HK1188733A - Bioerodible silicon-based devices for delivery of therapeutic agents - Google Patents

Abstract

This invention discloses bioerodible devices, such as implants for delivering therapeutic agents, particularly large molecules such as proteins and antibodies, in a controlled manner. The devices comprise a porous silicon-based carrier material impregnated with the therapeutic agent. The device may be used in vitro or in vivo to deliver the therapeutic agent, preferably in a controlled fashion over an intended period of time such as over multiple days, weeks or months. The device may be used for treating or preventing conditions of a patient such as chronic diseases.

Description

Bioerodible silicon-based devices for delivery of therapeutic agents

Related application

This application claims benefit of U.S. provisional application No.61/408,934 filed on 1/11/2010 and U.S. provisional application No.61/470,299 filed on 31/3/2011. The entire teachings of the referenced application are expressly incorporated herein by reference.

Background

There is a great deal of interest in the pharmaceutical industry to develop dosage forms that provide controlled release of therapeutic agents over a period of time. Releasing the active in this manner can help improve bioavailability and ensure that the proper concentration of the agent is maintained for a period of time without repeated administrations. This in turn also helps to minimize the effects of patient non-compliance, which is often a problem with other forms of administration.

Patients may be reluctant to comply with their treatment regimen, as compliance may lead to pain and trauma. For example, existing therapeutics can be very successful clinically in treating ophthalmic conditions such as age-related macular degeneration, diabetic macular edema, diabetic retinopathy, choroidal neovascularization, and other conditions that can lead to or are near blindness. The afflicted population is usually an elderly patient population who must adjust their activities of daily living to address these diseases in their early stages. However, as the disease progresses, permanent eye damage occurs and many clinically effective treatments are only prophylactic and not resilient. Therefore, to prevent blindness, it is almost essential to consistently follow a treatment regimen.

Unfortunately, treatment regimens typically require the patient to remain motionless, while the physician penetrates the patient's eye with a hypodermic needle to deliver the therapeutic agent into the eye, typically the vitreous of the eye. This can lead to trauma and pain, and patients may be reluctant to receive injections that may need to be given every week. The ability to provide the long-term benefits of each injection and thus reduce the pain and trauma suffered by the patient depends on the desired pharmacokinetics of the therapeutic agent and the implant loaded with and releasing the therapeutic agent.

Some known implants have active ingredients incorporated into the polymer and sol-gel system by entrapment during matrix phase synthesis. Microencapsulation techniques of biodegradable polymers include methods such as: film casting, molding, spray drying, extrusion, melt dispersion, interfacial deposition, phase separation (by emulsification and solvent evaporation), air suspension coating, pan coating, and in situ polymerization. Melt dispersion techniques are described, for example, in U.S. Pat. No.5,807,574 and U.S. Pat. No.5,665,428.

In an alternative method, the active ingredient is loaded after the porous matrix formation is complete. Such carrier systems typically have microscale, rather than nanoscale, pores to allow the agent to enter the pores. U.S. patent No.6,238,705, for example, describes loading a macroporous polymer composition by simple immersion in a solution of the active ingredient, while U.S. patent nos. 5,665,114 and 6,521,284 disclose the use of pressure to load the pores of an implantable prosthesis made of Polytetrafluoroethylene (PTFE). While this method is effective for small organic molecules, larger molecules such as proteins tend to aggregate in the macropores and cannot be effectively released in a controlled manner in vivo.

Within smaller pores, it has proven difficult to bind high concentrations of therapeutic agents due to blockage of narrow pores. The deposition of material to the orifice tends to prevent a high proportion of the material from occupying the pore system. The difficulty in achieving high active ingredient loadings limits the effectiveness of many of the presently known delivery systems.

Another concern when delivering therapeutic agents through implants is the biocompatibility of the implant following drug release. Bioerodible or resorbable implant materials are an attractive alternative to implants that need to be removed after drug release. The design and preparation of bioerodible implants for carrying therapeutic agents has begun to be explored. PCT publication No. wo2009/009563 describes a drug delivery system comprising a porous silicon material.

Accordingly, there is a continuing need to develop improved dosage forms for controlled release of therapeutic agents that are biocompatible and capable of delivering macromolecules in a sustained manner.

Summary of The Invention

Bioerodible devices, such as implants, for delivering therapeutic agents, particularly macromolecules such as proteins, antibodies, carbohydrates, polymers, or polynucleotides, in a controlled manner are disclosed. The device comprises a porous silicon-based carrier material loaded with a therapeutic agent. The device may be used in vitro or in vivo to deliver therapeutic agents, preferably in a controlled manner for a desired period of time, such as days, weeks or months. The carrier material is preferably formed of a bioerodible or resorbable material, for example a silicon-based material (such as elemental silicon or silicon dioxide), so that removal is not required after release of the therapeutic agent. In certain such embodiments, the carrier material and its decomposition products are biocompatible such that biological side effects resulting from bioerosion of the carrier material are minimized or not deleterious.

In certain embodiments, the support material comprises porous silica, such as mesoporous silica. The average pore size of the carrier material is typically selected such that it can carry the therapeutic agent, and exemplary pore sizes are 2-50nm in diameter, such as about 5 to about 40nm in diameter, about 15 to about 40nm in diameter, about 20 to about 30nm in diameter, about 2 to about 15nm in diameter, or about 5 to about 10nm in diameter.

In certain embodiments, the therapeutic agent is a protein having a molecular weight between 5,000amu and 200,000amu and may be between about 10,000 and about 150,000amu, between 10,000 and 50,000amu, between 50,000 and 100,000amu, or between 100,000 and 200,000 amu.

Alternatively, the size of the therapeutic agent can be characterized by a molecular radius (which can be determined, for example, by X-ray crystallography) or by a hydrodynamic radius. The therapeutic agent may be, for example, a protein having a molecular radius selected from 0.5nm to 20nm, such as about 0.5nm to 10nm, even about 1 to 8 nm. Preferably, the appropriate pore radius to allow entry of a particular agent (e.g., protein) is selected based on the pore-therapeutic agent (agent) difference, defined herein as the difference between the agent radius and the pore radius. For example, insulin with a hydrodynamic radius of 1.3nm has a pore-to-drug difference of 3.5nm from a pore with a minimum radius of 4.8 nm. The pore-agent difference can be used to determine the minimum average pore size suitable to accommodate a particular radius of protein. Pore-protein differences can generally be selected from about 3.0 to about 5.0 nm.

Typically, a device is selected having an average pore size that can accommodate the therapeutic agent. The average pore size of the carrier material can be selected based on the molecular weight or molecular radius of the therapeutic agent to be loaded into the pores of the carrier material. For example, therapeutic agents having a molecular weight selected from 100,000 to 200,000amu may be used with carrier materials having a larger average pore size, such as from about 15nm to about 40 nm. In certain embodiments, therapeutic agents having a molecular weight selected from 5,000 to 50,000amu may be used with carrier materials having a smaller average pore size, such as from about 2nm to about 10 nm.

In certain embodiments, the device is prepared by first forming a porous carrier material and then loading the pores with the therapeutic agent.

The invention includes a method of loading a therapeutic agent into the pores of a porous silicon-based carrier material comprising contacting the porous silicon-based carrier material with the therapeutic agent. An exemplary method for loading a therapeutic agent into pores of a porous silicon-based carrier material comprises selecting a porous silicon-based carrier material having pore diameters sized to allow a single protein to be loaded into the pores such that opposite sides of the protein engage opposite sides of the pores. A method for loading therapeutic agents into the pores of a porous silicon-based carrier material comprises selecting a porous silicon-based carrier having pore sizes dimensioned such that only a single agent is received within the width of a single pore at a time (i.e. longitudinal concatenation along the length of the pore is not excluded), e.g. if two agents are located side-by-side (laterally) in the pore, they cannot be accommodated.

The device may be placed on the skin or on the surface of the eye. Alternatively, the device may be disposed within a mammalian body, such as within a patient's eye or any other tissue or organ of the patient's body. In certain applications, the device is placed subcutaneously, subconjunctivally, or in the vitreous of the eye. The device may be used to treat or prevent a condition, such as a chronic disease, in a patient. In certain embodiments, these devices are used to treat or prevent eye diseases such as glaucoma, macular degeneration, diabetic macular edema, and age-related macular degeneration. The therapeutic agent may be released in a controlled manner, for example over a period of weeks or months, to treat or prevent an eye disease such as macular degeneration.

The present invention includes stabilized formulations and methods of stabilizing therapeutic agents in porous carrier materials as described herein. In certain embodiments, the invention includes stabilizing a biomolecule, such as an antibody, in the pores of a carrier material such that the half-life or the shelf-life of the biomolecule is greater than the half-life or the shelf-life of the biomolecule outside the carrier material.

The invention further comprises a syringe comprising a composition of a porous silicon-based carrier material, wherein the composition comprises less than 2% of biomolecules. These syringes may be used to administer therapeutic agents as follows: a. providing a syringe pre-loaded with a porous silicon-based carrier material; b. contacting the carrier material with a therapeutic agent; administering a carrier material to the patient. Step b may be performed by drawing the therapeutic agent into the syringe. Between steps b and c, an incubation time (e.g. 10min, 20min or 30 min) may be required to allow the therapeutic agent to adsorb into the pores of the carrier material. The therapeutic agent may be selected from small molecules or biomolecules.

Brief Description of Drawings

FIG. 1 shows the pore size distribution of a support material in which the pore size is in a non-uniform bimodal distribution.

FIG. 2 shows the results in PBS and SiO₂Lysozyme released from various silica matrices in saturated PBS. Dissolution medium-PBS: solid content of Davisil■，Davisil▲，DavisilO₂Saturated PBS: o. Davisil□，Davisil△，Davisil

Figure 3 shows the cumulative release of bevacizumab from a silica adsorbent in phosphate buffered saline.

Detailed description of the invention

SUMMARY

The sustained and controlled delivery of therapeutic agents to patients, particularly patients suffering from chronic disorders such as glaucoma or cancer, is becoming increasingly important in modern medical therapy. In order to maintain an almost constant presence of the active agent in the body, many therapies are most effective when administered at frequent intervals. Although frequent administration may be recommended, the inconvenience of patient compliance and associated difficulties may actually preclude treatment in this manner. Accordingly, sustained release devices that release therapeutic agents in a controlled manner are extremely attractive in areas such as cancer therapy and the treatment of other chronic diseases.

Devices for releasing therapeutic agents in vivo or in vitro may be formed from a variety of biocompatible or at least substantially biocompatible materials. One type of device employs a silicon-based carrier material. The silicon-based carrier material may comprise, for example, elemental silicon as well as silica, such as in the form of silica or silicates. Some silicon-based devices have proven to have high biocompatibility and beneficial degradation in biological systems, eliminating the need to remove the device after the therapeutic agent is released.

Tests have shown that high porosity silicon-based materials (e.g., 80% porosity) resorb faster than medium porosity silicon-based materials (e.g., 50% porosity), which resorb faster than bulk silicon-based materials, which exhibit little or no signs of bioerosion or resorption in biological systems. Furthermore, it will be appreciated that the average pore size of the support material will influence the rate of resorption. By adjusting the average pore size of the support material and the porosity of the material, the rate of bioerosion can be adjusted and selected.

Silicon-based devices are typically prepared using high temperatures and organic solvents or acidic media to form porous materials and load therapeutic agents into the pores. These conditions may be suitable for certain molecules such as salts, elements, and certain highly stable small organic molecules. However, for loading large organic molecules, such as proteins or antibodies, corrosion and/or harsh conditions during template preparation or loading may result in denaturation and inactivation (if not complete degradation) of the active agent. Loading macromolecules such as antibodies into a carrier material under mild conditions is a feature of the methods described herein that is particularly advantageous for large organic molecules such as proteins.

The particle size of the silicon-based carrier material may also affect the rate at which the pores of the carrier material can be loaded with therapeutic agent. Smaller particles (e.g., particles having a maximum diameter of 20 microns or less) may be loaded more rapidly than particles having a maximum diameter greater than 20 microns. This phenomenon is particularly evident when the pore diameter is similar in scale to the molecular diameter or size of the therapeutic agent. The rapid loading of the smaller particles may be attributed to the shorter average pore depth that the therapeutic agent must penetrate into the smaller particles.

Definition of

As used herein in the specification, "a" or "an" may mean one or more. As used in the claims herein, the terms "a" or "an" when used in conjunction with the term "comprising" may mean one or more than one. As used herein, "another" may mean at least a second or more.

The term "antibody" broadly encompasses naturally occurring antibody forms as well as recombinant antibodies, such as single chain antibodies, camelized, chimeric and humanized antibodies, and multispecific antibodies and fragments and derivatives of all of the foregoing, preferably fragments and derivatives having at least one antigen binding site. Antibody derivatives may comprise a protein or chemical moiety conjugated to an antibody. The term "antibody" is used in the broadest sense and covers fully assembled antibodies as well as recombinant peptides comprising them.

An "antibody fragment" includes a portion of an intact antibody, preferably the antigen binding or variable region of an intact antibody. Examples of antibody fragments include Fab, Fab ', F (ab')2, and Fv fragments; a diabody; linear antibodies (Zapata et al, Protein Eng.8(10):1057-1062 (1995)); a single chain antibody molecule; and multispecific antibodies formed from antibody fragments. Papain digestion of antibodies produces two identical antigen-binding fragments, referred to as "Fc" fragments, each of which has a single antigen-binding site, and a residual "Fc" fragment, the name of which reflects its ability to crystallize readily. Pepsin treatment produces F (ab')2 fragments that have two antigen binding sites and are still capable of cross-linking antigens.

As used herein, bioerosion refers to the gradual disintegration or breakdown of a structure or shell in a biological system over a period of time, e.g., by one or more physical or chemical degradation processes (e.g., enzymatic action, hydrolysis, ion exchange, or dissolution due to solubilization, emulsion formation, or micelle formation).

The terms "device" and "implant" are used substantially interchangeably herein to refer to the disclosed material, while the term "implant" is used preferentially to refer to a device that is implanted in a patient and not otherwise administered. The various descriptions of the device embodiments are intended to apply equally to implants, and vice versa.

The term "preventing" is art-recognized and is well known in the art when used with respect to a disorder such as local recurrence (e.g., pain), a disease such as cancer, a syndrome such as heart failure, or any other medical disorder, and includes administering a composition that reduces the frequency of, or delays the onset of, a symptom of a medical disorder in a subject relative to a subject not receiving the composition. Thus, preventing cancer includes, for example, reducing the number of detectable cancerous growths in a patient population receiving prophylactic treatment relative to an untreated control population, and/or delaying the appearance of detectable cancerous growths in a treated population by a statistically and/or clinically significant amount relative to an untreated control population. Preventing infection includes, for example, reducing the number of diagnoses of infection in the treated population relative to an untreated control population, and/or delaying the onset of symptoms of infection in the treated population relative to an untreated control population. Preventing pain includes, for example, reducing the magnitude of pain sensation experienced by a subject in the treated population relative to an untreated control population or alternatively delaying the pain sensation experienced by a subject in the treated population.

The term "prophylactic or therapeutic" treatment is art-recognized and includes the administration of one or more of the subject compositions to a host. If administered prior to clinical manifestation of the undesired condition (e.g., disease or other undesired state of the host animal), the treatment is prophylactic (i.e., it prevents the host from developing the undesired condition), whereas if administered after manifestation of the undesired condition, the treatment is therapeutic (i.e., it is intended to attenuate, alleviate or stabilize the existing undesired condition or side effects thereof).

Resorption, as used herein, refers to the erosion of a material when introduced into or onto a physiological organ, tissue or body fluid of a living human or animal.

For the subject treatment methods, a "therapeutically effective amount" of a compound refers to the amount of the compound in a preparation that, when administered (to a mammal, preferably a human) as part of a desired dosage regimen, may, for example, alleviate symptoms, alleviate the condition, or delay the onset of the condition at a reasonable benefit/risk ratio applicable to any drug treatment, depending on the condition or condition being treated or the clinically acceptable criteria for cosmetic purposes.

As used herein, the term "treating" includes reversing, reducing, or arresting the symptoms, clinical signs, and underlying pathology of the condition in a manner that ameliorates or stabilizes the condition in the subject.

Unless otherwise indicated, the term therapeutic macromolecule refers to a molecule having a molecular weight equal to or greater than 1000amu, preferably greater than 2000amu or even greater than 3000 amu. Unless otherwise indicated, a small molecule therapeutic molecule refers to a molecule having a molecular weight of less than 1000 amu.

Silicon-based materials and other bioerodible supports

The devices and methods described herein provide, inter alia, devices comprising a porous silicon-based carrier material, wherein at least one therapeutic agent is disposed in the pores of the carrier material. The methods use such devices to treat or prevent diseases, particularly chronic diseases. In addition, the method of making the device provides a device characterized by sustained and controlled release of therapeutic agents, particularly large molecules such as proteins or antibodies.

The device typically comprises a silicon-based carrier material, such as elemental silicon, silicon dioxide, silicon monoxide, silicates (containing anions with silicon, e.g. SiF)₆ ^2-、Si₂O₇ ^6-Or SiO₄ ^4-Compound(s) or any combination of such materials. In certain embodiments, the support material comprises a complete or partial framework of elemental silicon and the framework is substantiallyCovered or completely by a surface layer of silicon dioxide. In other embodiments, all or substantially all of the support material is silica.

While silicon-based materials are the preferred support materials for use in the present invention, additional bioerodible materials having certain common characteristics (e.g., porosity, pore size, particle size, surface characteristics, bioerodibility, and resorbability) with the silicon-based materials described herein can also be used in the present invention. Examples of additional materials that can be used as porous support materials are bioerodible ceramics, bioerodible metal oxides, bioerodible semiconductors, bone phosphates, calcium phosphates (e.g., hydroxyapatite), other inorganic phosphates, carbon black, carbonates, sulfates, aluminates, borates, aluminosilicates, magnesium oxide, calcium oxide, iron oxide, zirconium oxide, titanium oxide, and other similar materials.

In certain embodiments, the support material comprises silica, such as greater than about 50% silica, greater than about 60% silica by weight, greater than about 70% silica by weight, greater than about 80% silica by weight, greater than about 90% silica by weight, greater than about 95% silica by weight, greater than 99% silica by weight, or even greater than 99.9% silica by weight. Porous silicas are available from suppliers such as Davisil, Silicycle and Macherey-Nagel.

In certain embodiments, the support material comprises elemental silicon, greater than 60 wt.% silicon, greater than 70 wt.% silicon, greater than 80 wt.% silicon, greater than 90 wt.% silicon, or even greater than 95 wt.% silicon. Silicon is available from suppliers such as Vesta Ceramics.

The purity of silicon-based materials can be quantitatively assessed using techniques such as: energy dispersive X-ray analysis, X-ray fluorescence, inductively coupled optical emission spectroscopy, or glow discharge mass spectroscopy.

The carrier material may comprise other components such as metals, salts, minerals or polymers. The carrier material may have a coating disposed on at least a portion of the surface, for example, to improve biocompatibility of the device and/or to influence release kinetics.

The silicon-based carrier material may comprise elemental silicon or compounds thereof, such as silica or silicates, in amorphous form. In certain embodiments, the elemental silicon or compound thereof is present in crystalline form. In other embodiments, the carrier material comprises amorphous silicon dioxide and/or amorphous silicon. In certain embodiments, the silicon-based material has greater than about 60 wt% amorphous, greater than about 70 wt% amorphous, greater than about 80 wt% amorphous, greater than about 90 wt% amorphous, greater than about 92 wt% amorphous, greater than about 95 wt% amorphous, greater than about 99 wt% amorphous, or even greater than 99.9 wt% amorphous.

X-ray diffraction analysis can be used to identify the crystalline phase of silicon-based materials. Powder diffraction can be performed on, for example, a Scintag PAD-X diffractometer, such as one equipped with a liquid nitrogen cooled germanium solid state detector using Cu K-alpha radiation.

The silicon-based material may have a porosity of about 40% to about 95%, such as about 60% to about 80%. As used herein, porosity is a measure of the void space in a material and is the fraction of voids that make up the total volume of the material. In certain embodiments, the support material has a porosity of at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, or even at least about 90%. In particular embodiments, the porosity is greater than about 40%, such as greater than about 50%, greater than about 60%, or even greater than about 70%.

The carrier material of the device may have a thickness selected from about 20m²A/g to about 2000m²Surface area/weight ratio of/g, such as about 20m²G to about 1000m²In g or even about 100m²G to about 300m²(ii) in terms of/g. In certain embodiments, the surface area is greater than about 200m²A ratio of the total of the components of the composition to the total of the components of the composition greater than about 250m²/g or greater than about 300m²/g。

In certain embodiments, the therapeutic agent is distributed at a pore depth of at least about 10 microns, at least about 20 microns, at least about 30 microns, at least about 40 microns, at least about 50 microns, at least about 60 microns, at least about 70 microns, at least about 80 microns, at least about 90 microns, at least about 100 microns, at least about 110 microns, at least about 120 microns, at least about 130 microns, at least about 140 microns, or at least about 150 microns from the surface of the material. In certain embodiments, the therapeutic agent is substantially uniformly distributed in the pores of the carrier material.

The depth to which the therapeutic agent can be loaded into the carrier material is measured as the ratio of the depth to which the therapeutic agent penetrates the carrier material to the total width of the carrier material. In certain embodiments, the therapeutic agent is distributed to a depth of at least about 10% of the carrier material, at least about 20% of the carrier material, at least about 30% of the carrier material, at least about 40% of the carrier material, at least about 50% of the carrier material, or at least about 60% of the carrier material.

Quantification of the total loading can be achieved by a number of analytical methods, such as gravimetric analysis of pharmaceutical compositions, EDX (energy dispersive X-ray analysis), Fourier Transform Infrared (FTIR) spectroscopy or raman spectroscopy; or uv spectrophotometry, titrimetric analysis, HPLC or mass spectrometry of the therapeutic agent eluted in the solution. Quantification of loading uniformity can be obtained by combinatorial techniques capable of spatial resolution, such as cross-sectional EDX, Auger depth profiling, micro-raman spectroscopy, and micro-FTIR.

The porous silicon-based materials of the present invention can be classified according to the average diameter of the pore diameter. The silicon-based microporous material has an average pore size of less than 2nm, the silicon-based mesoporous material has an average pore size between 2-50nm and the silicon-based macroporous material has a pore size greater than 50 nm. In certain embodiments, greater than 50% of the pores of the silicon-based material have a pore size of 2-50nm, greater than 60% of the pores of the silicon-based material have a pore size of 2-50nm, greater than 70% of the pores of the silicon-based material have a pore size of 2-50nm, greater than 80% of the pores of the silicon-based material have a pore size of 2-50nm, or even greater than 90% of the pores of the silicon-based material have a pore size of 2-50 nm.

In certain embodiments, the support material comprises porous silica, such as mesoporous silica. In certain embodiments, the average pore size of the support material is selected from 2-50nm, such as from about 5 to about 40nm, from about 15 to about 40nm, such as from about 20 to about 30 nm. In certain embodiments, the average pore size is selected from about 2 to about 15nm, such as about 5 to about 10 nm. In certain embodiments, the average pore size is about 30 nm.

In certain embodiments, the support material has a population of pores with well-defined pore sizes, i.e., the pore size distribution of the support material is within a defined range. In certain embodiments, about 50% to about 99% of the pore diameters in a well-defined population of pores are within an average pore diameter of about 1nm to 15nm for that population, preferably within an average pore diameter of about 10nm, about 5nm, or even 3nm or 2nm for that population. In certain such embodiments, greater than about 50%, greater than about 60%, greater than about 70%, greater than about 80%, greater than about 90%, or even greater than about 95% of the pores in the support material have a pore size within the specified range. Similarly, a population of pores having a well-defined pore size may be a population in which greater than about 50%, greater than about 60%, greater than about 70%, greater than about 80%, greater than about 90%, or even greater than about 95% of the pores have a pore size within 20%, preferably within 15%, 10%, or even within 5% of the average pore size of the population.

Pore (e.g., mesopore) size distribution can be determined using established analytical methods such as gas adsorption, high resolution scanning electron microscopy, nuclear magnetic resonance cryo-porosimetry, and differential scanning calorimetry. In certain embodiments, more than one technique is used for a given sample.

Alternatively, a population of pores having a well-defined pore size may be a population having a standard deviation of pore size of less than 20%, preferably less than 15%, less than 10% or even less than 5% of the average pore size of the population.

The pore size can be preselected relative to the dimensional characteristics of the therapeutic agent to control the release rate of the therapeutic agent in the biological system. In general, too small a pore size hinders loading of the therapeutic agent, while too large a pore does not create a strong enough interaction with the therapeutic agent to exert the desired control over the release rate. For example, the average pore diameter of the support material may be selected from larger pores, e.g.15 nm to 40nm, for high molecular weight molecules (e.g.200,000-500,000 amu) and smaller pores, e.g.2 nm to 10nm, for lower molecular weight molecules (e.g.10,000-50,0000 amu). For example, for molecules having a molecular weight of about 14,000 to 15,000amu, such as about 14,700amu, an average pore diameter of about 6nm in diameter may be suitable. For molecules having a molecular weight of about 45,000 to 50,000amu, such as about 48,000amu, an average pore diameter of about 10nm in diameter may be selected. For molecules having a molecular weight of about 150,000amu, an average pore size of about 25-30nm in diameter may be selected.

The pore size can be preselected to suit the molecular radius of the therapeutic agent to control the release rate of the therapeutic agent in the biological system. For example, an average pore size of about 25nm to about 40nm in diameter may be suitable for molecules having a maximum molecular radius of about 6nm to about 8 nm. The molecular radius may be calculated by any suitable method, such as by using molecular physical dimensions based on X-ray crystallographic data or using hydrodynamic radii that represent the size of the molecular solution. Since solution state calculations depend on the nature of the solution in which the calculations are made, some measurements may preferably use molecular physical dimensions based on X-ray crystallographic data. As used herein, the maximum molecular radius reflects half of the maximum dimension of the therapeutic agent.

In certain embodiments, the average pore diameter is selected to limit aggregation of molecules (e.g., proteins) within the pores. Preventing aggregation of biomolecules, such as proteins, in the device is advantageous because it is believed that aggregation of biomolecules, such as proteins, in the device prevents controlled release of the molecules into the biological system. Thus, a pore that, for example, allows only one biomolecule to enter the pore at any one time due to the relationship between its size and the size of the biomolecule, would be superior to a pore that allows multiple biomolecules to enter and aggregate together within the pore. In certain embodiments, multiple biomolecules may be loaded into a well, but due to the depth of the well, proteins distributed throughout the depth of the well will aggregate to a lesser extent.

In certain embodiments, the carrier material comprises two or more different materials having different characteristics (e.g., pore size, particle diameter, or surface characteristics), each preselected to be suitable for a different therapeutic agent. For example, two different carrier materials may be compounded, one having a first population of pores with a pore size suitable for a first therapeutic agent and the other having a second population of pores with a pore size suitable for a second therapeutic agent. In certain other embodiments, the carrier material comprises a single material having two or more well-defined populations of pores, e.g., wherein the carrier material is made by molecular templating techniques, wherein the characteristics of the pores are pre-selected for two or more therapeutic agents, e.g., two therapeutic agents having different molecular radii. Thus, the carrier material can deliver two or more therapeutic agents in a controlled manner as described herein. In such embodiments, the loading of the therapeutic agent is preferably done in order of largest to smallest agent, such that the largest agent is selectively adsorbed into the largest pores (i.e., it does not fit into the smaller pores), such that the larger pores do not adsorb smaller agents.

For example, if the carrier material comprises a first well-defined population of pores having a diameter of about 6nm (i.e., suitable for molecules having a molecular weight of about 14,000 to 15,000 amu) and a second well-defined population of pores having a diameter of about 10nm (i.e., suitable for molecules having a molecular weight of about 45,000 to 50,000 amu), the latter therapeutic agent (i.e., a therapeutic agent having molecules having a molecular weight of about 45,000 to 50,000 amu) is preferably added to the carrier material prior to the addition of the smaller therapeutic agent (i.e., a therapeutic agent having molecules having a molecular weight of about 14,000 to 15,000 amu). Alternatively and additionally, in embodiments where two different porous materials together comprise a device, each carrier material may be separately loaded with a different therapeutic agent and these carrier materials may then be combined to create a device.

In certain embodiments where the carrier material has two or more well-defined distinct pore populations (e.g., the distinct pore populations do not substantially overlap), the differences between the characteristics of the distinct pore populations are preferably selected to limit adsorption of each distinct therapeutic agent to a certain pore population. In certain embodiments, the average pore size of two or more well-defined distinct populations of pores may be selected to limit adsorption of larger therapeutic agents into smaller pores. The average pore size difference can be defined as the difference between the average pore sizes of different populations of pores in the support material. For example, a difference in average pore diameter of at least 10nm may indicate that the support material may comprise at least two pore populations that differ in average pore diameter by at least 10nm ("average pore diameter difference"), e.g., the composition may comprise two pore populations having average pore diameters of 10nm and 20nm, three pore populations having average pore diameters of 10nm, 20nm, and 30nm, or four pore populations having average pore diameters of 10nm, 20nm, 30nm, and 40 nm. In certain embodiments, the average pore size difference is preferably at least about 5nm, at least about 10nm, at least 15nm, at least about 20nm, or at least about 30 nm. In certain embodiments, two or more well-defined pore populations have different average pore sizes such that the average pore size of any two pore populations differs from the smaller average pore size by at least 20%, preferably at least 30%, 40%, or even 50%.

In certain embodiments where the support material has a non-uniform pore size distribution, the support material has two or more well-defined populations of pores having different average pore sizes as described above. Similarly, with reference to fig. 1, a support material having a non-uniform pore size distribution can be characterized as a pore size distribution having at least two local maxima (e.g., in fig. 1, one at a pore size equal to a and one at a pore size equal to B), but as many as three or four local maxima, with the number of pores having the pore size of two adjacent local maxima (e.g., M in fig. 1)_XAAnd M_XB) Is the average of the pore diameters with two local maxima (e.g., M in FIG. 1_NABWhere the mean pore diameter of the two local maxima is AV_AB) Is at least three times, but preferably five times, ten times or even 20 times the number of pores. The pore size distribution can also be described by the following equation, which also applies where M_XAAnd M_XBIn certain embodiments where there is no equality, for example, the distribution is not exactly bimodal:

M_XA≥3(M_NAB) And M_XB≥3(M_NAB)，

Wherein M is_XANumber of particles of pore size a; m_XBNumber of particles of pore size B; and M_NABParticle number of pore size (a + B)/2, and wherein 3 may be replaced by any suitable multiplier as described above.

In certain embodiments, the therapeutic agent is selected from any agent suitable for treating or preventing a disease. In certain embodiments, the agent is selected from small molecule therapeutic agents, i.e., compounds having a molecular weight of less than 1000 amu. In a preferred embodiment, the therapeutic agent is selected from macromolecules having a molecular weight equal to or greater than 1000 amu. In certain embodiments, the therapeutic agent of the present invention is a biomolecule. As used herein, a biomolecule refers to any molecule produced by a living organism, including large polymer molecules such as proteins, polysaccharides, and nucleic acids; and small molecules such as primary metabolites, secondary metabolites, and natural products or synthetic variants thereof. In particular, proteins such as antibodies, ligands, and enzymes may be used as the therapeutic agents of the present invention. In particular embodiments, the biomolecules of the invention have a molecular weight in the range of about 10,000amu to about 500,000 amu. In certain embodiments, the therapeutic agent is selected from one or more monoclonal antibodies, such as ranibizumab (Lucentis) and bevacizumab (Avastin).

In certain embodiments, the therapeutic agent has a molecular weight between 10,000 and 50,000amu, between 50,000 and 100,000amu, or between 100,000 and 150,000 amu. In certain embodiments, the therapeutic agent is a protein having a molecular weight between 5,000amu and 200,000amu, such as a molecular weight of about 10,000 to about 150,000 amu.

Alternatively, the size of the therapeutic agent can be characterized by a molecular radius (which can be determined, for example, by X-ray crystallography) or by a hydrodynamic radius. The therapeutic agent may be, for example, a protein having a molecular radius selected from 0.5nm to 20nm, such as about 0.5nm to 10nm, even about 1nm to 8 nm.

Therapeutic agents having a molecular radius of 1 to 2.5nm may be advantageously used with carrier materials having a minimum pore radius of 4.5 to 5.8 nm. Therapeutic agents having a molecular radius of 7nm may be advantageously used with carrier materials having a minimum pore radius of 11 to 13nm, such as about 12 nm. For example, insulin having a hydrodynamic radius of 1.3nm may be used with a carrier material having an average minimum pore radius of 4.8 nm.

Protein-pore differences can be used to select suitable carrier materials to accommodate the therapeutic agent. This calculated value is the pore radius minus the molecular radius. Typically, the radius of the therapeutic agent will be the hydrodynamic radius or the maximum radius as determined via X-ray crystallography analysis. The pore radius will typically be the average pore radius of the support material. For example, insulin with a hydrodynamic radius of 1.3nm has a protein-pore difference of 3.5nm from the pore-protein difference of a pore with a minimum radius of 4.8 nm. In certain embodiments, the protein-pore difference is selected from 3 to 6nm, such as 3.2 to 4.5 nm. The protein-pore difference can be about 3.2nm, about 3.3nm, about 3.4nm, about 3.5nm, about 3.6nm, about 3.7nm, about 3.8nm, about 3.9nm, about 4.0nm, about 4.1nm, about 4.2nm, about 4.3nm, about 4.4nm, or about 4.5 nm.

In certain embodiments, the therapeutic agent is an antibody and the average pore size of the carrier material is selected from about 5nm to about 40nm, for example about 10nm to about 40nm, such as about 20nm to about 40nm, such as about 25nm to 35nm, such as about 30 nm. In certain embodiments, the therapeutic agent is an antibody selected from bevacizumab or ranibizumab and the average pore size of the carrier material is selected from about 5nm to about 40nm, such as 10nm to about 40nm, such as about 25nm to 35nm, such as about 30 nm. In certain embodiments, the therapeutic agent is bevacizumab and the average pore size of the carrier material is about 30 nm.

In certain embodiments, the walls of the support material separating the pores have an average width of less than 5nm, such as about 4.8nm, about 4.6nm, about 4.4nm, about 4.2nm, about 4.0nm, about 3.8nm, about 3.6nm, about 3.4nm, about 3.2nm, about 3.0nm, about 2.8nm, or even about 2.6 nm. In certain embodiments, the walls of the support material separating the pores have an average width of less than about 3nm, such as about 2.8nm, about 2.6nm, about 2.4nm, about 2.2nm, about 2.0nm, about 1.8nm, about 1.6nm, about 1.4nm, about 1.2nm, about 1.0nm, or even about 0.8 nm.

The dimensions and morphology of the device can be measured, for example, by Transmission Electron Microscopy (TEM) using, for example, a 2000JEOL electron microscope operating at 200 keV. Samples for TEM can be prepared by dispensing a large amount of porous support material via a dilute slurry onto a porous carbon film on a metal grid.

In certain embodiments, the space defined by the pores of the support material has a volume of from about 0.1mL/g to about 5mL/g per gram of support material. In certain embodiments, the pore volume is from about 0.2mL/g to about 3mL/g, such as from about 0.4mL/g to about 2.5mL/g, such as from about 1.0mL/g to about 2.5 mL/g.

In certain embodiments, the loading of carrier material is up to 70 weight percent, such as up to 40 weight percent, based on the combined weight of the carrier material and therapeutic agent. The loading is calculated by dividing the weight of loaded therapeutic agent by the total weight of loaded therapeutic agent and carrier material and multiplying by 100. In certain embodiments, the loading of the support material is greater than 10%, such as greater than 15%, greater than 20%, greater than 25%, greater than 30%, greater than 35%, greater than 40%, greater than 45%, or greater than 50%. In certain embodiments, the loading of the support material is less than 5%. The loading may be between about 5% and about 10%. In certain embodiments, the loading of the support material is between about 10% and about 20% by weight, between about 20% and about 30% by weight, between about 30% and about 40% by weight, between about 40% and about 50% by weight, or between about 50% and about 60% by weight.

The loading volume of the devices described herein can be evaluated based on the pore volume occupied by the therapeutic agent in the porous material. For carrier materials according to the invention, the percentage of the maximum loading capacity occupied by the therapeutic agent (i.e., the percentage of the total volume of pores in the porous carrier material occupied by the therapeutic agent) may be from about 30% to about 100%, such as from about 50% to about 90%. For any given carrier material, this value can be determined by dividing the volume occupied by the therapeutic agent during loading by the void volume of the carrier material prior to loading and multiplying by 100.

In certain embodiments, the devices of the present invention are particles having an average size, measured as the largest diameter, of from about 1 to about 500 microns, such as from about 5 to about 100 microns. In certain embodiments, the single device measured at its largest diameter is from about 1 to about 500 microns, such as from about 5 to about 500 microns.

To increase the loading rate of the particles of the invention, it may be advantageous to use relatively small particles. Because the smaller particles have shallower depth pores for the therapeutic agent to penetrate, the amount of time required to load the particles may be reduced. This can be particularly advantageous when the pore diameter is similar in dimension to the molecular diameter or size of the therapeutic agent. The smaller particles may be 1-20 microns, such as about 10-20 microns, for example about 15-20 microns, when measured on the largest dimension.

In some aspects, greater than 60%, greater than 70%, greater than 80%, or greater than 90% of the particles have a particle size of 1 to 20 microns, preferably 5 to 15 microns, when measured on the largest dimension. The average particle size of the particles may be between 1 and 20 microns, such as between 5-15 microns or about 15 microns, about 16 microns, about 17 microns, about 18 microns, about 19 microns.

The particle size distribution (including the mean particle diameter) can be measured, for example, using a Malvern particle size analyzer (model Mastersizer) from Malvern instruments, UK. A helium-neon gas laser beam may be projected through an optical cell containing a suspension of carrier material. Light impinging on the support material is scattered through an angle inversely proportional to the particle size. The photodetector array measures light intensities at a plurality of predetermined angles, and an electrical signal proportional to the measured light flux values is then processed by the microcomputer system according to a scattering pattern predicted by the refractive indices of the sample carrier material and the aqueous dispersant.

Larger devices/implants are also contemplated for controlled delivery of therapeutic agents. The devices/implants of the present invention may have an average size, measured on the largest dimension, of from about 1mm to about 5 cm. In certain embodiments, the device/implant has an average size, measured on the largest dimension, of about 5mm to about 3 cm. Implants larger than 1mm may be used for intramuscular, subcutaneous, intravitreal, or subdermal drug delivery, as measured on the largest scale.

In certain embodiments, the porous carrier materials described herein are used to stabilize sensitive therapeutic compounds, such as biomolecules, e.g., antibodies. In certain embodiments, biomolecules that are partially or completely unstable at elevated temperatures (such as at or above room temperature) can be stabilized at room temperature for extended periods of time. The biomolecules may be loaded into the carrier material such that the aqueous suspension of the biomolecules loaded into the carrier material is more stable than the aqueous solution of the corresponding biomolecules (i.e. the same aqueous solution with and without the addition of the porous carrier material). For example, the half-life of a biomolecule within the carrier material at room temperature (e.g., about 23 ℃) can be greater than the half-life of a biomolecule without the carrier material under the same conditions. In certain embodiments, the half-life of the biomolecule in the pores of the carrier material is at least twice as long as the biomolecule without the carrier material under the same conditions, more preferably at least 5 times, at least 10 times, at least 15 times, at least 20 times, at least 30 times, at least 40 times, at least 50 times, at least 60 times or at least 100 times as long as the biomolecule without the carrier material. For example, the half-life of the antibody within the pores of the support material may be at least 10 times longer, more preferably at least 20 times longer than that of the antibody without the support material.

Similarly, the biomolecules may have a longer shelf life within the pores of the support material than in a corresponding aqueous solution, preferably at least 2 times longer, at least 5 times longer, at least 10 times longer, at least 20 times longer, at least 30 times longer, at least 40 times longer, at least 50 times longer, at least 60 times longer, or at least 100 times longer. For example, the antibodies within the pores of the carrier material may have a longer useful life than antibodies without the carrier material, preferably at least 10 times longer or at least 20 times longer.

In certain embodiments, a porous device comprising a carrier material and a biomolecule, such as an antibody, exhibits stability at a temperature of 25 ℃ for at least 15 days or even about 1 month. Additionally or alternatively, in certain embodiments, the antibody-loaded device is stable at 25 ℃ for at least 6 months, at least 1 year, at least 1.5 years, at least 2 years, at least 2.5 years, at least 3 years, or at least 4 years. Stability can be assessed, for example, by the following method: high Performance Size Exclusion Chromatography (HPSEC), or comparing the biological activity of the biomolecule-loaded device after storage to a sample of a biomolecule-loaded device as-made or to the device activity measured before storage. For example, antibody activity can be assessed by various immunological assays, including, for example, enzyme-linked immunosorbent assays (ELISAs) and radioimmunoassays. Preferably, at the end of the storage period, the activity of the stored device is at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, at least 99.5%, at least 99.8% or even at least 99.9% of the activity of the corresponding ready-made device. Thus, the invention encompasses methods of treatment wherein the biomolecule-loaded devices have been stored at 25 ℃ for at least 6 months, at least 1 year, at least 1.5 years, at least 2 years, at least 2.5 years, at least 3 years, or at least 4 years prior to administration of the devices to a patient.

The invention further includes methods of stabilizing biomolecules. The method of the invention comprises loading biomolecules into the pores of a carrier material by any suitable method to form the device of the invention.

Preparation method

The invention also provides a method for preparing the silicon-based device. In certain embodiments, the porous silicon-based support material may be prepared by synthetic methods. For example, porous silica can be synthesized by reacting tetraethylorthosilicate with a template made from micellar rods. In certain embodiments, a collection of balls or rods filled with regularly arranged holes is obtained. The template can then be removed, for example by washing with a solvent adjusted to a suitable pH. In certain embodiments, the porous silicon-based support material may be prepared using a sol-gel process or a spray-drying process. In certain embodiments, the preparation of the support material involves one or more techniques suitable for preparing a porous silicon-based material.

The pores may be introduced into the silicon-based carrier material by techniques such as anodization, dye etching, or electrochemical etching. In one exemplary embodiment, anodization employs a platinum cathode immersed in a Hydrogen Fluoride (HF) electrolyte and a silicon wafer anode. Anodic corrosion, which creates pores in the material, is created by passing an electric current through the cell. In particular embodiments, a constant Direct Current (DC) is typically flowed to ensure a steady state tip concentration of HF, resulting in a more uniform porous layer.

In certain embodiments, the pores are incorporated into the silicon-based support material by dye etching with hydrofluoric acid, nitric acid, and water. In certain embodiments, a combination of one or more dyed etchants is used, such as hydrofluoric acid and nitric acid. In certain embodiments, a solution of hydrofluoric acid and nitric acid is used to form pores in a silicon-based material.

The porosity of the material can be determined by gravimetric measurements. BET analysis may be used to determine any one or more of pore volume, pore size distribution, and surface area of the support material. The BET theory (the initials incorporated by the authors of this theory) applies to the physical adsorption of gas molecules on solid surfaces and serves as the basis for important analytical techniques for measuring the specific surface area of materials (j.am. chem. soc., v.60, p.309 (1938)). BET analysis can be performed, for example, using a Micromeritics ASAP2000 Instrument available from Micromeritics Instrument Corporation, Norcross, Georgia. In an exemplary procedure, a sample of the support material may be vacuum degassed at a temperature, for example, greater than 200 ℃ for a period of time, such as about 2 hours or more, prior to taking the measurement. In certain embodiments, the pore size distribution curve is derived from analysis of adsorption branches at the isotherm output. Pore volume may be concentrated in P/P₀Single dot of = 0.985.

One or more drying techniques may be used to prepare the porous silicon-based materials of the present invention. For example, to prevent cracking of the porous silicon-based material, the material may be dried by supercritical drying, freeze drying, pentane drying or slow evaporation. Supercritical drying involves overheating the liquid pores beyond the critical point to avoid interfacial tension. Freeze-drying involves freezing and sublimating any solvent under vacuum. Pentane drying uses pentane as the drying liquid rather than water and therefore can reduce capillary stress due to lower surface tension. Slow evaporation is a technique that can be performed after water or ethanol washing and is effective in reducing the trap density of the solvent within the material.

The surface of the porous silicon-based material may be modified to exhibit properties such as improved stability, cell adhesion, or biocompatibility. Optionally, the material may be exposed to oxidizing conditions, such as by thermal oxidation. In one exemplary embodiment, the thermal oxidation process involves heating the silicon-based material to a temperature above 1000 ℃ to promote complete oxidation of the silicon-based material. Alternatively, the surface of the support material may be oxidized such that the support material comprises an elemental silicon framework partially, substantially or completely covered by an oxidized surface (such as a silicon dioxide surface).

The surface of the porous silicon-based material or a portion thereof may be derivatized. In an exemplary embodiment, the surface of the porous silicon-based material may be derivatized with organic groups such as alkanes or alkenes. In a particular embodiment, the surface of the support material may be derivatized by a hydrosilation reaction of silicon. In particular embodiments, the derivatized carrier material may serve as a biomaterial that is incorporated into living tissue.

Any one or more of electrostatic interactions, capillary interactions, and hydrophobic interactions may cause the therapeutic agent to be loaded into the pores of the carrier material. In certain embodiments, the carrier material and therapeutic molecule are placed in solution and large molecules (e.g., proteins or other antibodies) are drawn from the solution into the pores of the carrier material, which is reminiscent of the ability of the molecular sieve to draw water from the organic liquid. Hydrophobic drugs may be more suitable for loading into carrier materials formed primarily of silicon (e.g., greater than 50% of the material is silicon), while hydrophilic drugs may be more suitable for loading into carrier materials characterized primarily as silica (e.g., greater than 50% of the carrier material is silica). In certain embodiments, the loading of macromolecules into the pores of the support material is driven by external factors such as sonication or heating. The carrier material or a portion thereof can have an electrostatic charge and/or the therapeutic agent or a portion thereof can have an electrostatic charge. Preferably, the carrier material or a portion thereof has an electrostatic charge opposite to that of the therapeutic agent or a portion thereof, such that adsorption of the therapeutic agent into the pores of the carrier material is facilitated by electrostatic attraction forces. In certain embodiments, the therapeutic agent or carrier material may not have an electrostatic charge on its own, but rather may polarize and change its polarity in the vicinity of the carrier material or therapeutic agent, respectively, which facilitates adsorption of the therapeutic agent into the pores of the carrier material.

The carrier material may comprise a coating or surface modification to attract the therapeutic agent into the pores. In certain embodiments, the support material is fully or partially coated with or modified with a material comprising charged moieties in order to attract proteins or antibodies into the pores of the support material. In other embodiments, these portions may be attached directly to the carrier material. For example, the amine groups may be covalently attached to the surface of the carrier material such that the surface of the carrier material carries a positive charge when protonated at physiological pH, thereby attracting, for example, proteins or antibodies having a negatively charged surface. In other embodiments, the carrier material may be modified with carboxylic acid moieties such that the carrier material carries a negative charge when deprotonated at physiological pH values, thereby attracting proteins or antibodies with positively charged surfaces into the pores.

In certain embodiments, the support material may be a material other than porous silica. While silicon-based materials are the preferred support materials for use in the present invention, additional bioerodible materials having certain characteristics in common with the silicon-based materials described herein (e.g., porosity, pore size, particle size, surface characteristics, bioerodibility, and resorbability) can also be used in the present invention. Examples of additional materials that can be used as porous support materials are bioerodible ceramics, bioerodible metal oxides, bioerodible semiconductors, bone phosphates, calcium phosphates (e.g., hydroxyapatite), other inorganic phosphates, porous carbon blacks, carbonates, sulfates, aluminates, borates, aluminosilicates, magnesium oxide, calcium oxide, iron oxide, zirconium oxide, titanium oxide, and other similar materials. Many of these porous materials can be prepared using techniques (e.g., molding, oxidation, drying, and surface modification) similar to those described above for preparing porous silicon-based support materials.

In certain embodiments, the therapeutic agent may be incorporated into the carrier material after the carrier material is fully formed. Alternatively, the therapeutic agent may be incorporated into the carrier material at one or more stages of its preparation. For example, the therapeutic agent may be introduced into the carrier material before the drying stage of the carrier material or after the carrier material has been dried, or both. In certain embodiments, the therapeutic agent may be introduced into the carrier material after a thermal oxidation step of the carrier material. In certain aspects, the therapeutic agent is introduced as a final step in the device fabrication.

More than one therapeutic agent may be incorporated into the device. In certain such embodiments, each therapeutic agent may be individually selected from small organic molecules and macromolecules such as proteins and antibodies. For example, the ocular implant may be impregnated with two therapeutic agents for treating glaucoma, or one therapeutic agent for treating macular degeneration and another agent for treating glaucoma.

In certain aspects, for example when both a small molecule therapeutic agent and a larger molecule therapeutic agent (such as a protein) are incorporated into the device, the therapeutic agent may be incorporated into the carrier material at different stages of device fabrication. For example, the small molecule therapeutic agent may be incorporated into the carrier material prior to the oxidation or drying step, while the large molecule therapeutic agent may be incorporated after the oxidation or drying step. Similarly, a plurality of different therapeutic agents of the same or different types may be introduced into the final carrier material in a different order or substantially simultaneously. When the carrier material comprises a single material or a combination of materials having multiple pore sizes, the larger therapeutic agent is preferably added to the carrier material before the smaller therapeutic agent is added, so as not to fill the larger pores with the smaller therapeutic agent and interfere with the adsorption of the larger therapeutic agent. For example, if the carrier material comprises a single material or a combination of materials having some well-defined pores of about 6nm in diameter (i.e., suitable for molecules having a molecular weight of about 14,000amu to 15,000 amu) and some well-defined pores of about 10nm in diameter (i.e., suitable for molecules having a molecular weight of about 45,000amu to 50,000 amu), the latter therapeutic agent (i.e., a therapeutic agent having molecules having a molecular weight of about 45,000amu to 50,000 amu) is preferably added to the carrier material prior to the addition of the smaller therapeutic agent (i.e., a therapeutic agent having molecules having a molecular weight of about 14,000amu to 15,000 amu). Alternatively and additionally, in embodiments where two different porous materials together comprise a device, each carrier material may be separately loaded with a different therapeutic agent and these carrier materials may then be combined to create a device.

The therapeutic agent can be incorporated into the carrier material as a compound or solution with one or more pharmaceutically acceptable excipients. The therapeutic agent may be formulated for administration in any suitable manner (such as in the form of an implant) suitable for subcutaneous, intramuscular, intraperitoneal or epidermal introduction or implantation into an organ (such as the eye, liver, lung or kidney). The therapeutic agents according to the present invention may be formulated for parenteral administration by injection (e.g., intraocular, intravenous, intravascular, subcutaneous, intramuscular) or infusion, or for oral administration.

In certain embodiments, the porous silicon-based carrier material is loaded with one or more therapeutic agents at a point of service (such as a physician's office or hospital) prior to administration of the implant. For example, the porous silicon carrier material can be loaded with the therapeutic agent shortly before administration, such as 24 hours or less before administration, 3 hours or less before administration, 2 hours or less before administration, 1 hour or less before administration, or 30 minutes or less before administration.

The carrier material may be in any suitable form prior to loading with the therapeutic agent, such as in the form of a dry powder or granules, or formulated, for example, as an aqueous slurry with a buffer solution or other pharmaceutically acceptable liquid. The therapeutic agent may be in any suitable form, such as in the form of a solution, slurry or solid (such as a lyophilizate), prior to loading into the carrier material. The carrier material and/or therapeutic agent may be formulated with other components such as excipients, preservatives, stabilizers or therapeutic agents (e.g., antibiotics).

In some embodiments, carrier material already loaded with biomolecules (such as proteins or antibodies) may be formulated (and packaged and/or distributed), while in other embodiments, carrier material or carrier material preparations that are substantially free of biomolecules (e.g., containing less than 5% biomolecules or less than 2% biomolecules) are formulated (and packaged and/or distributed), e.g., for combination with biomolecules at the time of administration.

In certain embodiments, the biomolecule is a fusion protein. Fusion proteins contain at least two polypeptide domains that are not normally contiguous in nature. For example, the polypeptide domains may be derived from different organisms or different genes. In some embodiments, one such domain has therapeutic activity, and another domain is useful for production or to improve pharmacokinetic properties. Common domains in fusion proteins include, but are not limited to, polyhistidine, Glu-Glu, Glutathione S Transferase (GST), thioredoxin, protein A, protein G, and immunoglobulin heavy chain constant region (Fc), Maltose Binding Protein (MBP), which are particularly useful for isolating fusion proteins by affinity chromatography. The fusion protein may also comprise an "epitope tag", which is typically a short peptide sequence available for specific antibodies, such as FLAG, influenza virus Hemagglutinin (HA), and c-myc tags. In certain embodiments, the fusion polypeptide may contain one or more modifications capable of stabilizing the polypeptide. For example, such modifications can increase the in vitro half-life of the polypeptide, increase the circulating half-life of the polypeptide, or reduce proteolytic degradation of the polypeptide. In certain embodiments, the linker region is located between two polypeptide domains. Methods for producing fusion proteins are well known. One method can, for example, produce a hybrid gene such that the host cell directly expresses the fusion protein. As another example, one or more polypeptide domains may be produced separately and then covalently linked using a chemical cross-linker.

The therapeutic agent may be formulated (and packaged and/or distributed) as a solution at a concentration >50mg/mL, such as >60mg/mL, such as >75 mg/mL. In an exemplary embodiment, the therapeutic agent is bevacizumab and the bevacizumab may be formulated at a concentration of >50mg/mL, such as >60mg/mL, such as >75mg/mL, for example in a phosphate buffered solution. The therapeutic agent may be formulated (and packaged and/or distributed) with a surfactant, wherein the therapeutic agent has a maximum concentration of 50 mg/mL. Protein fragments (such as antibody fragments) can be formulated (and packaged and/or distributed) into solutions at concentrations >10mg/mL, >15mg/mL, or >20 mg/mL.

The therapeutic agent may be formulated (and packaged and/or distributed) with stabilizers, excipients, surfactants, or preservatives. In particular embodiments, the therapeutic agent is formulated (and packaged and/or distributed) substantially free of any one or more of stabilizers, excipients, surfactants, and preservatives, e.g., containing less than 1mg/mL, or preferably less than 0.1mg/mL of stabilizers, excipients, surfactants, or preservatives. The formulation of the therapeutic agent may contain less than 1mg/mL of surfactant, such as less than 0.1mg/mL of surfactant.

In certain embodiments, the carrier material may be sold and/or distributed in any suitable form, such as a dry powder or granules, or preloaded into any part of a syringe (such as the barrel of a syringe or the needle of a syringe) in slurry form (e.g., combined with a biocompatible liquid such as an aqueous solution). The preloaded syringe may contain other components in addition to the carrier material, such as excipients, preservatives, therapeutic agents (e.g., antibiotics), or stabilizers. The preloaded syringe may contain biomolecules, such as proteins and/or antibodies, or may contain a solution that is substantially free of biomolecules (e.g., less than 5% biomolecules or less than 2% biomolecules).

In certain embodiments, the porous silicon-based carrier material is loaded with one or more therapeutic agents within the barrel of the injector. In certain embodiments, the carrier material is located within the barrel of the syringe described above or it may be drawn into the syringe from a separate container. Where the carrier material is located in a syringe, a solution containing one or more therapeutic agents may be drawn into the syringe, thereby contacting the carrier material. Alternatively, the carrier material may be drawn into the syringe after the therapeutic agent or solution thereof is drawn into the syringe barrel. After combining the components, the mixture can be allowed to incubate for a period of time to allow the therapeutic agent to be loaded into the pores of the carrier material. In certain embodiments, the mixture is incubated for about 3 hours or less, about 2 hours or less, or about 1 hour or less, e.g., about 30 minutes, about 20 minutes, about 10 minutes, or about 5 minutes.

In certain embodiments, a device such as an implant can comprise a coating that modulates the release of a therapeutic agent. For example, the device can be coated with an excipient (such as cocoa butter) to allow a desired release profile of the therapeutic agent from the device.

Application method

In certain embodiments, the device is used to prevent or treat a condition in a patient. The various embodiments provided herein generally provide for the local delivery of a therapeutically effective amount of a therapeutic agent, i.e., to the site of pain, disease, etc., in a patient. In certain embodiments, the devices of the present invention can be delivered to any site on or within the body of a patient. For example, the devices of the present invention may be used on the surface of the skin or eye, or may be implanted under the skin, within muscle, within an organ, near bone, within the eye, or any other location that would be beneficial for controlled release of a therapeutic agent. The implant may be administered intravitreally, subcutaneously, subconjunctivally, intraperitoneally, intramuscularly, or subretinally. In certain embodiments, the implants of the invention are delivered to the ocular surface or into the eye, such as within the uvea of the eye or within the vitreous of the eye.

In certain embodiments, the devices of the present invention are used to treat intraocular diseases, such as posterior ocular diseases. Exemplary intraocular diseases include glaucoma, age-related macular degeneration (such as wet age-related macular degeneration), diabetic macular edema, geographic atrophy, choroidal neovascularization, uveitis, diabetic retinopathy, retinal vascular disease, and other types of retinal degeneration.

In certain embodiments, the devices of the present invention are used to treat diseases on the surface of the eye. Exemplary diseases include viral keratitis and chronic allergic conjunctivitis.

In certain embodiments, the method of treating an ocular condition comprises placing the device on the surface of the eye or within the eye, such as the vitreous of the eye or the aqueous humor. In certain embodiments, the implant is injected or surgically inserted into the eye of the patient. In certain embodiments, the implant is injected into the eye of the patient, for example, within the vitreous of the eye. In certain embodiments, the implant is injected in the form of a composition. In certain embodiments, the device composition comprises a plurality of devices. The device composition may comprise devices having an average size between about 1 micron and about 500 microns. In certain embodiments, the composition comprises devices having an average device size between 5 microns and 300 microns, such as between about 5 microns and 100 microns.

In certain embodiments, the invention includes methods of loading a therapeutic agent into a porous silicon-based carrier material prior to administration to a patient (such as shortly before administration to a patient). The therapeutic agent(s) and silicon-based carrier material may be packaged together as part of a kit or obtained separately by a medical practitioner, for example. The one or more therapeutic agents may be obtained in the form of a solution (such as an aqueous or organic solution), in the form of a lyophilizate for reconstitution, or in any other suitable form.

The practitioner may introduce the one or more therapeutic agents into the carrier material in any suitable manner, such as by incubating the therapeutic agent and carrier material in a vial or in the barrel, trocar, or needle of a syringe. In particular embodiments, where the therapeutic agent is loaded onto the carrier material in the vial, the carrier material may be incubated with the one or more therapeutic agents or solutions thereof in the vial for a period of time, such as less than 24 hours, less than 2 hours, less than 1 hour, or even about 30 minutes or less.

In other embodiments, the carrier material is preloaded into the barrel of a syringe and one or more therapeutic agents or solutions thereof are drawn into the syringe to form a mixture with the carrier material. The mixture in the syringe may be incubated for a period of time, such as 30 minutes or less. In certain embodiments, the particles are sterilized at one or more stages during device preparation (e.g., immediately prior to administration or prior to loading of the syringe). In certain embodiments, any method suitable for sterilizing an implant may be used in the preparation of the implant.

In certain aspects, the devices of the present invention can be used to administer any therapeutic agent in a sustained manner to a patient in need thereof. The implants of the present invention are not limited to ophthalmic and intraocular use and may be used on any part of the body. For example, the implants of the invention can be used to subcutaneously administer therapeutic agents, similar to Norplant contraceptive devices. In other embodiments, the implants of the invention are used to administer biomolecules over a sustained period of time to treat chronic diseases, such as arthritis. For example, the implants of the invention can be used to deliver a therapeutic agent, such as etanercept or adalimumab, to a patient in need of such therapy. The implant of the present invention may be located anywhere within the body, such as intramuscularly. The implant may comprise a plurality of small particles, such as a plurality of particles of 500 microns or less. The implant may comprise larger particles (such as greater than 500 microns) or particles of one or more sizes greater than 1mm, such as greater than 10 mm.

The method of administering a therapeutic agent may comprise: a. providing a syringe pre-loaded with a porous silicon-based carrier material; b. contacting the carrier material with a therapeutic agent; administering a carrier material to the patient. The porous silicon-based carrier material may be preloaded into any part of a syringe, such as the barrel of the syringe, an insert between the needle and the barrel, or the needle of the syringe. The porous material may be preloaded into a portion of the syringe that may be removably connected with other portions of the syringe (e.g., a cartridge). For example, the porous silicon material may be preloaded into an insert that is removably connected between the barrel and the needle of the syringe, with the remaining syringe components being selected from any commercially available syringe components. In such embodiments, the insert may include one or more filters to prevent particles from exiting the insert, such as a filter proximate to the connection point of the barrel and the porous carrier material located between the filter and the syringe needle. The filter can be used to contain the carrier material while in contact with the therapeutic agent for loading the therapeutic agent into the pores of the carrier material. The filter can then be removed, inverted, bypassed, or bypassed in order to administer the loaded carrier material to the patient.

The porous, silicon-based material may be preloaded into the needle of a syringe, the opening of which may be blocked by one or more separable blockers or filters to prevent particles from leaving the needle until the desired time. The blocker may be detached before or after the carrier material is loaded with the therapeutic agent to allow the loaded carrier material to be administered to the patient, for example, through a needle. The preloaded needle may be removably connected to any commercially available syringe barrel or may be secured to the syringe barrel.

Step b of the method for administering the therapeutic agent may be performed by drawing the therapeutic agent into the syringe (such as drawing the therapeutic agent in the form of a mixture or solution into the syringe barrel). The therapeutic agent may be a small molecule or a biomolecule. The therapeutic agent may be released to the patient over a period of up to four months, six months, or even up to twelve months after administration. In some embodiments, the therapeutic agent is released to the patient over a time course of 1 month to 6 months.

In certain embodiments, the device is loaded in vivo by separately administering the carrier material and the therapeutic agent to the patient. First, a carrier material or therapeutic agent, or a formulation containing a carrier material or therapeutic agent, is administered to a patient. Next, the carrier material or therapeutic agent, or a formulation containing the carrier material or therapeutic agent, which is not delivered in the first step, is administered to the same site of the patient, thereby causing the therapeutic agent to adsorb into the pores of the carrier material. The therapeutic agent is adsorbed into the pores of the carrier material within the first minutes, hours, or days after the second step until adsorption of the therapeutic agent into the pores of the carrier material reaches equilibrium with desorption of the therapeutic agent from the carrier material into the surrounding environment (e.g., on the body surface or in the body of a patient). Thereafter, the device may release a therapeutically effective amount of the therapeutic agent for a period of time longer than the initial rebalancing period of time (e.g., hours, days, weeks, months, or years).

In certain embodiments, the implant is injected subcutaneously or surgically inserted subcutaneously. In other embodiments, the device is delivered to the patient intravenously or intra-articularly.

In some embodiments, the composition is administered orally. Oral administration may be used, for example, to deliver an active agent to the stomach, small intestine, or large intestine. Formulations for oral administration may be capsules, cachets, pills, tablets, lozenges (using a flavored basis, usually sucrose and acacia or tragacanth), powders, granules, and the like, each containing a predetermined amount of the active ingredient. Solid dosage forms for oral administration (capsules, tablets, pills, dragees, powders, granules, etc.) may comprise the device together with one or more pharmaceutically acceptable carriers such as sodium citrate or calcium hydrogen phosphate and/or any of the following: (1) fillers or extenders such as starch, lactose, sucrose, glucose, mannitol, and/or silicic acid; (2) binders such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose and/or acacia; (3) humectants, such as glycerol; (4) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates and sodium carbonate; (5) dissolution retarders, such as paraffin; (6) absorption promoters, such as quaternary ammonium compounds; (7) wetting agents such as cetyl alcohol and glyceryl monostearate; (8) adsorbents such as kaolin and bentonite; (9) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate and mixtures thereof; and (10) a colorant. In the case of capsules, tablets and pills, the pharmaceutical compositions may also comprise buffering agents. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar (milk sugar) as well as high molecular weight polyethylene glycols and the like. The oral composition may further comprise sweetening agents, flavoring agents, and preservatives.

In certain embodiments, multiple implants are delivered to the patient, such as two implants, three implants, four implants, or five implants or more. These implants may be substantially the same in size or composition or may be of different sizes, consist of different carrier materials, or carry different therapeutic agents. The plurality of implants may be administered to the patient simultaneously or over a period of time and to one or more sites of the patient's body.

In certain embodiments, the therapeutic agent is released from the device into the surrounding biological system for days, weeks, months, or years. In certain such embodiments, the time course of therapeutic agent release is selected from one day to two years, such as two weeks to about one year, such as about one month to about one year. The device may release the drug into the eye over a time course of 1 day to 12 months, such as 1 day to 6 months, such as 1 week to 3 months. In certain embodiments, the therapeutic agent is released within two years, such as within 18 months, within 15 months, within 1 year, within 6 months, within 3 months, or even within 2 months. In certain embodiments, device release of the therapeutic agent occurs in a controlled manner such that a large percentage of the total infused therapeutic agent is not released immediately or within a short time frame (e.g., within minutes or hours after administration). For example, if the desired drug delivery time is 2 months, the total infused therapeutic agent may be released at a rate of, for example, about 1/60 infused therapeutic agents per day. In certain embodiments, controlled release involves release of the therapeutic agent over a time period of, for example, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, or 8 months, wherein the amount of release of the therapeutic agent is linear relative to the entire delivery process. In some embodiments, there may be a burst release effect of the therapeutic agent shortly after administration, followed by a substantially constant release over a subsequent period of time. The burst effect may last, for example, for 1-10 days, during which a certain percentage of the loaded drug is released. After the burst, the remaining therapeutic agent may be constantly released for a certain period of time. For example, in certain embodiments, less than 10% of the therapeutic agent is released within the first day after administration, and another constant release of 50% occurs for subsequent 2-30 days, e.g., at a substantially constant release rate. In another exemplary embodiment, less than 10% of the therapeutic agent is released 5 days prior to administration, followed by a constant release of 50% of the therapeutic agent over the next 25 days. By substantially constant release is meant that the rate at which the device releases the therapeutic agent is substantially constant over a period of time.

In certain embodiments, the therapeutic agent begins to be released immediately after administration. In certain embodiments, the therapeutic agent is released over a time course of about 3 to 8 months, such as over a time course of about 6 months. In certain embodiments, the supplemental devices of the present invention are administered to the patient at appropriate intervals to ensure a substantially continuous therapeutic effect. For example, successive doses of an implant that releases drug for a period of six months may be administered once a half year, i.e., once every six months.

Pharmacokinetics can be determined by serum and vitreous analysis using ELISA.

In certain embodiments, the device may be completely or partially bioerodible within a biological system. In certain embodiments, the device may be resorbable by a biological system. In certain embodiments, the device is bioerodible and resorbable in a biological system. In certain embodiments, the carrier material may have a partial bioactivity such that the material is incorporated into living tissue. In some embodiments, the carrier material does not substantially mineralize or attract mineral deposits after implantation. For example, in some embodiments, the carrier device is substantially non-calcified when placed in situ in a site where calcification is not desired.

In certain embodiments, the device is bioerodible in a biological system. In certain embodiments, greater than about 80% of the carrier material will bioerode in a biological system, such as greater than about 85%, greater than about 90%, greater than about 92%, greater than about 95%, greater than about 96%, greater than about 97%, greater than about 98%, greater than about 99%, greater than 99.5%, or even greater than 99.9%. In certain embodiments, in the case of bioerosion of the support material, it is partially or completely resorbed.

In certain embodiments, the device may be substantially bioerodible over a time course of 1 week to 3 years. In certain embodiments, substantially bioerosion refers to erosion of greater than 95% of the carrier material. In certain embodiments, substantial bioerosion occurs over a time course of from about 1 month to about 2 years, such as from about 3 months to 1 year. In certain embodiments, substantial bioerosion occurs within about 3 years, such as within about 2 years, within about 21 months, within about 18 months, within about 15 months, within about 1 year, within about 11 months, within about 10 months, within about 9 months, within about 8 months, within about 7 months, within about 6 months, within about 5 months, within about 4 months, within about 3 months, within about 2 months, within about 1 month, within about 3 weeks, within about 2 weeks, within about 1 week, or even within about 3 days. In certain embodiments, in the case of bioerosion of the support material, it is partially or completely resorbed.

In certain embodiments, the extent of bioerosion can be assessed by any suitable technique used in the art. In exemplary embodiments, bioerosion is assessed by in vitro assays to identify degradation products or by in vivo histology and analysis. The kinetics of biodegradability of the porous carrier material can be assessed in vitro by analysing the concentration of the main degradation products in the relevant body fluids. For porous silicon-based carrier materials in the posterior part of the eye, for example, the degradation products may comprise orthosilicic acid, quantified, for example, by molybdenum blue assay, and the body fluid may be a simulated or actual vitreous body fluid. The kinetics of in vivo biodegradability can be determined by implanting a known amount of porous silicon-based material into the relevant body part and monitoring its persistence over time using histology in combination with, for example, standard microanalysis techniques.

Examples

Material

Quality standards for commercial porous silica

Example 1

To establish the relationship between protein size and the desired pore size for drug loading, the amount of surface area occupied by the protein when adsorbed in monolayer coverage was correlated with cumulative surface area/pore size data from Barrett-Joyner-Halenda (BJH) analysis of the nitrogen adsorption data. The point at which the protein adsorption surface area data is equal to the cumulative total surface area from the nitrogen adsorption analysis defines the minimum available pore size that facilitates adsorption loading. The data in table 1 provides the minimum pore radius that can be used for a range of protein sizes. Subtracting the protein hydrodynamic radius from the minimum pore radius yields the protein-pore difference, which is the minimum amount of additional pore size needed to allow protein access. For the range of proteins studied, the average protein-pore difference was 3.9 nm.

TABLE 1 correlation between protein size and pore accessibility

Example 2

Adsorption of bevacizumab to DavisilThe kinetics of (5 mg) was established by incubating 5mg of adsorbent with 25. mu.L of 25mg/mL bevacizumab in phosphate buffer pH6.2 (Table 2). After the specified equilibration time, 1.975mL of phosphate buffer was added to the suspension and mixed by inversion for no more than 30 seconds, followed by removal of the particles by filtration through a 0.2 μm filter. The amount of protein in the filtrate was quantified using BCA assay (Thermo Scientific, USA). The amount of protein adsorbed was calculated by subtracting the amount in the filtrate from the starting concentration. Table 2 provides adsorption kinetics for a range of particle sizes. 8.4am (D)₅₀) Results in 95.6% adsorption in 30 minutes, compared to 15.8am (D)₅₀) And 54.5am (D)₅₀) The particle sizes of (a) resulted in adsorption of 73.7% and 19.8%, respectively.

TABLE 2 adsorption kinetics of bevacizumab for adsorbents with increased particle size

Example 3

The adsorption isotherm was obtained by equilibrating 1mL egg white lysozyme (Sigma) in 50mM phosphate buffer pH6.2 at a concentration ranging from 270. mu.M to 1. mu.M with 5mg of adsorbent. After 16 hours, the residual amount of lysozyme in the equilibrium solution was quantified by UV spectroscopy at 280 nm. Then, the amount of lysozyme adsorbed onto the adsorbent was plotted against the equilibrium concentration. The lysozyme monolayer adsorption capacity and Langmuir adsorption coefficient (K) were estimated using a standard linear transformation method.

To measure the extent and rate of lysozyme release, the adsorbent matrix was equilibrated with egg white lysozyme (Sigma) in 50mM phosphate buffer pH6.2 at room temperature for 16 hours (Table 3). SiO 2mL release by time measurement₂Amount of lysozyme in saturated phosphate buffered saline (pH 7.4). At each time point, the samples were centrifuged at 16,300g, and 1mL of supernatant was removed and replaced with fresh media. Then, the amount of released lysozyme was quantified by high pressure liquid chromatography. The rate of lysozyme release correlates with the adsorbent pore size and also with the strength of the interaction between lysozyme and adsorbent as measured by the Langmuir adsorption coefficient (K).

TABLE 3 relationship between adsorbent pore size, Langmuir adsorption coefficient and lysozyme release rate.

Example 4

Lysozyme (egg white, Sigma) was purified by mixing 50. mu.L of a solution of 25mg/mL with 10mg of adsorbent was equilibrated and loaded onto the silica adsorbent with increased pore size. After 16 hours, 3.95mL of phosphate buffered saline (PBS; pH7.4) or SiO was added₂Saturated phosphate buffered saline and the suspension incubated at 37 ℃. At each time point, the particles were sedimented by centrifugation at 16,300g, 2mL of supernatant removed and replaced with 2mL of fresh medium. The amount of lysozyme in the dissolution medium was then quantified by RP-HPLC. Lysozyme release kinetics were determined by regression analysis of cumulative release versus square root time. The results are given in figure 2 and table 4.

The adsorption isotherm was obtained by equilibrating 1mL egg white lysozyme (Sigma) in 50mM phosphate buffer pH6.2 at a concentration ranging from 270. mu.M to 1. mu.M with 5mg of adsorbent. After 16 hours, the residual amount of lysozyme in the equilibrium solution was quantified by UV spectroscopy at 280 nm. Then, the amount of lysozyme adsorbed onto the adsorbent was plotted against the equilibrium concentration. The lysozyme monolayer adsorption capacity and Langmuir adsorption coefficient (K) were estimated using a standard linear transformation method.

In phosphate buffered saline and SiO₂Experiments were performed in saturated phosphate buffered saline to demonstrate that lysozyme release occurs by two mechanisms. By SiO₂The saturated phosphate buffered saline prevented the porous silica matrix from dissolving out. Thus, any release of lysozyme occurs by a desorption process. In phosphate buffered saline, lysozyme release results from a combination of matrix-related dissolution and desorption. The results in table 4 demonstrate that the lysozyme desorption release component increases simultaneously with the increase in the matrix pore size. The lysozyme desorption rate was assumed to be inversely proportional to the adsorption strength between lysozyme and the porous matrix as determined by the Langmuir coefficient.

TABLE 4 silica adsorbents in phosphate buffered saline and SiO₂Release Rate of Lysozyme Release in saturated phosphate buffered saline

^aThe desorption component percent of lysozyme is in SiO₂Release in saturated PBS/Release of lysozyme in PBS X100

Example 5

Bevacizumab was adsorbed onto a silica adsorbent with increased pore size by equilibrating 25 μ L of 25mg/mL solution with 5mg of adsorbent. After 16 hours, 1.975mL of phosphate buffered saline (ph7.4) was added and the suspension was incubated at 37 ℃. At each time point, the particles were removed by centrifugation at 16,300g, 1mL of supernatant removed and replaced with 1mL of fresh medium (fig. 3, table 5). The amount of bevacizumab in the dissolution medium was then quantified by micro BCA assay (Thermo Scientific, USA). Bevacizumab release kinetics were determined by regression analysis of cumulative release versus square root time. The results demonstrate that the release rate of bevacizumab increases with increasing pore size of the adsorbent.

TABLE 5 Bevacizumab Release Rate from silica adsorbent in phosphate buffered saline

Example 6

The kinetics of protein adsorption into porous silica of various pore sizes was established by incubating 5mg of adsorbent with 25. mu.L of a 25mg/mL protein solution in phosphate buffer, pH 6.2. After the specified equilibration time, 1.975mL of phosphate buffer was added to the suspension and mixed by inversion for no more than 30 seconds, followed by removal of the particles by filtration through a 0.2 μm filter. The amount of protein in the filtrate was quantified using BCA assay (Thermo Scientific, USA) for bevacizumab and RP-HPLC for lysozyme. The amount of protein adsorbed was calculated by subtracting the amount in the filtrate from the starting concentration. Tables 5a, 5b, 6a and 6b provide adsorption kinetics for a range of porous silica pore sizes and particle sizes.

For both lysozyme and bevacizumab, it is evident that the increase in matrix pore size results in a faster protein adsorption rate. The results in tables 6a and 6b also demonstrate that a reduction in particle size results in an increase in protein adsorption rate.

Table 5a. effect of pore size on lysozyme adsorption.

Table 5b. effect of pore size on normalized lysozyme loading.

^aThe lysozyme normalized loading (%) is the lysozyme adsorption amount (μ g/mg)/24-hour lysozyme adsorption amount × 100

Table 6a. effect of pore size and particle size on bevacizumab adsorption.

Table 6b. effect of pore size and particle size on normalized bevacizumab loading.

^aThe normalized loading (%) of bevacizumab is bevacizumab adsorption (μ g/mg)/24 h bevacizumab adsorption × 100

Equivalent forms

Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the compounds described herein and methods of use thereof. Such equivalents are considered to be within the scope of this invention and are covered by the following claims. Those skilled in the art will also recognize that all combinations of the embodiments described herein are also within the scope of the invention.

Claims (66)

1. A device comprising a bioerodible porous silicon-based carrier material, wherein the carrier material is impregnated with at least one macromolecular therapeutic agent.

2. The device of claim 1, wherein the carrier material is resorbable.

3. The device of claim 1 or 2, wherein the therapeutic agent is distributed in the volume of the carrier material.

4. The device of claim 3, wherein the therapeutic agent is distributed throughout substantially the entire volume of the carrier material.

5. The device of any one of claims 1 to 4, wherein the therapeutic agent is selected from the group consisting of proteins, peptides, antibodies, carbohydrates, polymers, and polynucleotides.

6. The device of claim 5, wherein the therapeutic agent is an antibody.

7. The apparatus of any of claims 1 to 6, wherein the silicon-based carrier material is amorphous.

8. The device of any one of claims 1 to 7, wherein the support material has a porosity of at least about 40%.

9. The device of claim 8, wherein the support material has a porosity of at least about 70%.

10. The device of any one of claims 1 to 7, wherein the support material has a porosity in a range of about 40% to about 80%.

11. The device of any one of claims 1 to 10, wherein the average pore size is in the range of 2-50 nm.

12. The device of claim 11, wherein the average pore size is in the range of 10-50 nm.

13. The device according to any one of claims 1 to 12, wherein the surface area of the support material is between 20 and 1000m²Between/g.

14. The device of claim 13, wherein the surface area of the support material is between 100 and 300m²Between/g.

15. The device of any one of claims 1 to 14, wherein the average width of the walls separating the pores in the support material is less than 5 nm.

16. The device of claim 15, wherein the average width of the walls is less than 3 nm.

17. The device according to any one of claims 1 to 16, wherein the length of the carrier material measured at its longest point is between 1 and 500 microns.

18. The device of claim 17, wherein the length of the carrier material at its longest point is between 2 and 100 microns.

19. The device of any one of claims 1-18, wherein the loading of the carrier material is less than 5% by weight based on the combined weight of the carrier material and therapeutic agent.

20. The device of any one of claims 1-18, wherein the loading of the carrier material is from about 5% to about 10%.

21. The device of any one of claims 1-18, wherein the loading of the carrier material is from about 10% to about 20%.

22. The device of any one of claims 1-18, wherein the loading of the carrier material is from about 20% to about 30%.

23. The device of any one of claims 1-18, wherein the loading of the carrier material is from about 30% to about 40%.

24. The device of any one of claims 1-18, wherein the loading of the carrier material is from about 40% to about 50%.

25. The device of any one of the preceding claims, further comprising a second therapeutic agent impregnated in the carrier material.

26. A method for preparing the device of any one of the preceding claims, comprising contacting a porous silicon-based carrier material with a therapeutic agent.

27. The method of claim 26, wherein the average pore size of the carrier material is selected to allow entry of the therapeutic agent and to allow controlled release of the therapeutic agent in a biological medium for at least about 3 days.

28. The method of claim 27, wherein the average pore size is from about 15nm to about 40nm and the therapeutic agent has a molecular weight from about 100,000 to about 200,000 amu.

29. The method of claim 27, wherein the average pore size is from about 25nm to about 40nm and the therapeutic agent has a molecular radius from about 6 to about 8 nm.

30. The method of claim 27, wherein the average pore size is from about 2nm to about 10nm and the therapeutic agent has a molecular weight from about 5,000 to about 50,000 amu.

31. A method of treating or preventing a condition in a patient comprising administering to the patient the device of any one of claims 1-25.

32. The method of claim 31, wherein the device is applied to the surface of the patient's skin or eye.

33. The method of claim 31, wherein the device is administered subconjunctivally, intraperitoneally, intramuscularly, intravitreally, subcutaneously, or subretinally.

34. The method of claim 31, wherein the device is administered into the eye.

35. The method of claim 34, wherein the device is administered within the aqueous humor of the eye.

36. The method of claim 34, wherein the device is administered intravitreally of the eye.

37. The method of claim 31, wherein the disorder is selected from an ocular disorder.

38. The method of claim 37, wherein the disorder is selected from the group consisting of glaucoma, macular degeneration, diabetic macular edema, geographic atrophy, and age-related macular degeneration.

39. The method of any one of claims 31-38, wherein the device releases drug into the eye over a time course of 1 day to 12 months.

40. The method of claim 39, wherein the device releases the therapeutic agent over a time course of 1 month to 6 months.

41. The method of any one of claims 31-40, wherein the porous silicon-based carrier material is contacted with a solution comprising the therapeutic agent.

42. A composition comprising a bioerodible porous silicon-based support material, wherein the support material has at least two different pore populations, wherein the difference between the average pore diameters of the at least two different pore populations is at least about 5 nm.

43. The composition of claim 42, wherein the difference between the average pore sizes of the at least two different pore populations is between about 5nm and about 50 nm.

44. The composition of claim 42 or 43, wherein the ratio of the average pore diameters of the at least two different pore populations is at least about 1.5 to 1.

45. The composition of any one of claims 42 to 44, wherein the support material has two different pore populations.

46. A composition comprising a first plurality of bioerodible porous silicon-based particles having pores of a first average pore size and a second plurality of bioerodible porous silicon-based particles having pores of a second average pore size, wherein the first average pore size is at least 1.5 times the second average pore size.

47. A composition comprising a biomolecule loaded into pores of a porous silicon-based carrier material, wherein the biomolecule within the carrier material has a half-life at room temperature that is at least twice as long as the half-life of the biomolecule without the carrier material under the same conditions.

48. The composition of claim 47, wherein the half-life of the biomolecule within the carrier material is equal to or greater than 10 times the half-life of the biomolecule without the carrier material under the same conditions.

49. A composition comprising a biomolecule loaded into pores of a porous silicon-based carrier material, wherein the lifetime of the biomolecule within the carrier material is at least twice as long as the lifetime of the biomolecule without the carrier material under the same conditions.

50. The composition of claim 49, wherein the useful life of the biomolecule within the carrier material is at least 10 times as long as the useful life of the biomolecule without the carrier material under the same conditions.

51. The composition of claim 49 or 50, wherein the biomolecule within the carrier material is stable for at least 6 months at 25 ℃.

52. The composition of any one of claims 47-51, wherein the biomolecule is an antibody.

53. A syringe preloaded with a composition comprising a porous silicon-based carrier material, wherein said composition comprises less than 2% biomolecules.

54. The syringe of claim 53, wherein the composition is substantially free of biomolecules.

55. The syringe of claim 53 or 54, wherein the composition comprises an aqueous solution.

56. The syringe of any one of claims 53 to 55, wherein the syringe needle is pre-loaded with the silicon-based carrier material.

57. The syringe of any one of claims 53 to 55, wherein a removable insert in the syringe is loaded with the silicon-based carrier material.

58. The syringe of any one of claims 53 to 57, wherein the syringe further comprises one or more filters.

59. A method of administering a therapeutic agent comprising:

a) providing a syringe pre-loaded with a porous silicon-based carrier material;

b) contacting the carrier material with a therapeutic agent; and

c) administering the carrier material to a patient.

60. The method of claim 59, wherein step b is performed by drawing the therapeutic agent into the syringe.

61. The method of claim 59 or 60, wherein the therapeutic agent is a small molecule.

62. The method of any one of claims 59 to 61, wherein the therapeutic agent is released to the patient over a time course of up to four months.

63. The method of any one of claims 59 to 62, wherein the porous silicon-based carrier has a pore-therapeutic agent differential selected from about 3.0 to about 5.0 nm.

64. A method of loading a protein into the pores of a porous silicon-based carrier material, comprising: selecting a porous silicon-based carrier having pore sizes dimensioned to allow loading of a single protein into the pore such that opposing sides of the protein engage opposing sides of the pore.

65. The method of claim 64, wherein the porous silicon-based carrier has a pore-therapeutic agent differential selected from about 3.0 to about 5.0 nm.

66. A method of controlled release delivery of a therapeutic agent comprising:

a) administering to a patient a bioerodible porous silicon-based carrier material; and

b) administering a therapeutic agent to the same location of the patient's body.