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WO2000078359A2 - Compositions pour traiter des cellules tumorales resistant a la chimiotherapie et compositions de chimiotherapie ciblee - Google Patents

Compositions pour traiter des cellules tumorales resistant a la chimiotherapie et compositions de chimiotherapie ciblee Download PDF

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Publication number
WO2000078359A2
WO2000078359A2 PCT/US2000/016955 US0016955W WO0078359A2 WO 2000078359 A2 WO2000078359 A2 WO 2000078359A2 US 0016955 W US0016955 W US 0016955W WO 0078359 A2 WO0078359 A2 WO 0078359A2
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WO
WIPO (PCT)
Prior art keywords
peptide
doxorubicin
paclitaxel
cys
acm
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Ceased
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PCT/US2000/016955
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English (en)
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WO2000078359A3 (fr
Inventor
George Tuszynski
Taffy Williams
Paul Actor
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Individual
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Priority to AU54981/00A priority Critical patent/AU5498100A/en
Publication of WO2000078359A2 publication Critical patent/WO2000078359A2/fr
Anticipated expiration legal-status Critical
Publication of WO2000078359A3 publication Critical patent/WO2000078359A3/fr
Ceased legal-status Critical Current

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid

Definitions

  • compositions include a
  • compositions of this invention also include a paclitaxel-
  • compositions of the invention are of the same, resistant cancer resistant cancer and those with normal cancer.
  • the compositions of the invention are of the same, resistant cancer, resistant cancer and those with normal cancer.
  • Doxorubicin is the most commonly used anticancer chemotherapeutic
  • doxorubicin PNAS 93:7269-7273 (1996). It is an anthracycline derived from
  • Dox Doxorubicin
  • the cell and reach certain threshold levels.
  • Paclitaxel is a common chemotherapeutic agent often used to treat
  • TAXOLTM is a natural product with antitumor activity. It is obtained via a
  • Taxus baccata the Pacific Yew Tree
  • paclitaxel 5 ⁇ ,20-Epoxy-1 ,2 ⁇ ,4,7 ⁇ ,13 ⁇ -hexahydroxytax-
  • Paclitaxel acts as an antimicrotubule agent by promoting the assembly
  • paclitaxel induces abnormal arrays
  • cancer is the existence of drug resistance in tumors resulting in decreased cytotoxicity of chemotherapy agents. Some cancers are drug resistant prior
  • doxorubicin by a newly developed compound, oxalyl bis(N-phenyl)hydroxamic
  • MDR multidrug resistance
  • Paclitaxel is a substrate for the multidrug resistant pump, which is also
  • gP170 cells selected for high levels of resistance to this drug
  • tubulin genes and mutations in ⁇ -tubulin genes and mutations in ⁇ -tubulin. Id.
  • Other mechanisms for chemotherapy resistance include: glutathione
  • Another strategy proposes the use of liposome encapulated doxorubicin to
  • Doxorubicin conjugates have also been developed. Doxorubicin
  • doxorubicin has previously resulted in severe loss of cytotoxic activity. See
  • doxorubicin A previous paclitaxel-peptide conjugate has been constructed with a
  • bombesin/gastrin-releasing peptide receptor-recognizing peptide Gln-Trp-
  • a cancer chemotherapy agent is
  • a cancer chemotherapy agent is N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N
  • a peptide is selected, and the agent and peptide are coadministered
  • composition comprises a
  • FIG. 1 Doxorubicin is a drawing of doxorubicin.
  • FIG. 2 (Paclitaxel) is a drawing of paclitaxel.
  • FIG. 3 Doxorubicin - Peptide Conjugate
  • Dox-P peptide conjugate
  • the linkage is an amide linkage.
  • the peptide's amino terminus is modified with an Acm group
  • FIG. 4 (Paclitaxel - Peptide Conjugate) is a drawing of a paclitaxel-
  • ester-amide linkage with the ester linkage being between paclitaxel and the
  • FIG. 5 (Effect of Dox and Dox-P on Drug-Resistant CHO Cell Viability)
  • doxorubicin alone (Dox) is
  • FIG. 6 (Effect of Dox and Dox-P on CHO Cell Viability) demonstrates
  • FIG. 7 (Effect of Doxorubicin Peptide Conjugate on Adhesion of B16-
  • F10 Melanoma Cells show the effect of Dox-P on adhesion of B16-F10
  • FIG. 8 Effect of Doxorubicin (Dox), Doxorubicin-Peptide (Dox-P) on
  • FIG. 9 (Effect of Ala-Ser-Val-Thr-Ala-Arg on Doxorubicin Toxicity of
  • Doxorubicin Resistant CHO Cells shows the effect of coadministering the
  • doxorubicin and multidrug-resistant cells.
  • the present invention relates to chemotherapeutic agents conjugated
  • conjugates can be produced by linking the chemotherapeutic agent to
  • agents can also be coadministered with peptides of the present invention.
  • chemotherapeutic agents can be used in the present disclosure.
  • preferred compounds include those that
  • Acceptable agents including the following:
  • alkylating agents bisulfan, carboplatin, cisplatin, thiotepa
  • nitrogen mustards melphalan, cyclophospamide, chlorambucil
  • antibiotics • antibiotics (doxorubicin, bleomycin, danuorubicin, actinomycin D,
  • idarubicin idarubicin, fludarabine, floxuradine, 5-flurouracil, antracylcine, plicamycin,
  • vinca alkyloids (vincristine, vinblastine);
  • hormonal agonists and antagonists including: nilutamide
  • antiandrogens including: bicalutamide and flutamide;
  • antiestrogens including: anastrozole, toremitine, letrozole, and tamoxifen;
  • estrogens including: estradiol; gonadotropin releasing hormone analogs
  • leuprolide acetate and goserelin acetate including: leuprolide acetate and goserelin acetate; and progestins including:
  • paclitaxel camphothecan, topotecan, vincristine, vinblastine, colchicine
  • methotrexate methotrexate, mercaptopurine, irinotecan, B-methasone, dicarbazine,
  • vinurelbine L-asparginase, paclitaxel, docetaxal, tretinoin, temiposide, ricin,
  • cytoxin cytoxin, saintopin, ellipticin, azatoxin, SQZ, dinalin, and VP16).
  • This invention is most useful when the chemotherapy agent is highly
  • the invention is useful for treating resistant
  • cancers but it is particularly beneficial to use them to treat nonresistant cancers.
  • chemotherapy agents to the tumor cells
  • this invention allows for more effective treatment at lower doses.
  • composition of the present invention contains doxorubicin.
  • composition contains paclitaxel.
  • Peptides of the present invention include peptide sequences from
  • thrombospondin that bind to the thrombospondin receptor.
  • these include peptides binding to the TSP-1 receptor with an
  • Peptides can also be identified by their capacity to bind to the
  • thrombospondin receptor Such peptides can be developed and identified by
  • phage display peptide library kit such as that available from New
  • Phage display describes a selection
  • Phage display can be used to create a physical linkage between a vast library of random peptide sequences to the DNA
  • biopanning This technique is carried out by incubating a
  • the eluted phage is then amplified and taken
  • random peptides can be screened for their ability to bind
  • doxorubicin paclitaxel or other chemotherapeutic agents when conjugated
  • the peptides can be from 3 to 100 amino acids in length, preferably 3
  • thrombospondin protein include the heparin binding domains of the thrombospondin protein, which flank the Cys-Ser-Val-Thr-Cys-Gly (SEQ ID NO: 1) region.
  • Preferred peptides include the heparin binding domains of the thrombospondin protein, which flank the Cys-Ser-Val-Thr-Cys-Gly (SEQ ID NO: 1) region.
  • Val-Thr-Cys(Acm)-Gly (SEQ ID NO: 6) are expected to behave similarly in the
  • the first peptide is subject to oxidation at the sulfur atoms
  • peptide may be less stable.
  • blocking strategies include alkylation with
  • Peptides of the present invention include those with unnatural or non-
  • Such other moieties could include fluorine, chlorine, organic
  • Amino acids or peptides in the d-orientation can also be used, as can
  • compositions of the present invention was produced by:
  • composition of the invention was produced by linking the
  • thrombospondin can be used to form an amide with one end of a dicarboxylic
  • composition of the present invention was produced by linking
  • the ester linkage is between the paclitaxel and the succinyl linker
  • compositions of the present invention may be formulated in a
  • composition which may include carriers, thickeners, diluents,
  • buffers preservatives, surface active agents, liposomes, or lipid formulations
  • compositions may also include one or more
  • antimicrobial agents antimicrobial agents, antiinflammatory agents, anesthetics, and the like.
  • the pharmaceutical composition may be administered in a number of
  • Administration may be topically (including on the skin,
  • parenterally for example by intravenous drip, subcutaneous, intratumor,
  • formulations for topical administration may include ointments,
  • lotions, creams, gels, drops, suppositories, sprays, liquids and powders are included in the formulation.
  • compositions for oral may be necessary or desirable.
  • Compositions for oral may be necessary or desirable.
  • parenteral administration may include sterile aqueous solutions optionally
  • coadministered compound can then be compared to the chemotherapy agent
  • dosages are generally 10x below the lethal dose.
  • the LD 50 the dose that
  • composition's half life can also be determined in one or more animal models,
  • composition of the present invention can be used to adjust the dosages.
  • the dosages can be reduced in amount or
  • Optimal dosing schedules can also be calculated from measurements
  • Optimum dosages may vary depending on the potency of the composition,
  • Dox-P conjugate dosage is from about 5 mg/kg to about 30 mg/kg
  • compositions may be more preferably about 10 mg/kg to about 15 mg/kg and the compositions may
  • chemotherapy agents is about 60 to about 75 mg/m 2 given every 21 days,
  • paclitaxel-p conjugate dosage is from about 50 mg/m 2 to
  • This dosage can be any dosage that can be administered to a patient.
  • This dosage can be any dosage that can be administered to a patient.
  • compositions may be administered as a
  • paclitaxel-p can also be lower than the standard recommended dosages
  • Dox-P doxorubicin-peptide conjugate
  • the method begins with a derivatized thrombospondin peptide, which
  • the flask was transferred to a room temperature water bath and the
  • the solvent was removed at reduced pressure (about 10 "4 torr) while
  • Doxorubicin and multidrug-resistant CHO cells were cultured using
  • Example 1 at concentrations of 0.25, 0.5, 0.75, and 1.0 mM. Untreated cells
  • the assay incorporates a
  • the system incorporates an oxidation-reduction (redox) indicator that
  • absorbance-based instrumentation 570 nm and 600 nm.
  • Example 3 Effect of DOX and DOX-P on wild-type CHO cell viability
  • Example 2 The study of Example 2 was performed, except wild-type CHO cells
  • nonresistant cells is not hindered by the conjugation with the peptide.
  • the LD 50 the dose at which half of the cells die, was calculated from
  • Dox-P can overcome resistance.
  • F10 melanoma cells (a nonresistant cell line) to doxorubicin and the
  • Gly (SEQ ID NO: 6) peptide with a d orientation, or 1% bovine serum albumin
  • the non-adherent cells were removed and the wells were washed with a
  • the peptide to doxorubicin altered its ability to prevent wild-type, nonresistant
  • mice melanoma tumor cells were injected into mice.
  • the animals (5 in each group) were treated intraperitoneally 24 and 96 hours
  • Val-Thr-Cys(Acm)-Gly SEQ ID NO: 6
  • Dox peptide, or Dox
  • mice 18 ⁇ M/kg.
  • the mice were sacrificed and the melanoma tumor colonies on the
  • mice administered to mice at a concentration of 30 and 68 mg/kg, respectively. Both compound have the same number of doxorubicin molecules per weight
  • Thr-Ala-Arg (SEQ ID NO: 2) peptide shows a dose-response effect when
  • Val-Thr-Ala-Arg (SEQ ID NO: 2) peptide may cause this effect by binding to
  • the MDR pump inactivating it or decreasing its effectiveness. It may also
  • Ala-Ser-Val-Thr-Ala-Arg (SEQ ID NO: 2) peptide may also interact with
  • the membrane of the cell such that it modifies the membrane or MDR pump
  • a peptide's ability to bind to the thrombospondin receptor can be
  • response measure in arc seconds is proportional to receptor-
  • Thrombospondin can be used as a positive control.
  • Bovine Aorta Endothelial Cells (BAEC) and MDA-MB-231 cells, breast
  • carcinoma cells are transfected with purified DNA encoding for the receptor
  • TSP-1 thrombospondin
  • bovine serum albumin BSA
  • the stain is washed off and the cells are counted in a field of 1 mm
  • the data can be
  • paclitaxel-peptide conjugate (“Paclitaxel-P”) was produced by the following steps:
  • TAXOLTM paclitaxel
  • 2'-Succinyltaxol was prepared from paclitaxel by a method similar to
  • the activated ester N-hydroxysuccinimide ester of 2' succinyltaxol
  • a cytotoxicity assay can be used to evaluate the
  • cancer cells such as human breast cancer cells
  • paclitaxel-resistant cells paclitaxel-resistant cells
  • paclitaxel alone or the paclitaxel-
  • Untreated cells can be used as a negative control.
  • the assay quantitatively measures the proliferation of cell lines and can establish the
  • the assay incorporates a
  • the system incorporates an oxidation-reduction (redox) indicator that
  • the LD 50 the dose at which half of the cells die, can be calculated from
  • B16-F10 melanoma including B16-F10 melanoma, lewis lung carcinoma, human breast
  • paclitaxel resistant cells An example of a paclitaxel resistant cell line is the SKOV-3TR
  • F10 melanoma cells (a nonresistant cell line) to paclitaxel and the paclitaxel- peptide conjugate of Example 11. This study could also be performed with a
  • bovine serum albumin (BSA).
  • the stain is washed off and the cells are counted in a field of 1 mm
  • doxorubicin alters its ability to prevent wild-type, nonresistant tumor
  • mice melanoma tumor cells are injected into mice.
  • the melanoma tumor cells are injected into mice.
  • mice (5 in each group) are treated intraperitoneally 24 and 96 hours after
  • Val-Thr-Cys(Acm)-Gly (SEQ ID NO: 6) peptide, or paclitaxel, at a
  • the mice are sacrificed and the melanoma tumor colonies on the lung are counted.
  • the LD 50 can be determined experimentally by treating
  • mice with increasing doses of paclitaxel groups of mice with increasing doses of paclitaxel and identifying the
  • a toxicity study can be performed using mice to evaluate the
  • paclitaxel is administered to mice at a concentration which is lower than the
  • LD 50 which can be determined as in Example 15, preferably 10 fold lower
  • the paclitaxel-p conjugage is administered at a dose that
  • Paclitaxel and multidrug-resistant CHO cells are treated with a
  • peptides are co-administered with paclitaxel.
  • peptides are co-administered with paclitaxel.
  • peptide will not show any effect when coadministered with the paclitaxel.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Epidemiology (AREA)
  • Animal Behavior & Ethology (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

Cette invention se rapporte à des procédés et à des compositions servant au traitement du cancer et au traitement des cancers résistant à la chimiothérapie, ces compositions contenant un agent chimiothérapeutique conjugué à un peptide ou administré conjointement à un peptide.
PCT/US2000/016955 1999-06-21 2000-06-21 Compositions pour traiter des cellules tumorales resistant a la chimiotherapie et compositions de chimiotherapie ciblee Ceased WO2000078359A2 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU54981/00A AU5498100A (en) 1999-06-21 2000-06-21 Compositions for treating chemotherapy-resistant tumor cells and targeted chemotherapy compositions

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US14031099P 1999-06-21 1999-06-21
US60/140,310 1999-06-21

Publications (2)

Publication Number Publication Date
WO2000078359A2 true WO2000078359A2 (fr) 2000-12-28
WO2000078359A3 WO2000078359A3 (fr) 2002-01-24

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WO (1) WO2000078359A2 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2003035011A3 (fr) * 2001-10-26 2004-02-26 Uab Research Foundation Conjugues multimedicaments multiligands permettant une administration ciblee de medicaments

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5200397A (en) * 1990-02-22 1993-04-06 W. R. Grace & Co.-Conn. Use of peptide analogs of thrombospondin for the inhibition of angiogenic activity
DE69132708T2 (de) * 1990-09-24 2002-06-20 Philadelphia Health And Education Corp., Philadelphia Therapeutische Vervendung von Peptiden mit Thrombospondin - ähnlicher Aktivität
WO1992017499A1 (fr) * 1991-04-08 1992-10-15 Cornell Research Foundation, Inc. Hexapeptide particulier derive de la thrombospondine et utilisation de ce compose
US5770563A (en) * 1991-12-06 1998-06-23 The United States Of America As Represented By The Department Of Health And Human Services Heparin- and sulfatide binding peptides from the type I repeats of human thrombospondin and conjugates thereof
US5367059A (en) * 1992-05-14 1994-11-22 W. R. Grace & Co.-Conn. Cys-Ser-Val-Thr-Cys-Gly specific tumor cell adhesion receptor
JP2002501483A (ja) * 1997-02-13 2002-01-15 ボシュ アンド ロム サージカル，インコーポレイテッド 網膜色素上皮細胞を破壊するための方法
EP1109900A1 (fr) * 1999-06-21 2001-06-27 Inkine Pharmaceutical Company Inc. Angiocidine: recepteur d'adherence des cellules tumorales specifiques cys-ser-val-thr-cys-gly

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2003035011A3 (fr) * 2001-10-26 2004-02-26 Uab Research Foundation Conjugues multimedicaments multiligands permettant une administration ciblee de medicaments
US7311892B2 (en) 2001-10-26 2007-12-25 The Uab Research Foundation Multidrug multiligand conjugates for targeted drug delivery
US7807134B2 (en) 2001-10-26 2010-10-05 Ahmad Safavy Multidrug multiligand conjugates for targeted drug delivery

Also Published As

Publication number Publication date
WO2000078359A3 (fr) 2002-01-24
AU5498100A (en) 2001-01-09

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