/m², about 125 to aboutl50 mg/m², aboutl50 to about 175 mg/m², aboutl75 to about 200 mg/m², about 200 to about 225 mg/m², about 225 to about 250 mg/m², about 250 to about 300 mg/m², about 300 to about 350 mg/m², or about 350 to about 400 mg/m². In some embodiments, the effective amount of mTOR inhibitor (such as a limus drug, e.g, sirolimus) in the mTOR inhibitor nanoparticle composition is about 30 to about 300 mg/m², such as about 100 to about 150 mg/m², about 120 mg/m2, about 130 mg/m², or aboutl40 mg/m². In some embodiments, the amount of the mTOR inhibitor nanoparticle composition is administered weekly. In some embodiments, the amount of the mTOR inhibitor nanoparticle composition is administered weekly every 2 out of 3 weeks. In some embodiments, the amount of the mTOR inhibitor nanoparticle composition is on days 8 and 15 of a 21-day cycle, days 1 or 8 of a 21-day cycle, days 15 and 21 or a 21-day cycle, days 1 and 15 of a 21-day cycle, or days 1 and 21 of a 21-day cycle.

[0072] In some embodiments, the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab. In some embodiments, the amount of an anti-PD-1 antibody (such as nivolumab) is about 1 mg/kg to about 10 mg/kg. In some embodiments, the amount of an anti-PD-1 antibody (such as nivolumab) is at least about 1 mg/kg, such as at least about any of 2 mg/kg, 3 mg/kg, 4 mg/kg, 5 mg/kg, 6 mg/kg, 7 mg/kg, 8 mg/kg, 9 mg/kg, or 10 mg/kg. In some embodiments, the amount of an anti-PD-1 antibody (such as nivolumab) 1 mg/kg or less, such as any of 2 mg/kg or less, 3 mg/kg or less, 4 mg/kg or less, 5 mg/kg or less, 6 mg/kg or less, 7 mg/kg or less, 8 mg/kg or less, 9 mg/kg or less, or 10 mg/kg or less. In some embodiments, the amount of an anti-PD-1 antibody (such as nivolumab) is about any of 1 mg/kg, 2 mg/kg, 3 mg/kg, 4 mg/kg, 5 mg/kg, 6 mg/kg, 7 mg/kg, 8 mg/kg, 9 mg/kg, or 10 mg/kg. In some embodiments, the amount of an anti- PD-1 antibody is administered weekly, every two weeks, every three weeks, or monthly. In some embodiments, the amount of an anti-PD-1 antibody is administered every three weeks. In some embodiments, the amount of an anti-PD-1 antibody is administered every 21 days.

[0073] In some embodiments, the dosing frequencies for the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) include, but are not limited to, daily, every two days, every three days, every four days, every five days, every six days, weekly without break, three out of four weeks (such as on days 1, 8, and 15 of a 28-day cycle), once every three weeks, once every two weeks, or two out of three weeks. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered about once every 2 weeks, once every 3 weeks, once every 4 weeks, once every 6 weeks, or once every 8 weeks. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered at least about any of lx, 2x, 3x, 4x, 5x, 6x, or 7x (z.e., daily) a week. In some embodiments, the intervals between each administration are less than about any of 6 months, 3 months, 1 month, 20 days, 15, days, 14 days, 13 days, 12 days, 11 days, 10 days, 9 days, 8 days, 7 days, 6 days, 5 days, 4 days, 3 days, 2 days, or 1 day. In some embodiments, the intervals between each administration are more than about any of 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 8 months, or 12 months. In some embodiments, there is no break in the dosing schedule. In some embodiments, the interval between each administration is no more than about a week.

[0074] In some embodiments, the dosing frequency is once every two days for one time, two times, three times, four times, five times, six times, seven times, eight times, nine times, ten times, or eleven times. In some embodiments, the dosing frequency is once every two days for five times. In some embodiments, the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) is administered over a period of at least ten days, wherein the interval between each administration is no more than about two days, and wherein the dose of the mTOR inhibitor at each administration is about 0.25 mg/m² to about 250 mg/m², about 0.25 mg/m² to about 150 mg/m², about 0.25 mg/m² to about 75 mg/m², such as about 0.25 mg/m² to about 25 mg/m², or about 25 mg/m² to about 50 mg/m².

[0075] The administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/ albumin nanoparticle composition) can be extended over an extended period of time, such as from about a month up to about seven years. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered over a period of at least about any of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 18, 24, 30, 36, 48, 60, 72, or 84 months.

[0076] In some embodiments, the dosage of an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) in a nanoparticle composition can be in the range of 5-400 mg/m² when given on a 3-week schedule, or 5-250 mg/m²(such as 80-150 mg/m², for example 100-120 mg/m²) when given on a weekly schedule. For example, the amount of an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) is about 60 to about 300 mg/m² (e.g, about 260 mg/m²) on a 3-week schedule.

[0077] In some embodiments, the exemplary dosing schedules for the administration of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) include, but are not limited to, 100 mg/m², weekly, without break; 10 mg/m² weekly, 3 out of four weeks (such as on days 1, 8, and 15 of a 28-day cycle); 45 mg/m² weekly, 3 out of four weeks (such as on days 1, 8, and 15 of a 28-day cycle); 75 mg/m² weekly, 3 out of four weeks (such as on days 1, 8, and 15 of a 28-day cycle); 100 mg/m², weekly, 3 out of 4 weeks; 125 mg/m², weekly, 3 out of 4 weeks; 125 mg/m², weekly, 2 out of 3 weeks; 130 mg/m², weekly, without break; 175 mg/m², once every 2 weeks; 260 mg/m², once every 2 weeks; 260 mg/m², once every 3 weeks; 180-300 mg/m², every three weeks; 60-175 mg/m², weekly, without break; 20-150 mg/m² twice a week; and 150-250 mg/m² twice a week. The dosing frequency of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) may be adjusted over the course of the treatment based on the judgment of the administering physician.

[0078] In some embodiments, the individual is treated for at least about any of one, two, three, four, five, six, seven, eight, nine, or ten treatment cycles.

[0079] The mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) described herein allow infusion of the mTOR inhibitor nanoparticle composition to an individual over an infusion time that is shorter than about 24 hours. For example, in some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered over an infusion period of less than about any of 24 hours, 12 hours, 8 hours, 5 hours, 3 hours, 2 hours, 1 hour, 30 minutes, 20 minutes, or 10 minutes. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered over an infusion period of about 30 minutes.

[0080] In some embodiments, the exemplary dose of the mTOR inhibitor (in some embodiments a limus drug, e.g, sirolimus) in the mTOR inhibitor nanoparticle composition includes, but is not limited to, about any of 50 mg/m², 60 mg/m², 75 mg/m², 80 mg/m², 90 mg/m², 100 mg/m², 120 mg/m², 160 mg/m², 175 mg/m², 200 mg/m², 210 mg/m², 220 mg/m², 260 mg/m², and 300 mg/m². For example, the dosage of an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereol) in a nanoparticle composition can be in the range of about 100-400 mg/m² when given on a 3-week schedule, or about 10-250 mg/m² when given on a weekly schedule.

[0081] In some embodiments, the dosage of an mTOR inhibitor (such as a limus drug, e.g, sirolimus) is about 100 mg to about 400 mg, for example about 100 mg, about 200 mg, about 300 mg, or about 400 mg. In some embodiments, the limus drug is administered at about 100 mg weekly, about 200 mg weekly, about 300 mg weekly, about 100 mg twice weekly, or about 200 mg twice weekly. In some embodiments, the administration is further followed by a monthly maintenance dose (which can be the same or different from the weekly doses).

[0082] In some embodiments when the limus nanoparticle composition is administered intravenously, the dosage of an mTOR inhibitor (such as a limus drug, e.g, sirolimus) in a nanoparticle composition can be in the range of about 30 mg to about 400 mg. The mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) described herein allow infusion of the mTOR inhibitor nanoparticle composition to an individual over an infusion time that is shorter than about 24 hours. For example, in some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered over an infusion period of less than about any of 24 hours, 12 hours, 8 hours, 5 hours, 3 hours, 2 hours, 1 hour, 30 minutes, 20 minutes, or 10 minutes. In some embodiments, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) is administered over an infusion period of about 30 minutes to about 40 minutes.

[0083] The anti-PD-1 antibody (such as nivolumab) described herein can be infused to an individual over an infusion time that is shorter than about 24 hours. For example, in some embodiments, the anti-PD-1 antibody (such as nivolumab) is administered over an infusion period of less than about any of 24 hours, 12 hours, 8 hours, 5 hours, 3 hours, 2 hours, 1 hour, 30 minutes, 20 minutes, or 10 minutes. In some embodiments, the anti-PD-1 antibody (such as nivolumab) is administered over an infusion period of about 30 minutes.

[0084] In some embodiments, each dosage contains both an mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and an anti-PD-1 antibody to be delivered as a single dosage, while in other embodiments, each dosage contains either the mTOR inhibitor nanoparticle composition or the anti-PD-1 antibody to be delivered as separate dosages.

[0085] An mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and an anti-PD-1 antibody, in pure form or in an appropriate pharmaceutical composition, can be administered via any of the accepted modes of administration or agents known in the art. The compositions and/or agents can be administered, for example, orally, nasally, parenterally (such as intravenous, intramuscular, or subcutaneous), topically, transdermally, intravaginally, intravesically, intracistemally, or rectally. The dosage form can be, for example, a solid, semi-solid, lyophilized powder, or liquid dosage form, such as tablets, pills, soft elastic or hard gelatin capsules, powders, solutions, suspensions, suppositories, aerosols, or the like, preferably in unit dosage forms suitable for simple administration of precise dosages.

[0086] As discussed above, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the second therapeutic agent (e.g., anti-PD-1 antibody) can be administered in a single unit dose or separate dosage forms. Accordingly, the phrase “pharmaceutical combination” includes a combination of two drugs in either a single dosage form or a separate dosage forms, i.e., the pharmaceutically acceptable carriers and excipients described throughout the application can be combined with an mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and a second therapeutic agent (e.g., anti-PD-1 antibody) in a single unit dose, as well as individually combined with an mTOR inhibitor nanoparticle composition and a second therapeutic agent (e.g., an anti-PD-1 antibody) when these compounds are administered separately.

[0087] Auxiliary and adjuvant agents may include, for example, preserving, wetting, suspending, sweetening, flavoring, perfuming, emulsifying, and dispensing agents. Prevention of the action of microorganisms is generally provided by various antibacterial and antifungal agents, such as, parabens, chlorobutanol, phenol, sorbic acid, and the like. Isotonic agents, such as sugars, sodium chloride, and the like, may also be included. Prolonged absorption of an injectable pharmaceutical form can be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin. The auxiliary agents also can include wetting agents, emulsifying agents, pH buffering agents, and antioxidants, such as citric acid, sorbitan monolaurate, triethanolamine oleate, butylated hydroxy toluene, and the like.

[0088] Solid dosage forms can be prepared with coatings and shells, such as enteric coatings and others well-known in the art. They can contain pacifying agents and can be of such composition that they release the active compound or compounds in a certain part of the intestinal tract in a delayed manner. Examples of embedded compositions that can be used are polymeric substances and waxes. The active compounds also can be in microencapsulated form, if appropriate, with one or more of the above-mentioned excipients.

[0089] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs. Such dosage forms are prepared, for example, by dissolving, or dispersing, the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) or anti-PD-1 antibody described herein, or a pharmaceutically acceptable salt thereof, and optional pharmaceutical adjuvants in a carrier, such as, for example, water, saline, aqueous dextrose, glycerol, ethanol and the like; solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propyleneglycol, 1,3-butyleneglycol, dimethyl formamide; oils, in particular, cottonseed oil, groundnut oil, com germ oil, olive oil, castor oil and sesame oil, glycerol, tetrahydrofurfuryl alcohol, polyethyleneglycols and fatty acid esters of sorbitan; or mixtures of these substances, and the like, to thereby form a solution or suspension.

[0090] In some embodiments, depending on the intended mode of administration, the pharmaceutically acceptable compositions will contain about 1% to about 99% by weight of the compounds described herein, or a pharmaceutically acceptable salt thereof, and 99% to 1% by weight of a pharmaceutically acceptable excipient. In one example, the composition will be between about 5% and about 75% by weight of a compound described herein, or a pharmaceutically acceptable salt thereof, with the rest being suitable pharmaceutical excipients.

[0091] Actual methods of preparing such dosage forms are known, or will be apparent, to those skilled in this art. Reference is made, for example, to Remington's Pharmaceutical Sciences, 18th Ed., (Mack Publishing Company, Easton, Pa., 1990).

[0092] The mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) can be administered to an individual (such as a human) via various routes, including, for example, intravenous, intra-arterial, intraperitoneal, intrapulmonary, oral, inhalation, intravesicular, intramuscular, intra-tracheal, subcutaneous, intraocular, intrathecal, transmucosal, and transdermal. In some embodiments, sustained continuous release formulation of the composition may be used. In some embodiments, the composition is administered intravenously. In some embodiments, the composition is administered intraportally. In some embodiments, the composition is administered intraarterially. In some embodiments, the composition is administered intraperitoneally.

[0093] The anti-PD-1 antibody (such nivolumab) can be administered to an individual (such as a human) via various routes, including, for example, intravenous, intra-arterial, intraperitoneal, intrapulmonary, oral, inhalation, intravesicular, intramuscular, intra-tracheal, subcutaneous, intraocular, intrathecal, transmucosal, and transdermal. In some embodiments, the anti-PD-1 antibody is administered intravenously.

B. Treatment of undifferentiated pleomorphic sarcoma

[0094] In some embodiments, provided is a method of treating undifferentiated pleomorphic sarcoma in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody).

[0095] Undifferentiated pleomorphic sarcoma (UPS) is a high-grade, and often aggressive, soft-tissue sarcoma. Undifferentiated pleomorphic sarcoma usually appear as asymptomatic, unremarkable, rapidly growing cutaneous or subcutaneous nodule without superficial skin abnormalities. It is likely that the origin of undifferentiated pleomorphic sarcoma is mesenchymal stem cells. Undifferentiated pleomorphic sarcoma has been found to affect, amongst other areas, bones, soft tissues, and the retroperitoneum, and can also metastasize to other organs. Undifferentiated pleomorphic sarcoma can be diagnosed via histopathology of tumor samples, such as from a core needle technique or biopsy. Markers can be used to diagnose undifferentiated pleomorphic sarcoma, and include keratins, S100 protein, and/or SOXIO, smooth muscle actin (SMA), and desmin. MDM2 and CDK4 may also be helpful to distinguish undifferentiated pleomorphic sarcoma from dedifferentiated liposarcoma. Undifferentiated pleomorphic sarcoma exhibits atypical, pleomorphic spindle cells with abundant mitotic figures, and the tumor may display storiform, fascicular, or sheet-like configuration within a fibrous stroma. Robles-Tenorio & Solis-Ledesma, Slat Pear Is. Undifferentiated Pleomorphic Sarcoma, 2022.

[0096] In some embodiments, the undifferentiated pleomorphic sarcoma has phosphatase and tensin homolog (PTEN) loss. In some embodiments, the PTEN loss is a loss-of-function mutation or epigenetic silencing. PTEN loss is described in, e.g., Chang et al. , Biomolecules , 9, 2019; Vidotto et al., BrJ Cancer, 122, 2020, the contents of each of which are incorporated herein by reference in their entirety.

[0097] In some embodiments, the undifferentiated pleomorphic sarcoma has a tuberous sclerosis complex 2 (TSC2) mutation. In some embodiments, the TSC2 mutation is a missense mutation, nonsense mutation, deletion, splicing site mutation, insertion, substation, rearrangement, or frameshift, or a combination thereof. In some embodiments, the TSC2 mutation is an mTOR-activating aberration of TSC2. In some embodiments, the TSC2 mutation comprises a single-nucleotide variant (SNV). In some embodiments, the SNV comprises a mutation selected from the group consisting of C1503T, C2743G, C5383T, C3755G, G760T, C3442T, G880A, T707C, A4949G, or a deletion of any one or more of the amino acids at the position of 1405-1409, 1960-1970, 4999, 5002, 3521, 5208, 5238-5255. In some embodiments, the TSC2 mutation is a copy number variation of TSC2. In some embodiments, the the TSC2 mutation is a loss of function mutation. In some embodiments, the TSC2 mutation results in an aberrant expression level of TSC2. In some embodiments, the TSC2 mutation results in an aberrant activity level of a protein encoded by TSC2. In some embodiments, the TSC2 mutation results in a loss of heterozygosity of TSC2. Mutation analysis of TSC2 genes is known, e.g, Avgeris et al., Sci Rep, 7, 2017, the contents of which are incorporated herein by reference in their entirety.

[0098] In some embodiments, the individual is selected for the treatment on the basis of having a PTEN loss. In some embodiments, the individual is selected for the treatment on the basis of a TSC2 mutation. In some embodiments, the individual is selected for the treatment on the basis of having a PTEN loss and a TSC2 mutation. In some embodiments, the method further comprises selecting the individual on the basis of having a PTEN loss. In some embodiments, the method further comprises selecting the individual on the basis of having a TSC2 mutation. In some embodiments, the method further comprises selecting the individual on the basis of having a PTEN loss and a TSC2 mutation.

[0099] In some embodiments, there is provided a method of treating undifferentiated pleomorphic sarcoma having a PTEN loss in an individual (such as a human) comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g., sirolimus or a derivative thereof) and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody). In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g., sirolimus or a derivative thereof) and an albumin, wherein the mTOR inhibitor in the nanoparticles is associated (e.g, coated) with the albumin; and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with the albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm, for example about 100 nm), wherein the weight ratio of albumin and the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 9: 1 or less (such as about 9:1 or about 8:1); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the mTOR inhibitor is a limus drug. In some embodiments, the mTOR inhibitor is sirolimus or a derivative thereof. In some embodiments, the mTOR inhibitor nanoparticle composition comprises «a6-sirolimus. In some embodiments, the mTOR inhibitor nanoparticle composition is «a6-sirolimus. In some embodiments, the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab. In some embodiments, the anti-PD-1 antibody is nivolumab. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered sequentially. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered simultaneously. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered concurrently. In some embodiments, the mTOR inhibitor nanoparticle composition is administered 2 out of every 3 weeks, such as on days 8 and 15 of a 21 -day cycle. In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is from about 10 mg/m² to about 150 mg/m², such as about 100 mg/m². In some embodiments, the anti-PD-1 antibody is administered every three weeks, such as on day 1 of a 21-day cycle. In some embodiments, the anti-PD-1 antibody is administered for at least one cycle prior to administration of the mTOR inhibitor nanoparticle composition. In some embodiments, the anti-PD-1 antibody is administered at an amount of about 1 mg/kg to about 10 mg/kg, such as about 3 mg/kg.

[0100] In some embodiments, there is provided a method of treating undifferentiated pleomorphic sarcoma having a TSC2 mutation in an individual (such as a human) comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody). In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the mTOR inhibitor in the nanoparticles is associated (e.g, coated) with the albumin; and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with the albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm, for example about 100 nm), wherein the weight ratio of albumin and the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 9: 1 or less (such as about 9:1 or about 8:1); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the mTOR inhibitor is a limus drug. In some embodiments, the mTOR inhibitor is sirolimus or a derivative thereof. In some embodiments, the mTOR inhibitor nanoparticle composition comprises «a6-sirolimus. In some embodiments, the mTOR inhibitor nanoparticle composition is «a6-sirolimus. In some embodiments, the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab. In some embodiments, the anti-PD-1 antibody is nivolumab. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered sequentially. In some embodiments, th the nanoparticle composition and anti-PD-1 antibody are administered simultaneously. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered concurrently. In some embodiments, the mTOR inhibitor nanoparticle composition is administered 2 out of every 3 weeks, such as on days 8 and 15 of a 21 -day cycle. In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is from about 10 mg/m² to about 150 mg/m², such as about 100 mg/m². In some embodiments, the anti-PD-1 antibody is administered every three weeks, such as on day 1 of a 21-day cycle. In some embodiments, the anti-PD-1 antibody is administered for at least one cycle prior to administration of the mTOR inhibitor nanoparticle composition. In some embodiments, the anti-PD-1 antibody is administered at an amount of about 1 mg/kg to about 10 mg/kg, such as about 3 mg/kg.

[0101] In some embodiments, there is provided a method of treating undifferentiated pleomorphic sarcoma having a PTEN loss and a TSC2 mutation in an individual (such as a human) comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody). In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the mTOR inhibitor in the nanoparticles is associated (e.g, coated) with the albumin; and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with the albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm, for example about 100 nm), wherein the weight ratio of albumin and the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 9: 1 or less (such as about 9:1 or about 8:1); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the mTOR inhibitor is a limus drug. In some embodiments, the mTOR inhibitor is sirolimus or a derivative thereof. In some embodiments, the mTOR inhibitor nanoparticle composition comprises na6-sirolimus. In some embodiments, the mTOR inhibitor nanoparticle composition is na6-sirolimus. In some embodiments, the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab. In some embodiments, the anti-PD-1 antibody is nivolumab. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered sequentially. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered simultaneously. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered concurrently. In some embodiments, the mTOR inhibitor nanoparticle composition is administered 2 out of every 3 weeks, such as on days 8 and 15 of a 21 -day cycle. In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is from about 10 mg/m² to about 150 mg/m², such as about 100 mg/m². In some embodiments, the anti-PD-1 antibody is administered every three weeks, such as on day 1 of a 21-day cycle. In some embodiments, the anti-PD-1 antibody is administered for at least one cycle prior to administration of the mTOR inhibitor nanoparticle composition. In some embodiments, the anti-PD-1 antibody is administered at an amount of about 1 mg/kg to about 10 mg/kg, such as about 3 mg/kg. C. Treatment of leiomyosarcoma

[0102] In some embodiments, provided herein is a method of treating leiomyosarcoma in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g. an anti- PD-1 antibody).

[0103] Leiomyosarcoma is a subtype of soft tissue sarcoma, presented in most parts of the human body, with common locations including the abdomen, retroperitoneum, larger blood vessels, and the uterus. Leiomyosarcoma is a malignant mesenchymal tumor composed of cells that show distinct features of the smooth muscle lineage, and can be categorized as somatic soft tissue leiomyosarcoma, cutaneous leiomyosarcoma, or vascular leiomyosarcoma. Diagnosis can be performed on a tumor sample, such as obtained from a core needle biopsy. Leiomyosarcoma exhibit areas of high cellularity, commonly arranged in fascicles, and malignant cells are characterized by abundant pink to deep red cytoplasm on hematoxylin and eosin (H&E) staining, with cigar-shaped, centrally located nuclei. Such distinguishing features are lost in dedifferentiated tumors. EL-Naggar et al., Cancer Genomics, Chapter 22, 2014.

[0104] In some embodiments, the leiomyosarcoma is estrogen receptor-positive leiomyosarcoma.

[0105] In some embodiments, the individual is selected for the treatment on the basis of having the estrogen receptor-positive leiomyosarcoma. In some embodiments, the method further comprises selecting the individual on the basis of having the estrogen receptor-positive leiomyosarcoma.

[0106] In some embodiments, there is provided a method of treating estrogen receptorposition leiomyosarcoma in an individual (such as a human) comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g., sirolimus or a derivative thereol) and an albumin; and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereol) and an albumin, wherein the mTOR inhibitor in the nanoparticles is associated (e.g, coated) with the albumin; and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the method comprises administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) and an albumin, wherein the nanoparticles comprise the mTOR inhibitor associated (e.g, coated) with the albumin, wherein the nanoparticles have an average particle size of no greater than about 150 nm (such as no greater than about 120 nm, for example about 100 nm), wherein the weight ratio of albumin and the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 9: 1 or less (such as about 9:1 or about 8:1); and (b) an effective amount of an anti-PD-1 antibody. In some embodiments, the mTOR inhibitor is a limus drug. In some embodiments, the mTOR inhibitor is sirolimus or a derivative thereof. In some embodiments, the mTOR inhibitor nanoparticle composition comprises «a6-sirolimus. In some embodiments, the mTOR inhibitor nanoparticle composition is «a6-sirolimus. In some embodiments, the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab. In some embodiments, the anti-PD-1 antibody is nivolumab. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered sequentially. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered simultaneously. In some embodiments, the nanoparticle composition and anti-PD-1 antibody are administered concurrently. In some embodiments, the mTOR inhibitor nanoparticle composition is administered 2 out of every 3 weeks, such as on days 8 and 15 of a 21 -day cycle. In some embodiments, the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is from about 10 mg/m² to about 150 mg/m², such as about 100 mg/m². In some embodiments, the anti-PD-1 antibody is administered every three weeks, such as on day 1 of a 21-day cycle. In some embodiments, the anti-PD-1 antibody is administered for at least one cycle prior to administration of the mTOR inhibitor nanoparticle composition. In some embodiments, the anti-PD-1 antibody is administered at an amount of about 1 mg/kg to about 10 mg/kg, such as about 3 mg/kg.

D. Treatment of other cancers

[0107] In some aspects, provided is a method of treating a cancer in an individual in need thereof, wherein the cancer is selected from the group consisting of adenocarcinoma of ascending colon, cervical adenosquamous cell carcinoma, chondrosarcoma, chordoma, clear cell sarcoma, colorectal cancer, such as metastatic colorectal cancer, desmoplastic round cell tumor, desmoplastic small cell tumor, Ewing Sarcoma, myxoid liposarcoma, osteosarcoma, including osteogenic osteosarcoma, pleomorphic spindle cell sarcoma, serous carcinoma of endometrium, or synovial sarcoma, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and optionally (b) an effective amount of a second therapeutic agent (e.g., an anti-PD-1 antibody).

[0108] In some embodiments, the cancer is chondrosacoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of TP53 p.Y220C, TUBB3, AXL. In some embodiments, the cancer is chondrosacoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of BRCA2 truncation intron 7, EZH2 truncation intron 19, MLL2 R5048H, loss of RBI exons 3-9, and TP53 RllOdel. In some embodiments, the cancer is chondrosacoma, and the individual has an aberration at IDH1. In some embodiments, the cancer is chondrosacoma, and the individual has an aberration at IDH1. In some embodiments, the cancer is chondrosacoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of statuses selected from the group consisting of tumor mutational burden (TMB-low), mismatch repair (MMR)-proficient, PD-L1 -negative, MLH1 -positive, MLH2-positive, MLH6-positive, and PMS2-positive.

[0109] In some embodiments, the cancer is osteosarcoma, and the individual has a C17orf39 mutation and is MUTYH-positive. In some embodiments, the cancer is osteosarcoma, and the individual has an aberration at DPBl*04:01 and/or DPBI*04:02; and/or is DPw4-positive. In some embodiments, the cancer is chondrosacoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of BRCA2, FGFR1, KDM5A, MAF, MY018A, PCLO, RELN, and SF3B1; and/or the individual is RBI -positive and TP53-positive. In some embodiments, the cancer is osteosarcoma, and the individual is microsatellite (MSI)-stable, MMR-proficient, TMB-intermediate, and/or TP53- positive. In some embodiments, the cancer is osteosarcoma, and the individual is TMB- intermediate, PD-L1 -negative, MLH1 -positive, MLH2 -positive, MLH6-positive, PMS2-positive and/or has an aberration in TP53.

[0110] In some embodiments, the cancer is Ewing Sarcoma, and the individual has an aberration in EWSR1 (e.g., a gene fusion comprising at least a portion of the EWSR1 gene) and/or is KMT2D-proficient. In some embodiments, the cancer is Ewing Sarcoma, and the individual has an aberration in EWSR1 (e.g., a gene fusion comprising at least a portion of the EWSR1 gene, e.g., a EWSRl-FLIl fusion), and/or is tumor exonic mutational burden-high, MLH1 -positive, MLH2-positive, MLH6-positive, and/or PMS2-positive. In some embodiments, the cancer is Ewing Sarcoma, and the individual has an aberration in EWSR1 (e.g., a gene fusion comprising at least a portion of the EWSR1 gene) and/or EZH2 (e.g. a pathogenic variant of EZH2), and/or is MLH1 -positive, MLH2-positive, MLH6-positive, and/or PMS2-positive.

[oni] In some embodiments, the cancer is synovial sarcoma, and the individual is NY- ESO-1 -positive (e.g., NY-ESO-1 -positive in 99% of tumor cells).

[0112] In some embodiments, the cancer is desmoplastic round cell tumor, and the individual has an aberration in EWSR1 (e.g., a gene fusion comprising at least a portion of the EWSR1 gene, e.g., EWSR1-WT1 fusion) and/or is TMB-low.

[0113] In some embodiments, the cancer is clear cell sarcoma, and the individual is EWSR- positive.

[0114] In some embodiments, the cancer is chordoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of ARAF, ARID 1 A, ASXL1, BAP1, DNMT3A, FANCE, FGFR4, FLT3, FLT4, JAK3, KDM5C, NF1, PTCH, TERT, and TP53; and/or is MSI-stable, TMB-low, TSC2-positive, and/or INI-1 positive. [0115] In some embodiments, the cancer is serous carcinoma of endometrium, and the individual has an aberration at PIK3CA and/or is TP53-positive.

[0116] In some embodiments, the cancer is metastatic colorectal cancer, and the individual has an aberration in CRKL (e.g., amplification of the CRKL locus) and/or TP53 (e.g., a pathogenic variant of TP53, e.g., p.T253P); and/or is MSI-high, MMR-deficient (dMMR), PTEN-positive, MLH1 -positive, MLH2 -positive, MLH6-positive, and/or PMS2-positive.

[0117] In some embodiments, the cancer is cervical adenosquamous cell carcinoma, and the individual has an aberration in ERBB2 and/or AKT2 (e.g., amplification of the AKT2 locus); and/or is PD-L1 -positive and/or TMB-intermediate.

[0118] In some embodiments, the cancer is osteogenic osteosarcoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of BRCA2, FGFR1, KDM5A, MAF, MYO 18 A, PCLO, RELN, and SF3B1; and/or is RBI-positive and/or TP53 -positive.

[0119] In some embodiments, the cancer is adenocarcinoma of ascending colon, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of KRAS, NRAS, APC, BRCA, PIK3CA, SMAD4, and TP53; and/or is TMB-intermediate.

[0120] In some embodiments, the cancer is pleomorphic Spindle cell sarcoma, and the individual has an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from the group consisting of APC, ATRX, BRCA2, FBXW7, FLT1, GNA13, IRS2, JAK3, KMT2A, MED12, MLL2, PRKAR1 A, and SOX9; and/or is microsatellite stable (MSS), TMB-low, CCND3-positive, RBI-positive, VEGFA-positive, PDL-1 -positive, and/or TSC2- positive.

[0121] In some embodiments as used herein, an individual being “-positive” for a biomarker, e.g., “RBI -positive,” means the individual has detectable expression of a biomarker, such as detectable via standard biomarker measurement techniques including immunohistochemistry, mass spectrometry, PCR, or sequencing. In some embodiments, the expression of a biomarker is detectable when normalized reads per kilobase million (RPKM), normalized fragments per kilobase million (FPKM) or normalized transcripts per kilobase million (TPM) of the biomarker is greater than or equal to 1. In some embodiments as used herein, an individual being “- negative” for a biomarker, e.g., “PD-L1 -negative,” means the individual has no detectable expression of a biomarker, such as detectable via standard biomarker measurement techniques including immunohistochemistry, mass spectrometry, PCR, or sequencing.

E. Biomarkers

[0122] In certain aspects of the methods provided herein, the individual is selected for a treatment described herein on a basis of having an aberration at one or more (e.g., two or more, e.g., three or more) of the genes selected from AKT1, AKT2, APC, ARAF, ARID1A, ASXL1, ATRX, AXL, BAP1, BRCA, BRCA2, C17orf39, CCND3, CRKL, DNMT3A, ERBB2, EWSR1, EZH2, FANCE, FBXW7, FGFR1, FGFR4, FLU, FLT1, FLT3, FLT4, GNA13, HLA-A2, HLA- DPB1, IDH1, INI-1, IRS2, JAK3, KDM5A, KDM5C, KMT2A, KMT2D, KRAS, MAF, MED12, MLL2, MUTYH, MY018A, NF1, NRAS, NTRK1/2/3, NY-ESO-1, PCLO, PIK3CA, PRKAR1A, PTCH, PD-L1, PTEN, RBI, RELN, SF3B1, SMAD4, SOX9, TERT, TP53, TSC2, TUBB3, VEGFA, WT1. In some embodiments, the individual is selected for treatment on a basis of an aberration at PTEN and/or TSC2. In some embodiments, the aberration at PTEN is a PTEN loss. In some embodiments, the aberration at TSC2 is a TSC2 mutation. In some embodiments, the aberration at BRCA2 is a truncation in BRCA2 intron 7. In some embodiments, the aberration at EZH2 is a truncation in intron 19. In some embodiments the aberration at EWSR1 is a EWSR1 fusion. In some embodiments, the EWSR1 fusion is a EWSR1-FLI1 fusion. In some embodiments, the aberration at EWSR1 is a EWSR1 gene rearrangement. In some embodiments the aberration at FLU is a FLU fusion. In some embodiments, the FLU fusion is a EWSRl-FLIl fusion. In some embodiments, the aberration at MLL2 is a R5048H mutation. In some embodiments, the aberration at RBI is loss of RBI exons 3-9. In some embodiments, the aberration at TP53 is TP53 p.Y220C. In some embodiments, the aberration at TP53 is TP53 p.T253P. In some embodiments, the aberration at TP53 is TP53 RllOdel. In some embodiments, the aberration at AKT2 is a genetic amplification of AKT2. In some embodiments, the aberration at CRKL is a genetic amplification of CRKL. In some embodiments, the aberration in HLA-DPB1 is DPBl*04:01 and/or DPBI*04:02. In some embodiments, the individual is selected for treatment on a basis of not having a fusion in NTRK1/2/3.

[0123] In some embodiments, the individual is selected for treatment on a basis of having detectable expression of a biomarker, such as detectable via standard biomarker measurement techniques including immunohistochemistry, mass spectrometry, PCR, or sequencing. In some embodiments, the expression of a biomarker is detectable when normalized reads per kilobase million (RPKM), normalized fragments per kilobase million (FPKM) or normalized transcripts per kilobase million (TPM) of the biomarker is greater than or equal to 1. In some embodiments, the individual is selected for treatment on a basis of being DPw4-positive. In some embodiments, the individual is selected for treatment on a basis of being INI-1 positive. In some embodiments, the individual is selected for treatment on a basis of being NYESO positive and HLA-A2 positive. In some embodiments, the individual is selected for treatment on a basis of being PTEN-positive. In some embodiments, the individual is selected for treatment on a basis of not having detectable expression of a biomarker. In some embodiments, the individual is selected for treatment on a basis of being PD-L1 -negative. In some embodiments, the individual is selected for treatment on a basis of having a mutational status or genotype. In some embodiments, the individual is selected for treatment on a basis of being tumor mutational burden (TMB)-low. In some embodiments, the individual is selected for treatment on a basis of being TMB-intermediate. In some embodiments, the individual is selected for treatment on a basis of being tumor exonic mutational burden high. In some embodiments, the individual is selected for treatment on a basis of being microsatellite stable (MSS). In some embodiments, the individual is selected for treatment on a basis of having high microsatellite instability and/or mismatch repair (MMR)-deficient.

[0124] In some embodiments, tumor mutational burden (TMB) is the number of somatic (non-inherited) mutations per megabase (Mb) of genome. TMB can be used as a metric for predicting outcomes such as cancer treatment success and survival probability. TMB can be measured using high throughput sequencing techniques such as NGS. Different cancers may have variations in TMB levels. TMB levels can be categorized as high, intermediate and low, based on the number of mutations/Mb. For example, in some embodiments, TMB levels can be set as follows: (i) low - 1-5 mutations/Mb; (ii) intermediate - 6-15 mutations/Mb; and (iii) high - 16 or more mutations/Mb.

[0125] Mismatch repair (MMR) is the system for correcting base pair mismatches, insertion or deletion errors during DNA replication, and important for maintaining genomic stability. Defects in MMR are associated with genome instability and increased sporadic mutations in microsatellite regions. MMR status may be assessed by immunohistochemistry staining by detecting the presence or absence of proteins involved in MMR regulation: MLH1, MSH2, MSH6, and PMS2. MMR status may be categorized as MMR proficient (pMMR) or MMR deficient (dMMR). pMMR status is associated with a functional MMR pathway, while dMMR is associated with lacking one or more of the MMR proteins.

[0126] Microsatellite instability (MSI) is a phenotype that results when DNA mismatch repair is defective. MSI is a marker for cancer MMR status. MSI results in the insertion or deletion of repetitive sequences in the genome. MSI status is tested using PCR-based assays to determine the instability using five markers: BAT-25, BAT-26, D2S123, D5S346 and D17S250. MSI status can be subdivided into high microsatellite instability (MSI-H: two or more markers are unstable), low microsatellite instability (MSI-L: one marker is unstable), and microsatellite stable (MSS: no markers show instability).

[0127] As used herein, “based upon” includes assessing, determining, or measuring the individual’s characteristics as described herein (and preferably selecting an individual suitable for receiving treatment). When the status of an aberration is “used as a basis” for selection, assessing, measuring, or determining method of treatment as described herein, the aberration at one or more genes is determined before and/or during treatment, and the status (including presence, absence, expression level, activity level and/or phosphorylation level of the aberration) obtained is used by a clinician in assessing any of the following: (a) probable or likely suitability of an individual to initially receive treatment(s); (b) probable or likely unsuitability of an individual to initially receive treatment(s); (c) responsiveness to treatment; (d) probable or likely suitability of an individual to continue to receive treatment(s); (e) probable or likely unsuitability of an individual to continue to receive treatment(s); (I) adjusting dosage; or (g) predicting likelihood of clinical benefits. As used herein, when an individual is selected for treatment “on a basis” of having an aberration (including presence, absence, expression level, activity level and/or phosphorylation level of the aberration) at one or more genetic aberrations, individual may be selected for treatment on more than one basis, and the selection basis for treatment may comprise further criteria.

F. Techniques for determining a status of a cancer and/or individual

[0128] In certain aspects of the method provided herein, the undifferentiated pleomorphic sarcoma is characterized as having a PTEN loss and/ or a TSC2 mutation. In certain aspects of the method provided herein, the leiomyosarcoma is characterized as being estrogen receptorpositive leiomyosarcoma. In some embodiments, encompassed in the methods provided herein are steps for assessing and/ or determining the status of a cancer, such as undifferentiated pleomorphic sarcoma and/ or leiomyosarcoma, in an individual. In some embodiments, the method comprises obtaining a sample of the cancer, such as undifferentiated pleomorphic sarcoma and/ or leiomyosarcoma, from an individual. In some embodiments, the method comprises determining the presence, absence, or level of a biomarker, e.g, presence of a PTEN loss, such as via protein expression, sequencing, or an activity assay. In some embodiments, the method comprises determining the presence of a TSC2 mutation, such as via protein expression, sequencing, or an activity assay. In some embodiments, the method comprises determining the presence, absence, or level of an estrogen receptor, such as via protein expression, sequencing, or an activity assay.

[0129] The status of a cancer and/or an individual, such as in undifferentiated pleomorphic sarcoma and/ or leiomyosarcoma, can be assessed or determined by analyzing a sample from the individual. The assessment may be based on fresh tissue samples or archived tissue samples. Suitable samples include, but are not limited to, tumor tissue, normal tissue adjacent to the tumor tissue, normal tissue distal to the tumor tissue, or peripheral blood lymphocytes. In some embodiments, the sample is a tumor tissue. In some embodiments, the sample is a biopsy containing tumor cells, such as fine needle aspiration of solid tumor cells or laparoscopy obtained tumor cells. In some embodiments, the biopsied cells are centrifuged into a pellet, fixed, and embedded in paraffin prior to the analysis. In some embodiments, the biopsied cells are flash frozen prior to the analysis. In some embodiments, the sample is a plasma sample.

[0130] In some embodiments, the sample comprises a circulating metastatic cancer cell. In some embodiments, the sample is obtained by sorting circulating tumor cells (CTCs) from blood. In some further embodiments, the CTCs have detached from a primary tumor and circulate in a bodily fluid. In some further embodiments, the CTCs have detached from a primary tumor and circulate in the bloodstream. In some embodiments, the CTCs are an indication of metastasis.

[0131] In some embodiments, the sample is mixed with an antibody that recognizes a molecule (such as a protein) or fragment thereof. In some embodiments, the sample is mixed with a nucleic acid that recognizes nucleic acids (such as DNA or RNA) or fragment thereof. In some embodiments, the sample is used for sequencing analysis, such as next-generation DNA, RNA and/or exome sequencing analysis. [0132] Samples may be assessed before the start of the treatment, at any time during the treatment, and/or at the end of the treatment.

III. Compositions comprising nanoparticles comprising an mTOR inhibitor

[0133] The mTOR inhibitor nanoparticle compositions described herein comprise nanoparticles comprising (in various embodiments consisting essentially of or consisting of) an mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) and an albumin (such as human serum albumin). Nanoparticles of poorly water soluble drugs (such as macrolides) have been disclosed in, for example, U. S. Pat. Nos.5, 916, 596; 6,506,405; 6,749,868, 6,537,579, 7,820,788, and 8,911,786, 11,497,737, and also in U. S. Pat. Pub. Nos. 2006/0263434, and 2007/0082838; PCT Patent Application W008/137148, U.S. Patent Application No.: 62/927,047, each of which is incorporated herein by reference in their entirety.

[0134] In some embodiments, the composition comprises nanoparticles with an average or mean diameter of no greater than about 1000 nanometers (nm), such as no greater than about any of 900, 800, 700, 600, 500, 400, 300, 200, and 100 nm. In some embodiments, the average or mean diameters of the nanoparticles is no greater than about 200 nm. In some embodiments, the average or mean diameters of the nanoparticles is no greater than about 150 nm. In some embodiments, the average or mean diameters of the nanoparticles is no greater than about 100 nm. In some embodiments, the average or mean diameter of the nanoparticles is about 10 to about 400 nm. In some embodiments, the average or mean diameter of the nanoparticles is about 10 to about 150 nm. In some embodiments, the average or mean diameter of the nanoparticles is about 40 to about 120 nm. In some embodiments, the average or mean diameter of the nanoparticles are no less than about 50 nm. In some embodiments, the nanoparticles are sterile- filterable.

[0135] Methods of determining average particle sizes are known in the art, for example, dynamic light scattering (DLS) has been routinely used in determining the size of submicrometre-sized particles based. International Standard ISO22412 Particle Size Analysis - Dynamic Light Scattering, International Organisation for Standardisation (ISO) 2008 and Dynamic Light Scattering Common Terms Defined, Malvern Instruments Limited, 2011. In some embodiments, the particle size is measured as the volume-weighted mean particle size (Dv50) of the nanoparticles in the composition. [0136] In some embodiments, the nanoparticles comprise the mTOR inhibitor associated with the albumin. In some embodiments, the nanoparticles comprise the mTOR inhibitor coated with the albumin.

[0137] In some embodiments, the albumin has sulfhydryl groups that can form disulfide bonds. In some embodiments, at least about 5% (including for example at least about any one of 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%) of the albumin in the nanoparticle portion of the composition are crosslinked (for example crosslinked through one or more disulfide bonds).

[0138] In some embodiments, the nanoparticles comprising the mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) are associated (e.g, coated) with an albumin (such as human albumin or human serum albumin). In some embodiments, the composition comprises an mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) in both nanoparticle and non-nanoparticle forms (e.g, in the form of solutions or in the form of soluble albumin/nanoparticle complexes), wherein at least about any one of 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the mTOR inhibitor in the composition are in nanoparticle form. In some embodiments, the mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) in the nanoparticles constitutes more than about any one of 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the nanoparticles by weight. In some embodiments, the nanoparticles have a non-polymeric matrix. In some embodiments, the nanoparticles comprise a core of an mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) that is substantially free of polymeric materials (such as polymeric matrix).

[0139] In some embodiments, the composition comprises an albumin in both nanoparticle and non-nanoparticle portions of the composition, wherein at least about any one of 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the albumin in the composition are in non-nanoparticle portion of the composition.

[0140] In some embodiments, the weight ratio of the albumin to the mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) in the mTOR inhibitor nanoparticle composition is such that a sufficient amount of mTOR inhibitor binds to, or is transported by, the cell. While the weight ratio of an albumin to an mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) will have to be optimized for different albumin and mTOR inhibitor combinations, generally the weight ratio of an albumin to an mTOR inhibitor (such as a limus drug, e.g., rapamycin or a derivative thereof) (w/w) is about 0.01:1 to about 100:1, about 0.02:1 to about 50:1, about 0.05:1 to about 20:1, about 0.1:1 to about 20:1, about 1:1 to about 18:1, about 2: 1 to about 15:1, about 3:1 to about 12: 1, about 4:1 to about 10:1, about 5:1 to about 9: 1 , or about 9: 1. In some embodiments, the albumin to mTOR inhibitor (such as a limus drug, e.g., rapamycin or a derivative thereof) weight ratio is about any of 18:1 or less, 15:1 or less, 14:1 or less, 13:1 or less, 12:1 or less, 11:1 or less, 10: 1 or less, 9:1 or less, 8:1 or less, 7:1 or less, 6:1 or less, 5:1 or less, 4:1 or less, and 3:1 or less. In some embodiments, the weight ratio of the albumin (such as human albumin or human serum albumin) to the mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) in the composition is any one of the following: about 1:1 to about 18:1, about 1:1 to about 15:1, about 1:1 to about 12:1, about 1:1 to about 10:1, about 1:1 to about 9:1, about 1:1 to about 8:1, about 1: 1 to about 7:1, about 1:1 to about 6: 1, about 1 : 1 to about 5:1, about 1 : 1 to about 4:1, about 1 : 1 to about 3:1, about 1 : 1 to about 2:1, about 1 : 1 to about 1:1.

[0141] In some embodiments, the composition comprises nanoparticles comprising an mTOR inhibitor and an albumin, wherein the weight ratio of the albumin to the mTOR inhibitor in the composition is about 0.01:1 to about 100:1. In some embodiments, the composition comprises nanoparticles comprising an mTOR inhibitor (such as rapamycin) and an albumin, wherein the weight ratio of the albumin to the mTOR inhibitor (such as rapamycin) in the composition is about 18:1 or less (including for example any of about 1:1 to about 18:1, about 2:1 to about 15:1, about 3:1 to about 12:1, about 4:1 to about 10:1, about 5:1 to about 9:1, and about 9: 1). In some embodiments, the composition comprises nanoparticles comprising rapamycin, or a derivative thereof, and an albumin, wherein the weight ratio of the albumin to the rapamycin or derivative thereof in the composition is about 18:1 or less (including for example any of about 1:1 to about 18:1, about 2:1 to about 15:1, about 3:1 to about 12: 1, about 4:1 to about 10:1, about 5:1 to about 9:1, and about 9:1). In some embodiments, the mTOR inhibitor (such as rapamycin) is coated with albumin.

[0142] In some embodiments, the mTOR inhibitor nanoparticle composition (such as rapamycin/albumin nanoparticle composition) comprises one or more of the above characteristics.

[0143] The nanoparticles described herein may be present in a dry formulation (such as lyophilized composition) or suspended in a biocompatible medium. Suitable biocompatible media include, but are not limited to, water, buffered aqueous media, saline, buffered saline, optionally buffered solutions of amino acids, optionally buffered solutions of proteins, optionally buffered solutions of sugars, optionally buffered solutions of vitamins, optionally buffered solutions of synthetic polymers, lipid-containing emulsions, and the like.

[0144] In some embodiments, the pharmaceutically acceptable carrier comprises an albumin (such as human albumin or human serum albumin). The albumin may either be natural in origin or synthetically prepared. In some embodiments, the albumin is human albumin or human serum albumin. In some embodiments, the albumin is a recombinant albumin.

[0145] Human serum albumin (HSA) is a highly soluble globular protein of Mr 65K and consists of 585 amino acids. HSA is the most abundant protein in the plasma and accounts for 70-80 % of the colloid osmotic pressure of human plasma. The amino acid sequence of HSA contains a total of 17 disulfide bridges, one free thiol (Cys 34), and a single tryptophan (Trp 214). Intravenous use of HSA solution has been indicated for the prevention and treatment of hypovolemic shock (see, e.g, Tullis, JAMA, 237: 355-360, 460-463, (1977)) and Houser et al., Surgery, Gynecology and Obstetrics, 150: 811-816 (1980)) and in conjunction with exchange transfusion in the treatment of neonatal hyperbilirubinemia (see, e.g., Finlayson, Seminars in Thrombosis and Hemostasis, 6, 85-120, (1980)). Other albumins are contemplated, such as bovine serum albumin. Use of such non-human albumins could be appropriate, for example, in the context of use of these compositions in non-human mammals, such as the veterinary (including domestic pets and agricultural context). Human serum albumin (HSA) has multiple hydrophobic binding sites (a total of eight for fatty acids, an endogenous ligand of HSA) and binds a diverse set of drugs, especially neutral and negatively charged hydrophobic compounds (Goodman et al., The Pharmacological Basis of Therapeutics, 9^th ed, McGraw-Hill New York (1996)). Two high affinity binding sites have been proposed in subdomains IIA and IIIA of HSA, which are highly elongated hydrophobic pockets with charged lysine and arginine residues near the surface which function as attachment points for polar ligand features (see, e.g., Fehske et al., Biochem. Pharmcol., 30, 687-92 (198a), Vorum, Dan. Med. Bull., 46, 379-99 (1999), Kragh-Hansen, Dan. Med. Bull., 1441, 131-40 (1990), Curry et al., Nat. Struct. Biol., 5, 827-35 (1998), Sugio et al., Protein. Eng., 12, 439-46 (1999), He et al., Nature, 358, 209-15 (199b), and Carter et al., Adv. Protein. Chem., 45, 153-203 (1994)). Rapamycin and propofol have been shown to bind HSA (see, e.g., Paal et al., Eur. J. Biochem., 268(7), 2187-91 (200a), Purcell et al., Biochem. Biophys. Acta, 1478(a), 61-8 (2000), AAXmayex et al., Arzneimittelforschung, 45, 1053-6 (1995), and Garrido et al., Rev. Esp. Anestestiol. Reanim., 41, 308-12 (1994)). In addition, docetaxel has been shown to bind to human plasma proteins (see, e.g., Urien et al., Invest. New Drugs, 14(b), 147-51 (1996)).

[0146] An mTOR inhibitor (such as a limus drug, e.g. , rapamycin or a derivative thereol) is “stabilized” in an aqueous suspension if it remains suspended in an aqueous medium (such as without visible precipitation or sedimentation) for an extended period of time, such as for at least about any of 0.1, 0.2, 0.25, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 24, 36, 48, 60, or 72 hours. The suspension is generally, but not necessarily, suitable for administration to an individual (such as a human). Stability of the suspension is generally (but not necessarily) evaluated at a storage temperature (such as room temperature (such as 20-25 °C) or refrigerated conditions (such as 4 °C)). For example, a suspension is stable at a storage temperature if it exhibits no flocculation or particle agglomeration visible to the naked eye or when viewed using an optical microscope at 1000 times, at about fifteen minutes after preparation of the suspension. Stability can also be evaluated under accelerated testing conditions, such as at a temperature that is about 40 °C or higher.

[0147] The compositions described herein may be a stable aqueous suspension of the mTOR inhibitor, such as a stable aqueous suspension of the mTOR inhibitor at a concentration of any of about 0.1 to about 200 mg/ml, about 0.1 to about 150 mg/ml, about 0.1 to about 100 mg/ml, about 0.1 to about 50 mg/ml, about 0.1 to about 20 mg/ml, about 1 to about 10 mg/ml, about 2 mg/ml to about 8 mg/ml, about 4 to about 6 mg/ml, and about 5 mg/ml. In some embodiments, the concentration of the mTOR inhibitor is at least about any of 0.2 mg/ml, 1.3 mg/ml, 1.5 mg/ml, 2 mg/ml, 3 mg/ml, 4 mg/ml, 5 mg/ml, 6 mg/ml, 7 mg/ml, 8 mg/ml, 9 mg/ml, 10 mg/ml, 15 mg/ml, 20 mg/ml, 25 mg/ml, 30 mg/ml, 40 mg/ml, 50 mg/ml, 100 mg/ml, 150 mg/ml, or 200 mg/ml.

[0148] In some embodiments, the albumin is present in an amount that is sufficient to stabilize the mTOR inhibitor (such as a limus drug, e.g., rapamycin or a derivative thereol) in an aqueous suspension at a certain concentration. For example, the concentration of the mTOR inhibitor (such as a limus drug, e.g., rapamycin or a derivative thereof) in the composition is about 0.1 to about 100 mg/ml, including for example about any of 0.1 to about 50 mg/ml, about 0. 1 to about 20 mg/ml, about 1 to about 10 mg/ml, about 2 mg/ml to about 8 mg/ml, about 4 to about 6 mg/ml, or about 5 mg/ml. In some embodiments, the concentration of the mTOR inhibitor (such as a limus drug, e.g., rapamycin or a derivative thereof) is at least about any of 1.3 mg/ml, 1.5 mg/ml, 2 mg/ml, 3 mg/ml, 4 mg/ml, 5 mg/ml, 6 mg/ml, 7 mg/ml, 8 mg/ml, 9 mg/ml, 10 mg/ml, 15 mg/ml, 20 mg/ml, 25 mg/ml, 30 mg/ml, 40 mg/ml, and 50 mg/ml. In some embodiments, the albumin is present in an amount that avoids use of surfactants (such as Cremophor), so that the composition is free or substantially free of surfactant (such as Cremophor).

[0149] In some embodiments, the composition, in liquid form, comprises from about 0.1% to about 50% (w/v) (e.g., about 0.5% (w/v), about 5% (w/v), about 10% (w/v), about 15% (w/v), about 20% (w/v), about 30% (w/v), about 40% (w/v), or about 50% (w/v)) of an albumin. In some embodiments, the composition, in liquid form, comprises about 0.5% to about 5% (w/v) of albumin.

[0150] In some embodiments, the albumin allows the composition to be administered to an individual (such as a human) without significant side effects. In some embodiments, the albumin (such as human serum albumin or human albumin) is in an amount that is effective to reduce one or more side effects of administration of the mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) to a human. The term “reducing one or more side effects” of administration of the mTOR inhibitor (such as a limus drug, e.g, rapamycin or a derivative thereof) refers to reduction, alleviation, elimination, or avoidance of one or more undesirable effects caused by the mTOR inhibitor, as well as side effects caused by delivery vehicles (such as solvents that render the limus drugs suitable for injection) used to deliver the mTOR inhibitor. Such side effects include, for example, myelosuppression, neurotoxicity, hypersensitivity, inflammation, venous irritation, phlebitis, pain, skin irritation, peripheral neuropathy, neutropenic fever, anaphylactic reaction, venous thrombosis, extravasation, and combinations thereof. These side effects, however, are merely exemplary and other side effects, or combination of side effects, associated with limus drugs (such as a limus drug, e.g, rapamycin or a derivative thereof) can be reduced.

[0151] In some embodiments, the composition is a dry (such as lyophilized) composition that can be reconstituted, resuspended, or rehydrated to form generally a stable aqueous suspension of the nanoparticles comprising an mTOR inhibitor and an albumin. In some embodiments, the composition is a liquid (such as aqueous) composition obtained by reconstituting or resuspending a dry composition. In some embodiments, the composition is an intermediate liquid (such as aqueous) composition that can be dried (such as lyophilized).

A. mTOR inhibitors

[0152] The methods described herein in some embodiments comprise administration of nanoparticle compositions of mTOR inhibitors. “mTOR inhibitor” used herein refers to an inhibitor of mTOR. mTOR is a serine/threonine-specific protein kinase downstream of the phosphatidylinositol 3-kinase (PI3K)/Akt (protein kinase B) pathway, and a key regulator of cell survival, proliferation, stress, and metabolism. mTOR pathway dysregulation has been found in many human carcinomas, and mTOR inhibition produced substantial inhibitory effects on tumor progression.

[0153] The mammalian target of rapamycin (mTOR) (also known as mechanistic target of rapamycin or FK506 binding protein 12-rapamycin associated protein 1 (FRAP1)) is an atypical serine/threonine protein kinase that is present in two distinct complexes, mTOR Complex 1 (mTORCl) and mTOR Complex 2 (mT0RC2). mTORCl is composed of mTOR, regulatory- associated protein of mTOR (Raptor), mammalian lethal with SEC 13 protein 8 (MLST8), PRAS40 and DEPTOR (Kim et al. (2002). Cell 110: 163-75; Fang et al. (2001). Science 294 (5548): 1942-5). mTORCl integrates four major signal inputs: nutrients (such as amino acids and phosphatidic acid), growth factors (insulin), energy and stress (such as hypoxia and DNA damage). Amino acid availability is signaled to mTORCl via a pathway involving the Rag and Ragulator (LAMTOR1-3) Growth factors and hormones (e.g, insulin) signal to mTORCl via Akt, which inactivates TSC2 to prevent inhibition of mTORCl. Alternatively, low ATP levels lead to the AMPK-dependent activation of TSC2 and phosphorylation of raptor to reduce mTORCl signaling proteins.

[0154] Active mTORCl has a number of downstream biological effects including translation of mRNA via the phosphorylation of downstream targets (4E-BP1 and p70 S6 Kinase), suppression of autophagy (Atgl3, ULK1), ribosome biogenesis, and activation of transcription leading to mitochondrial metabolism or adipogenesis. Accordingly, mTORCl activity promotes either cellular growth when conditions are favorable or catabolic processes during stress or when conditions are unfavorable. [0155] mTORC2 is composed of mTOR, rapamycin-insensitive companion of mTOR (RICTOR), G[3L, and mammalian stress-activated protein kinase interacting protein 1 (mSINl). In contrast to mTORCl, for which many upstream signals and cellular functions have been defined (see above), relatively little is known about mTORC2 biology. mT0RC2 regulates cytoskeletal organization through its stimulation of F-actin stress fibers, paxillin, RhoA, Rael, Cdc42, and protein kinase C a (PKCa). It had been observed that knocking down mT0RC2 components affects actin polymerization and perturbs cell morphology (Jacinto et al. (2004). Nat. Cell Biol. 6, 1122-1128; Sarbassov et al. (2004). Curr. Biol. 14, 1296-1302). This suggests that mTORC2 controls the actin cytoskeleton by promoting protein kinase Ca (PKCa) phosphorylation, phosphorylation of paxillin and its relocalization to focal adhesions, and the GTP loading of RhoA and Rael. The molecular mechanism by which mT0RC2 regulates these processes has not been determined.

[0156] In some embodiments, the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) is an inhibitor of mTORCl. In some embodiments, the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) is an inhibitor of mT0RC2. In some embodiments, the mTOR inhibitor (such as a limus drug, e.g, sirolimus or a derivative thereof) is an inhibitor of both mTORCf and mT0RC2.

[0157] In some embodiments, the mTOR inhibitor is a limus drug, which includes sirolimus and its analogs. Examples of limus drugs include, but are not limited to, temsirolimus (CCI- 779), everolimus (RAD001), ridaforolimus (AP-23573), deforolimus ( MK-8669), zotarolimus (ABT-578), pimecrolimus, and tacrolimus (FK-506). In some embodiments, the limus drug is selected from the group consisting of temsirolimus (CCI-779), everolimus (RAD001), ridaforolimus (AP-23573), deforolimus (MK-8669), zotarolimus (ABT-578), pimecrolimus, and tacrolimus (FK-506). In some embodiments, the mTOR inhibitor is an mTOR kinase inhibitor, such as CC-115 or CC-223.

[0158] In some embodiments, the mTOR inhibitor is sirolimus. Sirolimus is macrolide antibiotic that complexes with FKBP-12 and inhibits the mTOR pathway by binding mTORCl.

[0159] In some embodiments, the mTOR inhibitor is selected from the group consisting of sirolimus (rapamycin), BEZ235 (NVP-BEZ235), everolimus (also known as RAD001, Zortress, Certican, and Afinitor), AZD 8055, temsirolimus (also known as CCI-779 and Torisel), CC-115, CC-223, PI-103, Ku-0063794, INK 128, AZD2014, NVP-BGT226, PF-04691502, CH5132799, GDC-0980 (RG7422), Torin 1, WAY-600, WYE-125132, WYE-687, GSK2126458, PF- 05212384 (PKI-587), PP-121, OSI-027, Palomid 529, PP242, XL765, GSK1059615, WYE-354, and ridaforolimus (also known as deforolimus).

[0160] BEZ235 (NVP-BEZ235) is an imidazoquilonine derivative that is an mTORCl catalytic inhibitor (Roper J, et al. PLoS One, 2011, 6(9), e25132). Everolimus is the 40-O-(2- hydroxy ethyl) derivative of sirolimus and binds the cyclophilin FKBP-12, and this complex also mTORCl. AZD8055 is a small molecule that inhibits the phosphorylation of mTORCl (p70S6K and 4E-BP1). Temsirolimus is a small molecule that forms a complex with the FK506-binding protein and prohibits the activation of mTOR when it resides in the mTORCl complex. PI-103 is a small molecule that inhibits the activation of the rapamycin-sensitive (mTORCl) complex (Knight et al. (2006) Cell. 125: 733-47). KU-0063794 is a small molecule that inhibits the phosphorylation of mTORCl at Ser2448 in a dose-dependent and time-dependent manner. INK 128, AZD2014, NVP-BGT226, CH5132799, WYE-687, and are each small molecule inhibitors of mTORCl. PF-04691502 inhibits mTORCl activity. GDC-0980 is an orally bioavailable small molecule that inhibits Class I PI3 Kinase and TORC1. Torin 1 is a potent small molecule inhibitor of mTOR. WAY-600 is a potent, ATP-competitive and selective inhibitor of mTOR. WYE-125132 is an ATP-competitive small molecule inhibitor of mTORCl. GSK2126458 is an inhibitor of mTORCl. PKI-587 is a highly potent dual inhibitor of PI3Ka, PI3Ky and mTOR. PP-121 is a multi-target inhibitor of PDGFR, Hck, mTOR, VEGFR2, Src and Abl. OSI-027 is a selective and potent dual inhibitor of mTORCl and mTORC2 with IC50 of 22 nM and 65 nM, respectively. Palomid 529 is a small molecule inhibitor of mTORCl that lacks affinity for ABCB1/ABCG2 and has good brain penetration (Lin et al. (2013) Int J Cancer DOI:

10.1002/ij c. 28126 (e-published ahead of print). PP242 is a selective mTOR inhibitor. XL765 is a dual inhibitor of mTOR/PI3k for mTOR, pl 10a, pl 10[3, pl lOy and pl 106. GSK1059615 is a novel and dual inhibitor of PI3Ka, PI3KJ3, PI3K8, PI3Ky and mTOR. WYE-354 inhibits mTORCl in HEK293 cells (0.2 pM-5 pM) and in HUVEC cells (10 nM-lpM). WYE-354 is a potent, specific and ATP-competitive inhibitor of mTOR. Deforolimus (Ridaforolimus, AP23573, MK-8669) is a selective mTOR inhibitor. C. Other components in the Nanoparticle Composition

[0161] In some embodiments, the composition is suitable for administration to a human. In some embodiments, the composition is suitable for administration to a mammal such as, in the veterinary context, domestic pets and agricultural animals. The following formulations and methods are merely exemplary and are in no way limiting. Formulations suitable for oral administration can consist of (a) liquid solutions, such as an effective amount of the compound dissolved in diluents, such as water, saline, or orange juice, (b) capsules, sachets or tablets, each containing a predetermined amount of the active ingredient, as solids or granules, (c) suspensions in an appropriate liquid, and (d) suitable emulsions. Tablet forms can include one or more of lactose, mannitol, com starch, potato starch, microcrystalline cellulose, acacia, gelatin, colloidal silicon dioxide, croscarmellose sodium, talc, magnesium stearate, stearic acid, and other excipients, colorants, diluents, buffering agents, moistening agents, preservatives, flavoring agents, and pharmacologically compatible excipients. Lozenge forms can comprise the active ingredient in a flavor, usually sucrose and acacia or tragacanth, as well as pastilles comprising the active ingredient in an inert base, such as gelatin and glycerin, or sucrose and acacia, emulsions, gels, and the like containing, in addition to the active ingredient, such excipients as are known in the art.

[0162] Examples of suitable carriers, excipients, and diluents include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, saline solution, syrup, methylcellulose, methyl- and propylhydroxybenzoates, talc, magnesium stearate, and mineral oil. The formulations can additionally include lubricating agents, wetting agents, emulsifying and suspending agents, preserving agents, sweetening agents or flavoring agents.

[0163] Formulations suitable for parenteral administration include aqueous and nonaqueous, isotonic sterile injection solutions, which can contain anti-oxidants, buffers, bacteriostats, and solutes that render the formulation compatible with the blood of the intended recipient, and aqueous and non-aqueous sterile suspensions that can include suspending agents, solubilizers, thickening agents, stabilizers, and preservatives. The formulations can be presented in unit-dose or multi-dose sealed containers, such as ampules and vials, and can be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid excipient, for example, water, for injections, immediately prior to use. Extemporaneous injection solutions and suspensions can be prepared from sterile powders, granules, and tablets of the kind previously described. Injectable formulations are preferred.

[0164] In some embodiments, the composition is formulated to have a pH range of about 4.5 to about 9.0, including for example pH ranges of about any of 5.0 to about 8.0, about 6.5 to about 7.5, and about 6.5 to about 7.0. In some embodiments, the pH of the composition is formulated to no less than about 6, including for example no less than about any of 6.5, 7, or 8 (such as about 8). The composition can also be made to be isotonic with blood by the addition of a suitable tonicity modifier, such as glycerol.

D. Albumin-based nanoparticle compositions of rapamycin

[0165] The methods described herein are particularly suitable for albumin-based nanoparticle compositions described herein in more details. The nanoparticle composition in some embodiments includes (a) nanoparticles that include rapamycin and albumin, and (b) a non-nanoparticle portion that includes rapamycin and albumin. The rapamycin and the albumin of the nanoparticles are associated with each other in the nanoparticles. For example, the nanoparticles may include a coating having the albumin, which surrounds a core comprising the rapamycin. In the non-nanoparticle portion of the composition, the rapamycin and the albumin may or may not associated with each other (z.e., the rapamycin may be in a reversible binding equilibrium with the albumin), but do not associate with each other in a manner that forms nanoparticles. That is, the nanoparticle composition may include nanoparticle-bound albumin and nanoparticle-bound rapamycin in the nanoparticle portion of the composition, and non- nanoparticle albumin and non-nanoparticle rapamycin in the non-nanoparticle portion of the composition. As used herein, “in the nanoparticles” is used synonymously with “in the nanoparticle portion.” The albumin of the nanoparticles may be further distinguishable from the albumin in the non-nanoparticle portion of the composition; for example, the oligomeric profile of the albumin in the nanoparticles may differ from the oligomeric profile of the albumin in the non-nanoparticle portion of the composition. The oligomer profile means the percentage of various albumin species compared with the total albumin in the composition. The types of albumin species includes albumin monomers, dimers, trimers, oligomers, and polymers. As used herein, “albumin monomers” or “monomeric albumin” refers to an albumin species having one, and only one, albumin unit; “albumin dimers” or “dimeric albumin” refers to an albumin species having two, and only two, albumin units; “albumin trimers” or “trimeric albumin” refers to albumin species having three, and only three, albumin units; “albumin polymers” refers to albumin species having a higher molecular weight than albumin monomers and albumin dimers; “albumin oligomers” or “oligomeric albumin” refers to lower molecular weight polymeric albumin species associated with a UV-based size-exclusion chromatography peak observed between a peak associated with albumin dimers and higher molecular weight polymeric albumin species.

[0166] The albumin of the nanoparticles associates with the rapamycin of the nanoparticles so that a nanoparticle suspension has a high concentration of rapamycin, which allows the composition to be used as a pharmaceutical composition for treating certain diseases, such as cancer. Manufactured nanoparticles (which may be made, for example, using the methods described herein) may be formulated, filtered, or otherwise processed to obtain the pharmaceutical composition, which may be suitable for medical use in a human individual.

[0167] Generally, to make the rapamycin pharmaceutical compositions described herein, rapamycin is dissolved in an organic solvent. Suitable organic solvents include, for example, ketones, esters, ethers, chlorinated solvents, and other solvents known in the art. For example, the organic solvent can be a mixture of methylene chloride/ethanol, chloroform/ethanol, or chloroform//c/7-butanol (for example with a ratio of about any one of 1:9, 1:8, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2, 1: 1, 2:1, 3:1, 4:1, 5:1, 6: 1, 7:1, 8:1, or 9:1 or with a ratio of about any one of 3:7, 5:7, 4:6, 5:5, 6:5, 8:5, 9:5, 9.5:5, 5:3, 7:3, 6:4, or 9.5:0.5). In some embodiments, the organic solvent comprises between about 10% and about 50% tert-butanol by volume. In some embodiments, the organic solvent comprises about any of 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% tert-butanol by volume. In some embodiments, the organic solvent comprises about any of 10-15%, 15-20%, 20-25%, 25-30%, 30-35%, 35-40%, 40-45%, or 45-50%, or any combination of such ranges, of tert-butanol by volume. In some embodiments, the organic solvent comprises between about 50% and about 90% chloroform by volume. In some embodiments, the organic solvent comprises about any of 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, or 90% chloroform by volume. In some embodiments, the organic solvent comprises about any of 50- 55%, 55-60%, 60-65%, 65-70%, 70-75%, 75-80%, 80-85%, or 85-90%, or any combination of such ranges, of chloroform by volume. In some embodiments, the organic solvent comprises between about 10% and about 50% tert-butanol by volume and between about 50% and about 90% chloroform by volume. In some embodiments, the organic solvent comprises chloroform and tert-butanol at a volumetric ratio of about 1:1 to about 1:9, such as about any of 1:1, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, and 9:1.

[0168] Albumin (such as recombinant albumin, for example NOVOZYME™ recombinant albumin or INTRIVIA™ recombinant albumin disclosed herein) is dissolved in an aqueous solution (such as water) and combined with the rapamycin solution to form a crude emulsion. The mixture is subjected to high pressure homogenization (e.g, using an Avestin, APV Gaulin, MICROFLUIDIZER™ such as a MICROFLUIDIZER™ Processor M-l 10EH from Microfluidics, Stansted, or Ultra Turrax homogenizer). The emulsion may be cycled through the high pressure homogenizer for between about 2 to about 100 cycles, such as about 5 to about 50 cycles or about 6 to about 20 cycles (e.g, about any one of 6, 8, 10, 12, 14, 16, 18 or 20 cycles). The organic solvent can then be removed by evaporation utilizing suitable equipment known for this purpose, including, but not limited to, rotary evaporators, falling film evaporators, wiped film evaporators, spray driers, and the like that can be operated in batch mode or in continuous operation. In some embodiments, the evaporator is a wiped film evaporator. The solvent may be removed at reduced pressure (such as at about any one of 25 mm Hg, 30 mm Hg, 40 mm Hg, 50 mm Hg, 100 mm Hg, 200 mm Hg, or 300 mm Hg). The amount of time used to remove the solvent under reduced pressure may be adjusted based on the volume of the formulation. For example, for a formulation produced on a 300 mL scale, the solvent can be removed at about 1 to about 300 mm Hg (e.g, about any one of 5-100 mm Hg, 10-50 mm Hg, 20-40 mm Hg, or 25 mm Hg) for about 5 to about 60 minutes (e.g, about any one of 7, 8, 9, 10, 11, 12, 13, 14, 15 16, 18, 20, 25, or 30 minutes). The dispersion obtained can be further lyophilized.

[0169] The nanoparticle compositions described herein (such a pharmaceutical composition) may have distinct characteristics for any one or more (in any combination) of the following: (1) the oligomeric status of the albumin associated with (such as in) the nanoparticles, such as the percentage of albumin monomers, dimers, and/or polymers (or trimers) of the albumin associated with (such as in) the nanoparticles; (2) the oligomeric status of the albumin associated with (such as in) the non-nanoparticle portion of the composition, such as the percentage of albumin monomers, dimers, and/or polymers (or trimers) of the albumin associated with (such as in) the non-nanoparticle portion of the composition; (3) the oligomeric status of the total albumin in the composition, such as the percentage of albumin monomers, dimers, and/or polymers (or trimers) of the total albumin in the composition; (4) the particle size profile of the nanoparticles, such as the average particle size, poly dispersity index, and/or size distribution; (5) the portion (e.g, weight percentage) of the nanoparticles that is albumin and/or the portion (e.g, weight percentage) of the nanoparticles that is rapamycin; (6) the weight ratio of the albumin to the rapamycin in the nanoparticles; (7) the weight ratio of the albumin to the rapamycin in the non-nanoparticle portion of the composition; (8) the weight ratio of the albumin to the rapamycin in the non-nanoparticle portion of the composition (9) the weight ratio of the total albumin to the total rapamycin in the composition; (10) the portion (e.g, weight percentage) of rapamycin that is in the nanoparticles (or the non-nanoparticle portion of the composition) compared to the total rapamycin in the composition; (11) the portion (e.g, weight percentage) of albumin that is in the non-nanoparticle portion (or in the nanoparticles) compared to the total albumin in the composition; (12) the concentration of albumin in the composition; (13) the concentration of albumin in the non-nanoparticle portion of the composition; (14) the concentration of albumin in the composition that is associated with (such as in) the nanoparticles; (15) the concentration of rapamycin in the composition; (16) the concentration of rapamycin in the non-nanoparticle portion of the composition; (17) the concentration of rapamycin in the composition that is associated with (such as in) the nanoparticles; (18) the osmolality of the composition; (19) the viscosity of the composition; (20) the pH of the composition; (21) the stability of the nanoparticles in the composition; (22) the amount of residual solvent in the composition; (23) the zeta potential of the nanoparticles in the composition; (24) the crystalline status of the rapamycin in the nanoparticles; (25) the particle morphology of the nanoparticles, such as the shape, sphericity, thickness of the coating, and/or surface-to-volume ratio; (26) the weight percentage of seco-rapamycin in the nanoparticles, as compared to the sum of seco-rapamycin and rapamycin, by weight; (27) the presence, percentage, or concentration of albumin stabilizer (such as sodium caprylate and/or N-acetyltryptophanate) in the composition; (28) the recovery of rapamycin following filtration; (29) in vitro release kinetics of the nanoparticles; (30) the portion of total rapamycin in the composition that is both in the non-nanoparticle portion of the composition and not bound to albumin; and/or (31) the weight percentage of seco-rapamycin in the composition, as compared to the sum of seco-rapamycin and rapamycin, by weight. In some embodiments, the oligomeric status (such as the percentage of albumin monomers, dimers, or polymers (or trimers)) of the nanoparticles, the non-nanoparticles portion, or the total composition is assessed by sizeexclusion chromatography using a saline mobile phase coupled with a multiple angle light scattering (MALS) detector). [0170] The nanoparticle compositions described herein (such a pharmaceutical composition) may have distinct characteristics for any one or more (in any combination) of the following: (1) the oligomeric status of the albumin associated with (such as in) the nanoparticles, such as the percentage of albumin monomers, dimers, oligomers, and/or polymers (other than oligomers) of the albumin associated with (such as in) the nanoparticles; (2) the oligomeric status of the albumin associated with (such as in) the non-nanoparticle portion of the composition, such as the percentage of albumin monomers, dimers, oligomers, and/or polymers (other than oligomers) of the albumin associated with (such as in) the non-nanoparticle portion of the composition; (3) the oligomeric status of the total albumin in the composition, such as the percentage of albumin monomers, dimers, oligomers, and/or polymers (other than oligomers) of the total albumin in the composition; (4) the particle size profile of the nanoparticles, such as the average particle size, poly dispersity index, and/or size distribution; (5) the portion (e.g, weight percentage) of the nanoparticles that is albumin and/or the portion (e.g, weight percentage) of the nanoparticles that is rapamycin; (6) the weight ratio of the albumin to the rapamycin in the nanoparticles; (7) the weight ratio of the albumin to the rapamycin in the non-nanoparticle portion of the composition; (8) the weight ratio of the albumin to the rapamycin in the non-nanoparticle portion of the composition (9) the weight ratio of the total albumin to the total rapamycin in the composition; (10) the portion (e.g, weight percentage) of rapamycin that is in the nanoparticles (or the non-nanoparticle portion of the composition) compared to the total rapamycin in the composition; (11) the portion (e.g, weight percentage) of albumin that is in the non-nanoparticle portion (or in the nanoparticles) compared to the total albumin in the composition; (12) the concentration of albumin in the composition; (13) the concentration of albumin in the non- nanoparticle portion of the composition; (14) the concentration of albumin in the composition that is associated with (such as in) the nanoparticles; (15) the concentration of rapamycin in the composition; (16) the concentration of rapamycin in the non-nanoparticle portion of the composition; (17) the concentration of rapamycin in the composition that is associated with (such as in) the nanoparticles; (18) the osmolality of the composition; (19) the viscosity of the composition; (20) the pH of the composition; (21) the stability of the nanoparticles in the composition; (22) the amount of residual solvent in the composition; (23) the zeta potential of the nanoparticles in the composition; (24) the crystalline status of the rapamycin in the nanoparticles; (25) the particle morphology of the nanoparticles, such as the shape, sphericity, thickness of the coating, and/or surface-to-volume ratio; (26) the weight percentage of seco- rapamycin in the nanoparticles, as compared to the sum of seco-rapamycin and rapamycin, by weight; (27) the presence, percentage, or concentration of albumin stabilizer (such as sodium caprylate and/or N-acetyltryptophanate) in the composition; (28) the recovery of rapamycin following filtration; (29) in vitro release kinetics of the nanoparticles; (30) the portion of total rapamycin in the composition that is both in the non-nanoparticle portion of the composition and not bound to albumin; and/or (31) the weight percentage of seco-rapamycin in the composition, as compared to the sum of seco-rapamycin and rapamycin, by weight. As used herein, “albumin oligomers” or “oligomeric albumin” refers to lower molecular weight polymeric albumin species associated with a UV-absorbance-based size-exclusion chromatography peak observed between a peak associated with albumin dimers and higher molecular weight polymeric albumin species. In some embodiments, the oligomeric status (such as the percentage of albumin monomers, dimers, oligomers, or polymers (other than oligomers)) of the nanoparticles, the non- nanoparticle portion, or the total composition is assessed by size-exclusion chromatography using a mobile phase containing an aqueous portion and a miscible organic portion (such as an aqueous buffer containing 7.5% methanol) coupled with a UV detector. In some embodiments, the percentage of albumin in the nanoparticle portion that is in the form of monomeric, dimeric, oligomeric, or polymeric albumin (other than oligomeric albumin) is determined by separating the nanoparticles from the non-nanoparticle portion, dissolving the nanoparticles, and subjecting the dissolved nanoparticles to size-exclusion chromatography. In some embodiments, the sizeexclusion chromatography uses a mobile phase containing an aqueous portion and a miscible organic portion (such as an aqueous buffer containing 7.5% methanol) coupled with a UV detector.

[0171] In some embodiments, the nanoparticle composition has one or more of the following distinct characteristics: (1) about 80% to about 95% (or as further provided herein) of the total albumin in the composition is in the form of monomeric albumin; (2) about 4% to about 15% (or as further provided herein) of the total albumin in the composition is in the form of dimeric albumin; (3) about 0.5% to about 5% (or as further provided herein) of the total albumin in the composition is in the form of polymeric albumin (or trimeric albumin); (4) the weight ratio of the total albumin to the total rapamycin in the composition is about 1:1 to about 10:1 (or as further provided herein); (5) about 90% or more (or as further provided herein) of the total rapamycin in the composition is in the nanoparticles; (6) about 90% or more (or as further provided herein) of the total albumin in the composition is in the non-nanoparticle portion of the nanoparticles; (7) the composition comprises tert-butanol at a concentration of less than about 10 pg/mL or less than about 10 ppm (or as further provided herein); (8) the composition comprises chloroform at a concentration of less than about 5 pg/mL or less than about 5 ppm (or as further provided herein); (9) the composition comprises an albumin stabilizer (such as sodium caprylate and/or N-acetyltryptophanate); (10) at least about 80% or more (or as further provided herein) of the rapamycin in the composition is recoverable after filtering the composition with a 0.2 micron filter; (11) the composition is stable for at least 24 hours; and/or (12) less than about 5% of the total rapamycin in the composition is both in the non-nanoparticle portion of the composition and unbound to albumin in the non-nanoparticle portion of the composition. In some embodiments, the nanoparticle composition may be a nanoparticle suspension, and the nanoparticle composition may have one or more of the following distinct characteristics (in addition to or in alternative to any one of the previously described district characteristics): (1) the concentration of albumin in the composition is about 30 mg/mL to about 100 mg/mL (or as further provided herein); (2) the concentration of rapamycin in the composition is about 1 mg/mL to about 15 mg/mL (or as further provided herein, such as about 1 mg/mL to about 7 mg/mL); (3) the osmolality of the composition is about 300 mOsm/kg to about 350 mOsm/kg (or as otherwise provided herein); (4) the viscosity of the composition is about 1.2 cP to about 1.5 cP (or as otherwise provided herein); and/or (5) the pH of the composition is about 6.0 to about 7.5 (or as otherwise provided herein).

[0172] In some embodiments, the nanoparticles of the composition have one or more of the following distinct characteristics: (1) about 70% to about 85% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin monomers; (2) about 9% to about 20% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin dimers; (3) about 5% to about 15% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin polymers (or albumin trimers); (4) the nanoparticles have a volume weighted mean particle size and/or Z-av erage particle size of about 200 nm or less (or as otherwise provided herein, such as between about 50 nm and about 200 nm); (5) the nanoparticles have a poly dispersity index of less than about 0.2 (or as otherwise provided herein, such as between about 0.03 and about 0.2); (6) the span of the particle size distribution ((Dv95- Dv5)/Dv50) is about 0.8 to about 1.2 (or as otherwise provided herein); (7) the nanoparticles are about 25% to about 45% albumin by weight (or as otherwise provided herein); (8) the nanoparticles are about 55% to about 75% rapamycin by weight (or as otherwise provided herein); (9) the weight ratio of albumin to rapamycin in the nanoparticles is about 1 : 1 to about 1:4 (or as otherwise provided herein); (10) the zeta potential of the nanoparticles in the composition is about -25 mV to about -50 mV (or as otherwise provided herein); (11) the nanoparticles have an amorphous morphology; (12) the rapamycin in the nanoparticles has an amorphous morphology; (13) the vinyl chain of the rapamycin in the nanoparticles interacts with the albumin in the nanoparticles; (14) at least a portion (such as at least 20%, or as otherwise provided herein) of the nanoparticles in the composition are non-spherical; (15) the nanoparticles comprise less than about 2.5% seco-rapamycin (or as otherwise provided herein, such as between about 0.2% and about 2.5%) compared to the sum of seco-rapamycin and rapamycin by weight; and/or (16) the composition comprises less than 3% seco-rapamycin (or as otherwise provided herein, such as between about 0.2% and about 2.5%) compared to the sum of seco-rapamycin and rapamycin by weight. In some embodiments, the nanoparticle composition may be a nanoparticle suspension, and in some embodiments the concentration of the albumin in the nanoparticle suspension that is in the nanoparticles is about 1.8 mg/mL to about 3 mg/mL (or as otherwise provided herein).

[0173] In some embodiments, the nanoparticles of the composition have one or more of the following distinct characteristics: (1) about 25% to about 50% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin monomers; (2) about 5% to about 16% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin dimers; (3) about 1% to about 4.5% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin oligomers; (4) about 42% to about 60% (or as otherwise provided herein) of the albumin in the nanoparticles is in the form of albumin polymers (other than oligomers); (5) the nanoparticles have a volume weighted mean particle size and/or Z-average particle size of about 200 nm or less (or as otherwise provided herein, such as between about 50 nm and about 200 nm); (6) the nanoparticles have a poly dispersity index of less than about 0.2 (or as otherwise provided herein, such as between about 0.03 and about 0.2); (7) the span of the particle size distribution ((Dv95-Dv5)/Dv50) is about 0.8 to about 1.2 (or as otherwise provided herein); (8) the nanoparticles are about 25% to about 45% albumin by weight (or as otherwise provided herein); (9) the nanoparticles are about 55% to about 75% rapamycin by weight (or as otherwise provided herein); (10) the weight ratio of albumin to rapamycin in the nanoparticles is about 1:1 to about 1:4 (or as otherwise provided herein); (11) the zeta potential of the nanoparticles in the composition is about -25 mV to about -50 mV (or as otherwise provided herein); (12) the nanoparticles have an amorphous morphology; (13) the rapamycin in the nanoparticles has an amorphous morphology; (14) the vinyl chain of the rapamycin in the nanoparticles interacts with the albumin in the nanoparticles; (15) at least a portion (such as at least 20%, or as otherwise provided herein) of the nanoparticles in the composition are non-spherical; (16) the nanoparticles comprise less than about 2.5% seco-rapamycin (or as otherwise provided herein, such as between about 0.2% and about 2.5%) compared to the sum of seco-rapamycin and rapamycin by weight; and/or (17) the composition comprises less than about 3% seco-rapamycin (or as otherwise provided herein, such as between about 0.2% and about 3%) compared to the sum of seco-rapamycin and rapamycin, by weight. In some embodiments, the nanoparticle composition may be a nanoparticle suspension, and in some embodiments the concentration of the albumin in the nanoparticle suspension that is in the nanoparticles is about 1.8 mg/mL to about 3 mg/mL (or as otherwise provided herein).

[0174] In some embodiments, the non-nanoparticle portion of the composition has one or more of the following distinct characteristics: (1) about 80% to about 95% (or as otherwise provided herein) of the albumin in the non-nanoparticle portion of the composition is in the form of albumin monomers; (2) about 5% to about 14% (or as otherwise provided herein) of the albumin in the non-nanoparticle portion of the composition is in the form of albumin dimers; and/or (3) about 1% to about 5% (or as otherwise provided herein) of the albumin in the non- nanoparticle portion of the composition is in the form of albumin polymers (or albumin trimers). In some embodiments, the nanoparticle composition may be a nanoparticle suspension, and the non-nanoparticle portion of the nanoparticle suspension may have one or more of the following distinct characteristics (in addition to or in alternative to any one of the previously described district characteristics): (1) the concentration of albumin in the non-nanoparticle portion of the composition is between about 30 mg/mL and about 100 mg/mL (or as otherwise provided herein); and/or (2) the concentration of rapamycin in the non-nanoparticle portion is about 20 pg/mL to about 55 pg/mL (or as otherwise provided herein).

[0175] In some embodiments, the non-nanoparticle portion of the composition has one or more of the following distinct characteristics: (1) about 80% to about 95% (or as otherwise provided herein) of the albumin in the non-nanoparticle portion of the composition is in the form of albumin monomers; (2) about 5% to about 16% (or as otherwise provided herein) of the albumin in the non-nanoparticle portion of the composition is in the form of albumin dimers; about 0.5% to about 4% (or as otherwise provided herein) of the albumin in the non-nanoparticle portion of the composition is in the form of albumin oligomers; and/or (4) about 0.5% to about 3% (or as otherwise provided herein) of the albumin in the non-nanoparticle portion of the composition is in the form of albumin polymers (other than oligomers). In some embodiments, the nanoparticle composition may be a nanoparticle suspension, and the non-nanoparticle portion of the nanoparticle suspension may have one or more of the following distinct characteristics (in addition to or in alternative to any one of the previously described district characteristics): (1) the concentration of albumin in the non-nanoparticle portion of the composition is between about 30 mg/mL and about 100 mg/mL (or as otherwise provided herein); and/or (2) the concentration of rapamycin in the non-nanoparticle portion is about 20 pg/mL to about 55 pg/mL (or as otherwise provided herein).

[0176] The compositions (such as pharmaceutical compositions) described herein can be in liquid (e.g, as a nanoparticle suspension) or powder forms. For example, in some embodiments, the composition is a liquid nanoparticle suspension (for example prior to lyophilization). In some embodiments, the composition is a reconstituted suspension (e.g, in an aqueous solution such as a saline solution). In some embodiments, the composition is dried, such as lyophilized. In some embodiments, the composition is sterile. In some embodiments, the composition is contained in a sealed container, such as a sealed vial (e.g, a glass vial) or sealed bag.

[0177] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin. In some embodiments, about 0.5% to about 5% of the albumin in the non-nanoparticle portion or the total albumin in the nanoparticle composition is in the form of polymeric albumin (or trimeric albumin). In some embodiments, about 4% to about 14% of the albumin in the non-nanoparticle portion or the total albumin in the nanoparticle composition is in the form of dimeric albumin. In some embodiments, about 80% to about 95% of the albumin in the non-nanoparticle portion or the total albumin in the nanoparticle composition is in the form of monomeric albumin. In some embodiments, the weight ratio of the albumin to the rapamycin in the composition is about 1 : 1 to about 10:1. In some embodiments, about 90% or more of the albumin in the composition is in the non-nanoparticle portion. In some embodiments, about 90% or more of the rapamycin in the composition is in the nanoparticles. In some embodiments, the concentration of albumin in the nanoparticle composition that is in the non-nanoparticle portion or the concentration of total albumin in the nanoparticle composition is about 30 mg/mL to about 100 mg/mL. In some embodiments, the osmolality of the composition is about 300 mOsm/kg to about 350 mOsm/kg. In some embodiments, the viscosity of the composition is about 1.2 cP to about 1.5 cP. In some embodiments, the pH of the composition is about 6.0 to about 7.5. In some embodiments, the composition is stable at 4 °C and/or 25 °C for at least 24 hours. In some embodiments, the rapamycin in the nanoparticles has an amorphous morphology. In some embodiment, the nanoparticle composition is a nanoparticle suspension. In some embodiments, the nanoparticle composition is a dried composition. In some embodiments, the nanoparticle composition is sterile, for example by filtration. In some embodiments, the nanoparticle composition is contained within a sealed container, such as a sealed vial or a sealed bag. In some embodiments, the nanoparticle composition comprises less than 10 pg/mL tert-butanol and/or comprises less than 5 pg/mL chloroform.

[0178] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin; and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0179] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 25% to about 50% of the albumin in the nanoparticles is in the form of monomeric albumin; and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0180] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0181] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 25% to about 50% of the albumin in the nanoparticles is in the form of polymeric albumin (other than oligomeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0182] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin; and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0183] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 5% to about 16% of the albumin in the nanoparticles is in the form of dimeric albumin; and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0184] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0185] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 25% to about 50% of the albumin in the nanoparticles is in the form of monomeric albumin, about 1% to about 4.5% of the albumin in the nanoparticles is in the form of oligomeric albumin, about 5% to about 16% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 25% to about 50% of the albumin in the nanoparticles is in the form of polymeric albumin (other than oligomeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0186] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm), comprising rapamycin and albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0187] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm), comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0188] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm), comprising about 55% to about 65% (by weight) rapamycin and about 25% to about 45% (by weight) albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0189] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm), comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein the albumin comprises about 25% to about 45% of the nanoparticles by weight and the rapamycin comprises about 55% to about 75% of the nanoparticles by weight, wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0190] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm), comprising about 55% to about 75% (by weight) rapamycin and about 25% to about 45% (by weight) albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL).

[0191] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm), comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein the albumin comprises about 25% to about 45% of the nanoparticles by weight and the rapamycin comprises about 55% to about 75% of the nanoparticles by weight, wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL).

[0192] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm) and a zeta potential of about -25 mV to about -50 mV, comprising about 55% to about 75% (by weight) rapamycin and about 25% to about 45% (by weight) albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL).

[0193] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm) and a zeta potential of about -25 mV to about -50 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein the albumin comprises about 25% to about 45% of the nanoparticles by weight and the rapamycin comprises about 55% to about 75% of the nanoparticles by weight, wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL).

[0194] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm) and a zeta potential of about -25 mV to about -50 mV, comprising about 55% to about 75% (by weight) rapamycin and about 25% to about 45% (by weight) albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL); and wherein about 3% or less of the rapamycin in the nanoparticle composition is free rapamycin.

[0195] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm) and a zeta potential of about -25 mV to about -50 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein the albumin comprises about 25% to about 45% of the nanoparticles by weight and the rapamycin comprises about 55% to about 75% of the nanoparticles by weight, wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL); and wherein about 3% or less of the rapamycin in the nanoparticle composition is free rapamycin. [0196] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm) and a zeta potential of about -25 mV to about -50 mV, comprising about 55% to about 75% (by weight) rapamycin and about 25% to about 45% (by weight) albumin (such as human albumin), wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL); and wherein the sum of seco-rapamycin and rapamycin in the nanoparticles is less than 3% (such as about 0.2% to about 3%) seco-rapamycin, by weight. In some embodiments, the sum of seco-rapamycin and rapamycin in the composition is less than 3% (such as about 0.2% to about 3%) seco-rapamycin, by weight.

[0197] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 200 nm or less (such as about 50 nm to about 200 nm) and a zeta potential of about -25 mV to about -50 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein the albumin comprises about 25% to about 45% of the nanoparticles by weight and the rapamycin comprises about 55% to about 75% of the nanoparticles by weight, wherein about 70% to about 85% of the albumin in the nanoparticles is in the form of monomeric albumin, about 9% to about 20% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 5% to about 15% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL); and wherein the sum of seco-rapamycin and rapamycin in the nanoparticles is less than 3% (such as about 0.2% to about 3%) seco- rapamycin, by weight. In some embodiments, the seco-rapamycin is less than 3% (such as about 0.2% to about 3%) of the sum of seco-rapamycin and rapamycin in the composition.

[0198] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin; and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin. In some embodiments, about 1.5% to about 3% of the albumin in the non-nanoparticle portion or the total albumin in the nanoparticle composition is in the form of polymeric albumin (or trimeric albumin). In some embodiments, about 7% to about 11% of the albumin in the non-nanoparticle portion in the nanoparticle composition is in the form of dimeric albumin. In some embodiments, about 7% to about 11% of the total albumin in the nanoparticle composition is in the form of dimeric albumin. In some embodiments, about 83% to about 92% of the albumin in the non-nanoparticle portion or the total albumin in the nanoparticle composition is in the form of monomeric albumin. In some embodiments, the weight ratio of the albumin to the rapamycin in the composition is about 7:1 to about 9:1. In some embodiments, about 95% or more of the albumin in the composition is in the non-nanoparticle portion. In some embodiments, about 98% to about 99.5% of the rapamycin in the composition is in the nanoparticles. In some embodiments, the concentration of albumin in the nanoparticle composition that is in the non-nanoparticle portion or the concentration of total albumin in the nanoparticle composition is about 35 mg/mL to about 45 mg/mL.

[0199] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0200] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin; and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0201] In some embodiments, the nanoparticle composition comprises (a) nanoparticles comprising rapamycin and albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11 % of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin. [0202] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm, comprising rapamycin and albumin (such as human albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0203] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm, comprising rapamycin and albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0204] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0205] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a zeta potential of about -33 mV to about -39 mV, comprising rapamycin and albumin (such as human albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0206] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a zeta potential of about -33 mV to about -39 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin; and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin

[0207] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a zeta potential of about -33 mV to about -39 mV, comprising rapamycin and albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0208] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a zeta potential of about -33 mV to about -39 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0209] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about -33 mV to about -39 mV, comprising rapamycin and albumin (such as human albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0210] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about -33 mV to about -39 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin; and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin

[0211] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about -33 mV to about -39 mV, comprising rapamycin and albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0212] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about -33 mV to about -39 mV, comprising a coating comprising albumin (such as human albumin) and a core comprising rapamycin, wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non- nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0213] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm, comprising about 62% to about 68% (by weight) rapamycin and about 32% to about 38% (by weight) albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin.

[0214] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm, comprising about 62% to about 68% (by weight) rapamycin and about 32% to about 38% (by weight) albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL).

[0215] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about -33 mV to about -39 mV, comprising about 62% to about 68% (by weight) rapamycin and about 32% to about 38% (by weight) albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL).

[0216] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about -33 mV to about -39 mV, comprising about 62% to about 68% (by weight) rapamycin and about 32% to about 38% (by weight) albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL); and wherein about 1% or less of the rapamycin in the nanoparticle composition is free rapamycin.

[0217] In some embodiments, the nanoparticle composition comprises (a) nanoparticles having a Z-average particle size of about 85 nm to about 95 nm and a zeta potential of about of about -33 mV to about -39 mV, comprising about 62% to about 68% (by weight) rapamycin and about 32% to about 38% (by weight) albumin (such as human albumin), wherein about 74% to about 80% of the albumin in the nanoparticles is in the form of monomeric albumin, about 12% to about 17% of the albumin in the nanoparticles is in the form of dimeric albumin, and about 7% to about 11% of the albumin in the nanoparticles is in the form of polymeric albumin (or trimeric albumin); and (b) a non-nanoparticle portion comprising albumin (such as human albumin) and rapamycin; wherein the concentration of the rapamycin in the nanoparticle composition is about 1 mg/mL to about 100 mg/mL (such as about 1 mg/mL to about 15 mg/mL); and wherein the sum of seco-rapamycin and rapamycin in the nanoparticles is less than 1% (such as about 0.5% to about 1%) seco-rapamycin, by weight. In some embodiments, seco- rapamycin is greater than about 0.2% (such as about 0.2% to about 3%) of the sum of seco- rapamycin and rapamycin in the composition.

[0218] Also provided herein are commercial batches of the nanoparticle compositions (such as the pharmaceutical compositions) for use of any one of the treatment methods described here. “Commercial batch” as used herein refers to a batch size that is at least about 20 grams (by mass of rapamycin). Commercial batches are produced at a larger scale than experimental or bench- scale batches. The increased scale is associated with longer production times, including longer steps (such as evaporation steps) or longer hold times between steps.

[0219] In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered subcutaneously. In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered intravenously. In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered at a dose between about 1 mg/m² and about 150 mg/m², between about 5 mg/m² and about 75 mg/m², e.g, via intravenous infusion. In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered at a dose of about any one of 5, 7.5, 10, 15, 30, 56, 75 or 100 mg/m², e.g, via intravenous infusion. In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered to the individual having cancer in one or more 21 -day cycles (e.g, three-week cycles). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered to the individual once during each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered during Week 1, Week 2, or Week 3 during each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered on Day 1, Day 8, or Day 15 of each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered to the individual twice during each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered during Week 1 and Week 2 during each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered during Week 2 and Week 3 during each 21-day cycle (e.g, three- week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered during Week 1 and Week 3 during each 21 -day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered on Day 1 and Day 8 of each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered on Day 1 and Day 15 of each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered on Day 8 and Day 15 of each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered to the individual three times during each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered during Week 1, Week 2, and Week 3 during each 21-day cycle (e.g, three-week cycle). In some embodiments, the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is administered on Day 1, Day 8, and Day 15 of each 21-day cycle (e.g, three- week cycle). In some embodiments, the dosage of the mTOR inhibitor nanoparticle composition (e.g, a sirolimus/albumin nanoparticle composition, such as FYARRO™) is modified (e.g, if the individual experiences one or more adverse effects). Details regarding dosage modification for FYARRO™ and circumstances under which dosage modifications are made are detailed at www(dot)accessdata(dot)fda(dot)gov/drugsatfda_docs/label/2021/2133121bl.pdf.

V. Articles of Manufacture and Kits

[0220] In some embodiments, there is provided an article of manufacture containing materials useful for the treatment according to the methods provided herein, such as for undifferentiated pleomorphic sarcoma (including undifferentiated pleomorphic sarcoma having a PTEN loss and/ or a TSC2 mutation) or leiomyosarcoma (including estrogen receptor-positive leiomyosarcoma), the article of manufacture, such as a medicament or medicament combination, comprising an mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition, e.g, na/i-sirohmus) and an anti-PD-1 antibody (e.g, nivolumab). The article of manufacture can comprise a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, etc. The containers may be formed from a variety of materials such as glass or plastic. Generally, the container holds a composition which is effective for treating a disease or disorder described herein, and may have a sterile access port (for example the container may be an intravenous solution bag or a vial having a stopper pierceable by a hypodermic injection needle). At least one active agent in the composition is a) a nanoparticle formulation of an mTOR inhibitor; or b) an anti-PD-1 antibody. The label or package insert indicates that the composition is used for treating the particular condition in an individual. The label or package insert will further comprise instructions for administering the composition to the individual. Articles of manufacture and kits comprising combination therapies described herein are also contemplated.

[0221] Package insert refers to instructions customarily included in commercial packages of therapeutic products that contain information about the indications, usage, dosage, administration, contraindications and/or warnings concerning the use of such therapeutic products. In some embodiments, the package insert indicates that the composition is used for treating a solid tumor (such as bladder cancer, renal cell carcinoma, or melanoma).

[0222] Additionally, the article of manufacture may further comprise a second container comprising a pharmaceutically-acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer's solution and dextrose solution. It may further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles, and syringes.

[0223] Kits are also provided that are useful for various purposes, e.g., for treatment of an undifferentiated pleomorphic sarcoma (including undifferentiated pleomorphic sarcoma having a PTEN loss and/ or a TSC2 mutation) or leiomyosarcoma (including estrogen receptor-positive leiomyosarcoma). Kits of the invention include one or more containers comprising an mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) (or unit dosage form and/or article of manufacture), and in some embodiments, further comprise an anti- PD-1 antibody (such as described herein) and/or instructions for use in accordance with any of the methods described herein. The kit may further comprise a description of selection of individuals suitable for treatment. Instructions supplied in the kits of the invention are typically written instructions on a label or package insert (e.g, a paper sheet included in the kit), but machine-readable instructions (e.g., instructions carried on a magnetic or optical storage disk) are also acceptable. [0224] The kits of the invention are in suitable packaging. Suitable packaging includes, but is not limited to, vials, bottles, jars, flexible packaging (e.g., sealed Mylar or plastic bags), and the like. Kits may optionally provide additional components such as buffers and interpretative information. The present application thus also provides articles of manufacture, which include vials (such as sealed vials), bottles, jars, flexible packaging, and the like.

[0225] The instructions relating to the use of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody generally include information as to dosage, dosing schedule, and route of administration for the intended treatment. The containers may be unit doses, bulk packages (e.g., multi-dose packages) or subunit doses. For example, kits may be provided that contain sufficient dosages of an mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and an anti-PD-1 antibody as disclosed herein to provide effective treatment of an individual for an extended period, such as any of a week, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks, 3 months, 4 months, 5 months, 7 months, 8 months, 9 months, or more. Kits may also include multiple unit doses of the mTOR inhibitor nanoparticle composition (such as sirolimus/albumin nanoparticle composition) and the anti-PD-1 antibody and instructions for use, packaged in quantities sufficient for storage and use in pharmacies, for example, hospital pharmacies and compounding pharmacies.

[0226] Those skilled in the art will recognize that several embodiments are possible within the scope and spirit of this invention. The invention will now be described in greater detail by reference to the following non-limiting examples. The following examples further illustrate the invention but, of course, should not be construed as in any way limiting its scope.

EXAMPLES

Example 1

[0227] This example demonstrates a phase I/II investigation study administering nab- sirolimus and nivolumab to individual having a cancer. Patients and Methods

Study design

[0228] This was an open-label, single-center, dose-finding phase IB study using a fixed dose of nivolumab, an anti-PD-1 antibody, and escalating doses of m/i-sirolimus given intravenously. Patients were enrolled from September 2017 to July 2021. The study was conducted in accordance with the Declaration of Helsinki and approved by the Western Institutional Review Board (Protocol Code 20151429 on September 8, 2017) for studies involving humans.

[0229] The primary endpoint was determination of the MTD of m/i-sirolimus when combined with nivolumab. Secondary endpoints included the disease control rate [DCR: complete response (CR) plus partial response (PR) plus stable disease] as determined by local radiological assessment using Response Evaluation Criteria in Solid Tumors (RECIST) vl.l; the objective response rate (CR plus PR), and progression-free (PFS), and overall (OS) survival. Secondary endpoints also included determination of the median PFS and median OS.

[0230] The exploratory endpoint evaluated the correlation between response based on immune-related response criteria (irRECIST) and that based on RECIST vl.l using Pearson’s coefficient of correlation.

Dose-escalation phase I part of the study

[0231] The study employed the standard “cohort of three” design, wherein three patients were treated at each dose level with expansion to six patients per cohort when dose-limiting toxicity (DLT) was observed in one out of the three initially enrolled patients at each dose level. If no DLT occurred after two doses, escalation to the next dose level was permitted. The MTD was defined as the highest safely tolerated dose at which no more than one patient experienced DLT, with the next higher dose level having at least two patients who experienced DLT. Patients in the dose-escalation study were able to continue treatment at their designated dose levels for up to 18 3-week cycles or until significant disease progression or unacceptable toxicity occurred. No intra-patient dose escalation took place.

[0232] The dose of nivolumab given was 3 mg/kg intravenously over 30 min q 3 weeks (day 1 of every 21 -day cycle) beginning in cycle 1. /6-sirolimus was administered on days 8 and 15 beginning in cycle 2. Nivolumab was given first based on preclinical findings of increased efficacy when administered before wo/i-sirolimus. Escalating doses of wo/i-sirolimus were given intravenously over 30 min for 2 out of every 3 weeks starting from 56 mg/m² (n=3-6); 75 mg/m² (n=3-6) and 100 mg/m² (n=3-6).

[0233] Note that DLT included colitis, hepatitis, or pneumonitis of grade 3 or more; any grade 1-2 colitis, hepatitis or pneumonitis that recurred, worsened or persisted with oral steroids longer than 14 days; symptoms of adrenal crisis; any grade 4 hematological toxicity; or any non- hematological toxicity of grade 3 or more, according to the National Cancer Institute Common Terminology Criteria for Adverse Events (CTCAE) v4.03 (14). When DLT developed in more than one patient at 56 mg/m², the dose of wo/i-sirolimus was de-escalated to 45 mg/m² and to 30 mg/m² when a DLT developed at 45 mg/m².

Expansion phase IB/ II part of study

[0234] Following dose escalation and identification of the maximum tolerated dose (MTD), patients continued to receive wo/i-sirolimus and nivolumab at the MTD, namely, 100 mg/m2 and 3 mg/kg, respectively. Additional patients added to the study also received m/i-sirolimus and nivolumab at the MTD. The purposes of the expansion phase was to assess overall safety and efficacy in a great number of patients. Patients in the expansion phase of the study continued treatment for up to 18 3-week treatment cycles or until significant disease progression or unacceptable toxicity occurred. The study was conducted in compliance with the International Conference on Harmonisation Good Clinical Practices.

[0235] After three or more treatment cycles, the principal investigator recommended surgical debulking, complete surgical removal or a biopsy. If residual disease was present either by histopathological examination or by computerized tomography (CT) scan/magnetic resonance imaging (MRI), repeat treatment cycles were given 2-4 weeks after surgery, when the surgical incision had healed, and if the patient had less than grade 1 toxicity.

[0236] Treatment was continued in the presence of increased tumor size by CT scan or MRI, indicating progressive disease by RECIST vl.l when there were no signs or symptoms indicating unequivocal progression, no worsening of Eastern Cooperative Oncology Group (ECOG) score attributable to progressive disease, no tumor growth at critical sites that was life- threatening, when the patient had signed an informed consent form that they were aware of alternative therapies but wished to defer these therapies, or when there was clinical benefit, as determined by the investigator.

Study population

[0237] The study was designed to enroll up to 40-50 patients. Patients who failed to become evaluable for the secondary endpoint with a follow-up CT/MRI were replaced. Tumors with histology of Ewing sarcoma, PEComa, epithelioid sarcoma, desmoid tumor, chordoma, nonsmall-cell lung cancer, small-cell lung cancer, urothelial carcinoma, melanoma, renal cell carcinoma, squamous cell carcinoma of head and neck, hepatocellular carcinoma, classical Hodgkin’s lymphoma, microsatellite instability/mismatch repair deficiency (MSI-H/dMMR) metastatic colorectal cancer, and tumors with genetic mutations sensitive to mTOR inhibitors, were confirmed locally by the institution prior to enrollment.

Patient eligibility

[0238] Patients were eligible for inclusion in this study only when all of the following criteria were met: a. Histologically confirmed diagnosis of Ewing sarcoma, PEComa, epithelioid sarcoma, desmoid tumor, chordoma, non-small cell lung cancer, small cell lung cancer, urothelial carcinoma, melanoma, renal cell carcinoma, squamous cell carcinoma of head and neck, hepatocellular carcinoma, classical Hodgkin’s lymphoma, MSIH/dMMR metastatic colorectal cancer, and tumors with genetic mutations sensitive to mTOR inhibitors that were either metastatic or locally advanced and for which surgery was not a recommended option; b. One or more measurable target lesions by CT scan or MRI, measurable disease by RECIST vl.l, confirmed by investigator; c. Previous treatment completed after 5 half-lives or >28 days prior to enrollment, whichever was shorter; d. >40 kg for those aged 12-17 years, or 18 years or older, with Eastern Cooperative Oncology Group (ECOG) performance status 0 or 1; e. Blood chemistry levels at screening (obtained <14 days prior to enrollment, local laboratory): Total bilirubin <1.5*upper limit of normal (ULN) mg/dl, aspartate aminotransferase/alanine aminotransferase <2.5*ULN (<5*ULN if attributable to liver metastases), alkaline phosphatase <3 /ULN (or >3xULN when due to bone metastases), serum creatinine <1.5xULN; f. Blood counts at screening (obtained <14 days prior to enrollment, local laboratory): Absolute neutrophil count >1 ,5x 10⁹/l, platelet count >100,000/mm3 (100*10⁹/l), hemoglobin >9 g/dl, serum triglyceride <300 mg/dl, serum cholesterol <350 mg/dl; g. Males and females of child-bearing age had to agree to use effective contraception 28 days before treatment with study drugs, while on study, and have a negative serum pregnancy test (human chorionic gonadotrophin) result at screening and to agree to have pregnancy testing during the study period, and after the end of treatment with study drugs. A second form of birth control was required even in the case of tubal ligation. Male patients were required to practice abstinence or agree to use a condom during sexual contact with a pregnant female or a female of childbearing potential while participating in the study. A second form of birth control was required even in the case of successful vasectomy; h. Life expectancy of >3 months, as determined by the investigator; i. Ability to understand and provide signed informed consent; j. Willingness and ability to comply with scheduled visits, laboratory tests, and other study procedures.

[0239] Patients were not eligible for inclusion in this study when any of the following criteria applied: a. Known active uncontrolled or symptomatic central nervous system (CNS) metastases. For patients with controlled and asymptomatic CNS metastases, prior treatment for CNS metastases must have been completed >28 days (including radiotherapy/surgery) prior to the start of treatment in this study and should not be receiving chronic corticosteroid therapy for the CNS metastases; b. Active gastrointestinal bleeding; c. Uncontrolled pre-existing thyroid abnormality; d. Uncontrolled serious medical or psychiatric illness; e. ‘Currently active’ second malignancy, with the exception of patients with non-melanoma skin cancer, carcinoma in situ of the cervix, resected incidental prostate cancer (staged pT2 with Gleason Score <6 and postoperative prostate-specific antigen <0.5 ng/ml), or other adequately treated carcinoma in-situ; f. Patients were not considered to have a currently active malignancy when they had completed therapy and were free of disease for >1 year); g. Liver-directed therapy within 2 months of enrollment. Prior treatment with radiotherapy [including radio-labeled spheres or CyberKnife, hepatic arterial embolization (with or without chemotherapy) or cryotherapy/ablation] was allowed when these therapies did not affect the areas of measurable disease being used for this protocol; h. Infection requiring systemic anti-infective treatment that was completed <14 days prior to enrollment (with the exception of uncomplicated urinary tract infection or upper respiratory tract infection); i. Uncontrolled diabetes mellitus as defined by HbAlc >8% despite adequate therapy; j. Unstable coronary artery disease or myocardial infarction during the preceding 6 months; k. Receipt of any concomitant antitumor therapy; l. History of interstitial lung disease, pneumonitis, or pulmonary hypertension; m. Use of strong inhibitors and inducers of cytochrome P450 3A4 (CYP3A4) in the 14 days prior to receiving the first dose of «a6-sirolimus. Additionally, use of any known CYP3A4 substrate with narrow therapeutic window (such as fentanyl, alfentanil, astemizole, cisapride, dihydroergotamine, pimozide, quinidine, terfanide) within the 14 days prior to receiving the first dose of «a6-sirolimus; n. Active hepatitis B or hepatitis C; o. Non-oncology vaccine therapy used for prevention of infectious disease within 4 weeks of trial enrollment; p. Autoimmune disease including rheumatoid arthritis, systemic progressive sclerosis (scleroderma), systemic lupus erythematosus, autoimmune vasculitis and motor neuropathy considered to be of autoimmune origin (e.g. Guillain-Barre syndrome); q. Systemic immunosuppression, including human immunodeficiency virus positive status with or without acquired immune deficiency syndrome; r. Skin rash (psoriasis, eczema) affecting >25% body surface area; s. Inflammatory bowel disease (Crohn’s or ulcerative colitis); t. Ongoing or uncontrolled diarrhea within 4 weeks of trial enrollment; u. Recent history of acute diverticulitis, intraabdominal abscess or gastrointestinal obstruction within 6 months of trial enrollment (known risk factors for bowel perforation); v. Current, active or previous history of heavy alcohol abuse; w. Pituitary endocrinopathy; a. Adrenal insufficiency or excess.

Length of study

[0240] The study was designed to take approximately 32 months from the first patient enrolled to last patient follow-up, including approximately 24 months of enrollment, an estimated 6 months of treatment (or until treatment was no longer tolerated) and an end of treatment visit at 4 weeks (±7 days) after the last treatment.

[0241] The study was designed to end at either the date of the last visit of the last patient, or the date of receipt of the last data point from the last patient that was required for primary, secondary, or exploratory analysis, as pre-specified in the protocol.

[0242] The study treatment was designed to end for a patient as of the date of the last dose of nivolumab or m6-sirolimus. End of treatment visit for a patient occurred when safety assessments and procedures were performed after the last treatment, which had to be at least 4 weeks (±7 days) after the last dose of nivolumab or m6-sirolimus.

[0243] The study was designed with a follow-up period based on the on-study time period after the end of treatment visit. All patients that discontinued the study drug and did not withdrawal full consent to participate in the study continued in the follow-up phase for survival and initiation of anticancer therapy. Follow-up took place approximately every 12 weeks (±3 weeks), until death, withdrawal of consent, or study closure, whichever was the earliest. This evaluation may have been made by record review with/without telephone contact.

Statistical considerations

[0244] Safety analyses: Demographic and baseline information (e.g., extent of prior therapy) for study patients was tabulated. The number of patients studied for the dose-escalation part of the study was nine, and was 22 for the dose-expansion part.

[0245] For the phase I part of the study, the following information was reported for all adverse events observed: Dose level, type (organ affected or laboratory determination, such as absolute neutrophil count), severity [by CTCAE version 4.03 and most extreme abnormal values for laboratory determinations] and relatedness to study treatment. For each dose, the number and percentage of patients experiencing any grade 3, 4, or 5 adverse event were reported, as well as the number and percentage of patients who experienced selected, specific types of adverse event. In addition, the DLTs were summarized by dose level and the MTD was determined.

[0246] For the phase I part of the study, the entire treated population (full analysis set) was that analyzed for all safety analyses. Summary tables provided herein include the number and percentage of patients with adverse events, serious adverse events, fatal adverse events and other adverse events of interest. Safety was analyzed in all patient groups together (metastatic and locally advanced disease). Frequency tables, graphs, and summary statistics were used to analyze outcome data.

[0247] The incidence of all treatment-emergent adverse events were tabulated. Tables of fatal adverse events, serious adverse events, treatment-related adverse events, and adverse events leading to withdrawal from the study drugs were also collected. [0248] For nivolumab and m/j-sirolimus exposure, summary statistics were used for the total number of doses, average dose administered, and duration of each treatment.

[0249] Efficacy analyses'. The disease control rate (CR, PR, SD), objective response rate (ORR), progression free survival (PFS) and overall survival (OS) were assessed by local radiological assessment using RECIST vl.l and irRECIST.

[0250] The focus of the study was to estimate the DCR in patients treated with nivolumab and m/j-sirolimus. Patients who disease progressed before receiving /7o6-sirolimus were replaced and were not included in the statistical analysis. The number and percentage of patients achieving response was summarized.

[0251] . nalysis of other efficacy endpoints, median PFS, and median OS were assessed for all tumor subtypes together.

[0252] The number of patients in each category was relatively small; therefore, median PFS and median OS for these patients were summarized by descriptive statistics.

[0253] Exploratory analysis: Response based on RECIST vl.l was correlated with that based on irRECIST (10) using Pearson’s coefficient of correlation.

Results

Demographic information

[0254] A total of 34 patients were included in this study. Table 1 shows the patients enrolled according to sex and race. There were 19 males and 15 females, of whom most (22/34) were White not of Hispanic origin. Table 2 shows the patients enrolled according to age group and sex. The majority (56%) were young adults between 18 and 39 years of age. Table 3 shows the histological tumor subtypes of patients enrolled in the study. Most patients (15/34) had osteosarcoma or Ewing sarcoma.

Table 1. Patients enrolled in the SOC-1701 study, according to race and sex.

Table 2. Patients enrolled in the SOC-1701 study, according to age group and sex.

Table 3. Histological cancer subtypes of patients enrolled in the SOC-1701 study.

Summary of safety analysis

[0255] Thirty-one out of 34 patients who received at least one dose of «a6-sirolimus were evaluable for the safety analysis. [0256] In the phase I part of the study, no DLT occurred. The phase IB part of the study used the standard dose of 3 mg/kg nivolumab every 3 weeks and the MTD dose of 100 mg/m² nab- sirolimus on days 8 and 15 of each cycle.

AEs by body system

[0257] Table 4 shows AEs by body system observed in the phase I part and expanded phase IB part of the study, toxicity grade, and attribution to treatment regimen. 25 out of 31 (80.6%) patients who received at least one dose of /7o6-sirolimus had at least one treatment-related AE. The number of patients in the dose 3+ expanded phase IB category who had at least one treatment-related AE was 20 out of 25 (80%). Grade 3 adverse events considered to be related to nivolumab included increased thyroid-stimulating hormone (3.2%), while grade 3 adverse events considered to be related to «a6-sirolimus included thrombocytopenia (9.7%), oral mucositis (3.2%), increased thyroid-stimulating hormone (3.2%), acute dehydration (3.2%), hypertriglyceridemia (3.2%) and hypophosphatemia (3.2%).

Attorney Reference: 63877-20227.40

Table 4. Adverse events related to study therapy, by grade.

sf-5924244

SUBSTITUTE SHEET (RULE 26) Attorney Reference: 63877-20227.40

Summary of efficacy analysis

[0258] Table 5 shows the best overall response, PFS and OS data of patients enrolled in the study. sf-5924244

SUBSTITUTE SHEET (RULE 26) Table 5. Efficacy analysis of best overall response, progression-free survival (PFS) and overall survival (OS) (n=31/34).

(ir)RECIST: (Immune-related) Response Evaluation Criteria for Solid Tumors; PD: Progressive disease; PR: Partial response; SD: stable disease.

[0259] Thirty-one out of 34 patients who received at least one treatment cycle and had a follow-up CT scan were evaluable for best overall response, PFS and OS. Three patients did not receive na6-sirolimus.

[0260] The best responders (i.e. those with PR) were patients with undifferentiated pleomorphic sarcoma whose tumors had loss of phosphatase and tensin homolog (PTEN) and a tuberous sclerosis complex 2 (TSC2) mutation, and leiomyosarcoma whose tumor was estrogen receptor-positive.

[0261] As of the data cut-off date, two patients (one with TSC2 and another with TP 53 mutation) were still alive after 113 and 104 weeks, two patients with A4ES7/I-WTI fusion were still alive after 104 and 106 weeks, and one patient with /V wCA was still alive after 98 weeks. Summary of exploratory endpoint analysis.

[0262] Correlation between RECISTvl. 1 and irRECIST: Response was correlated according to RECISTvl.1 and irRECIST using Pearson correlation. The Pearson correlation coefficient was 1.00 (/?<0.0001), which represented a perfect correlation.

[0263] Deaths: Table 6 shows the list of deaths, Table 7 shows the list of patients who withdrew due to an adverse event, and Table 8 shows patients who were lost to follow-up, therefore their survival status is not known. As of the data cut-off date, 23 patients died from disease progression, eight with disease progression have been lost to follow-up and five patients were still alive. No patient died within 28 days of receiving /7o6-sirolimus.

Table 6. Deaths during the study¹.

All deaths were due to disease progression, not attributable to a study drug.

Table 7. Patients who withdrew from the study due to adverse event.

Table 8. List of patients who were lost to follow-up.

[0264] Table 9 shows the sarcoma histology, mutation(s) if known, number of cycles treated, best overall response rate (BORR), progression free survival (PFS), and overall survival (OS) of each patient.

[0265] Best responders (PR) were patients with UPS who also had PTEN loss and TSC2 mutation, and LMS who was ER+. Eight patients had a PFS of greater than 17.8 weeks (4.1 months = median PFS of trabectedin alone). These patients had Ewings (EWSR1 fusion), chondrosarcoma (IDH1 mutation), chondrosarcoma (MSH1,2,6, PMS+), serous carcinoma of endometrium (P1K3CA mutation or TP53 mutation). One patient with osteosarcoma and one patient with desmoplastic small cell tumor had no mutation. A number of responders had mutations in TSCl/2/mTOR and were low to intermediate TMB and MMR deficient (dMMR).

[0266] The results demonstrate that treatment with nivolumab plus m/i-sirolimus was safe with no unexpected adverse events. Unexpectedly, it was found that the best responders were patients with undifferentiated pleomorphic sarcoma with a PTEN loss and a TSC2 mutation and estrogen receptor-positive leiomyosarcoma.

Claims

CLAIMS What is claimed is:

1. A method of treating undifferentiated pleomorphic sarcoma in an individual in need thereof, the method comprising administering to the individual:

(a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and

(b) an effective amount of an anti-PD-1 antibody.

2. The method of claim 1, wherein the undifferentiated pleomorphic sarcoma has phosphatase and tensin homolog (PTEN) loss.

3. The method of claim 2, wherein the PTEN loss is a loss-of-function mutation or epigenetic silencing.

4. The method of claim 2 or 3, wherein the individual is selected for the treatment on the basis of having the PTEN loss.

5. The method of any one of claims 2-4, further comprising selecting the individual on the basis of having the PTEN loss.

6. The method of any one of claims 1-5, wherein the undifferentiated pleomorphic sarcoma has a tuberous sclerosis complex 2 (TSC2) mutation.

7. The method of claim 4, wherein the TSC2 mutation is a missense mutation, nonsense mutation, deletion, splicing site mutation, insertion, substation, rearrangement, or frameshift, or a combination thereof.

8. The method of claim 6 or 7, wherein the individual is selected for the treatment on the basis of the TSC2 mutation.

9. The method of any one of claims 6-8, further comprising selecting the individual on the basis of having the TSC2 mutation.

10. A method of treating leiomyosarcoma in an individual in need thereof, the method comprising administering to the individual: (a) an effective amount of a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin; and

(b) an effective amount of an anti-PD-1 antibody.

11. The method of claim 10, wherein the leiomyosarcoma is estrogen receptor-positive leiomyosarcoma.

12. The method of claim 11, wherein the individual is selected for the treatment on the basis of having the estrogen receptor-positive leiomyosarcoma.

13. The method of claim 11 or 12, further comprising selecting the individual on the basis of having the estrogen receptor-positive leiomyosarcoma.

14. The method of any one of claims 10-13, wherein the leiomyosarcoma is PTEN positive.

15. The method of any one of claims 10-14, wherein the leiomyosarcoma has an intermediate tumor mutation burden.

16. The method of any one of claims 1-15, wherein the undifferentiated pleomorphic sarcoma or the leiomyosarcoma is locally advanced, advanced, malignant, advanced malignant, or metastatic.

17. The method of any one of claims 1-16, wherein the undifferentiated pleomorphic sarcoma or the leiomyosarcoma is relapsed, refractory, or resistant to a prior treatment.

18. The method of claim 17, wherein the prior treatment comprises a composition comprising nanoparticles comprising an mTOR inhibitor and an albumin.

19. The method of any one of claims 1-18, wherein the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is from about 10 mg/m² to about 150 mg/m²

20. The method of claim 19, wherein the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 100 mg/m².

21. The method of claim 19, wherein the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 75 mg/m².

22. The method of claim 19, wherein the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 56 mg/m²

23. The method of claim 19, wherein the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 45 mg/m².

24. The method of claim 19, wherein the amount of the mTOR inhibitor in the mTOR inhibitor nanoparticle composition is about 30 mg/m².

25. The method of any one of claims 1-24, wherein the mTOR inhibitor nanoparticle composition is administered weekly.

26. The method of any one of claims 1-25, wherein the mTOR inhibitor nanoparticle composition is administered 2 out of every 3 weeks.

27. The method of any one of claims 1-26, wherein the mTOR inhibitor nanoparticle composition is administered on days 8 and 15 of a 21-day cycle.

28. The method of any one of claims 1-27, wherein the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered concurrently to the individual.

29. The method of any one of claims 1-27, wherein the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered sequentially to the individual.

30. The method of any one of claims 1-27, wherein the mTOR inhibitor nanoparticle composition and the anti-PD-1 antibody are administered simultaneously to the individual.

31. The method of any one of claims 1-30, wherein the mTOR inhibitor is a limus drug.

32. The method of claim 31, wherein the limus drug is sirolimus.

33. The method of any one of claims 1-32, wherein the average diameter of the nanoparticles in the composition is no greater than about 150 nm.

34. The method of claim 33, wherein the average diameter of the nanoparticles in the composition is no greater than about 120 nm.

35. The method of any one of claims 1-34, wherein the weight ratio of the albumin to the mTOR inhibitor in the nanoparticle composition is no greater than about 9:1.

36. The method of any one of claims 1-35, wherein the nanoparticles comprise the mTOR inhibitor associated with the albumin.

37. The method of claim 36, wherein the nanoparticles comprise the mTOR inhibitor coated with the albumin.

38. The method of any one of claims 1-37, wherein the mTOR inhibitor nanoparticle composition is administered intravenously, intraarterially, intraperitoneally, intravesicularly, subcutaneously, intrathecally, intrapulmonarily, intramuscularly, intratracheally, intraocularly, transdermally, orally, or by inhalation.

39. The method of claim 38, wherein the mTOR inhibitor nanoparticle composition is administered intravenously.

40. The method of any one of claims 1-39, wherein the anti-PD-1 antibody is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, atezolizumab, dostarlimab, durvalumab, and avelumab.

41. The method of any one of claims 1-40, wherein the individual is human.