FORMULATIONS AND METHODS OF USE

BACKGROUND

The development of new delivery vehicles that deliver certain therapeutic agents to a patient (e.g., targeted to a particular tissue or cell type or targeted to a specific diseased tissue, but not normal tissue), or that control the release of drugs in a subject has long been recognized as beneficial. For example, “small particle therapies” (e.g., microparticles, nanoparticles, liposomes, and micelles) have proven useful for such purposes, due to the small size, and their ability to evade recognition in the body allowing for targeted and controlled delivery, while remaining stable for an effective amount of time. A common disadvantage associated with small particle therapies is that after intravenous administration they are rapidly cleared from the bloodstream, which limits the duration of the therapeutic effect.

Surface modification of such small particles through the association or coating of the particles with suitable potentiating agents has been shown to modify their physicochemical characteristics and increase the stability and in vivo performance. This surface modification, for example, increases the hydrodynamic size of the particle and reduces renal clearance and/or reduces recognition and clearance or inactivation by the immune system, and results in improved pharmacokinetic and pharmacodynamic properties. One type of potentiating agent is poly(ethylene glycol) and the associated surface modification technique is referred to as “PEGylation.” PEGylation is a useful, effective strategy for reducing biospecific interactions for pharmaceuticals.

Because the shelf-life of small particle therapies is often limited, these particles may be lyophilized to a dry, stable state. Lyophilization, which is sometimes referred to as “freeze-drying,” is an attractive method to enhance the physico-chemical stability of small particles. Lyophilization has become a preferred method because it enables moisture removal at relatively low temperatures under sterile conditions. Furthermore, the technique imparts the valuable product attribute of rapid reconstitution with a suitable reconstitution reagent at the point of use.

In a typical lyophilization process, multiple vials containing a liquid drug formulation are loaded on temperature-controlled shelves within a sterile chamber and cooled to low temperatures until completely solidified. FIG. 1 illustrates a typical freezing process. A conventional lyophilization process lasts about 72 hours. Following the thermal treatment (freezing step), chamber pressure is reduced and shelf temperature is adjusted to enable removal of the frozen solvent (drying) through sublimation in a step termed “primary drying.” When sublimation is complete, the shelf temperature is raised for a “secondary drying” to remove additional unfrozen solvent still bound to the solid product. When sufficient solvent has been removed, the drying process is concluded by stoppering the vials or bottles in the chamber, generally under a sub-ambient pressure of inert gas. The final dry product is called a “lyophilized preparation” (or also a “cake”) and usually occupies roughly the same volume as Initial liquid because of its high porosity. Lyophilization produces certain types of mechanical stress that can damage small particles. For example, such stresses can alter the desired conformation of the small particles or deform the small particles. Following lyophilization and reconstitution, the size of small particles can change, or the small particles may aggregate, which reduces the effectiveness of the small particles. To combat these problems, lyoprotectants may be added to the liquid formulation to protect small particles from stress and damage during lypophilization. Conventional lyoprotectants are typically added in relatively large concentrations.

The lyophilization of small particle therapeutics is time-consuming and challenging. The lyophilization of small particles that have been subjected to a surface modification with a potentiating agent, e.g., PEGylated particles, is challenging because of the tendency of such particles to agglomerate. Particle aggregation and fusion, both during and after lyophilization, is frequently observed even in the presence of common lyoprotectants such as sugars (e.g., sucrose, trehalose, mannose). Further, more complex lyoprotectants, such as polysaccharides like dextrans, have a well-known incompatibility with PEG. It is believed that this incompatibility causes a phase separation between the dextran and the PEG, which may cause the sugar molecules to no longer act as a stabilizer resulting in aggregation of the particles. Similar stability problems occur during freeze-drying of solutions of PEGylated nanoparticles and excipients that are incompatible with PEG. Another view is that this aggregation may be the result of the crystallization of PEG polymer chains upon the surfaces of the particles upon freezing. Aggregation is particularly problematic because it can reduce effectiveness or render the small particles unsuitable for their intended use. Aggregation of PEGylated nanoparticles during lyophilization tends to increase as the concentration of the PEGylated nanoparticles increases in a liquid formulation that is to be lyophilized. It can also be difficult to resuspend lyophilized preparations of PEGylated nanoparticles to produce a solution that contains a high concentration of PEGylated nanoparticles. The ability to lyophilize liquid formulations that contain a high concentration PEGylated nanoparticles, and to resuspend the resulting lyophilized preparations to produce a solution that contains a high concentration of PEGylated nanoparticles, is desirable to allow administration of the PEGylated nanoparticles in convenient ways, for example, intravenously, e.g., as an intravenous bolus.

A need exists for improved lyophilized preparations of PEGylated nanoparticles.

SUMMARY

Invention relates to the production, use and administration of small particulates, liposomes, and micelles for therapeutic agent delivery. In particular, at least in some embodiments, the present invention relates to the production and lyophilization of PEGylated nanoparticles, microparticles, micelles, and liposomes for use and administration into a subject.

In some aspects, the invention is a liquid (e.g., pre-lyophilization, or resuspended after lyophilization) or lyophilized formulation that comprises a particulate construct (e.g., nanoparticle or microparticle) and a lyoprotectant that comprises a cyclic oligosaccharide. The particulate construct comprises a polymer, a therapeutic agent and a potentiating agent. The therapeutic agent and the potentiating agent are associated with the polymer, and preferable are covalently bonded to the polymer, directly or through a suitable linker moiety. If desired, the particulate construct can further comprise one or more additional components, such as a stabilizing polymer, an excipient, a surfactant, and the like. If desired, the particulate construct can comprises a targeting ligand.

The polymer can be, for example, a polyester, a polysaccharide, a polyamide, a polyether, a polycarbonate, a polyacrylate, a polylactic acid, a polyglycolic acid, a polydioxanone, a poly-(lactide-co-glycolide), a polyethylenimine, a copolymer of methoxy-poly(ethylene glycol)-block-poly(lactide-co-glycolide), a chitin, a chitosan, a poly(glycolide), a poly(ε-caprolactone), a poly(hydroxy ester ether), a poly(hydroxybutyrate), a poly(anhydride), a poly(orthoester), a polyamine acids), a poly(ethylene oxides), a poly(phosphazene), a poly etheresters, a polyester amides, a polyamide, derivatives, copolymers, blends, and other combinations thereof. In some embodiments, the polymer is a biodegradable block copolymer with a weight in the range of from about 1 kDa to about 21 kDa. For example, the biodegradable block copolymer can comprises poly-(lactide-co-glycolide).

The cyclic oligosaccharide can comprise a polysaccharide moiety selected from the group consisting of: an α-cyclodextrin, a β-cyclodextrin, a 2-hydroxypropyl-β-cyclodextrin, a β-cyclodextrin sulfobutylether, any derivative thereof, and any combination thereof. In some embodiments, the polysaccharide is incorporated within the backbone of a polymer. If desired, at least one occurrence of the cyclic oligosaccharide can be oxidized.

In preferred aspects, the formulation further comprises a wetting agent, such as a monosaccharide, a disaccharide, a surfactant, an amino acid, any derivative thereof, and any combination thereof. In some preferred embodiments, the wetting agent is a disaccharide selected from the group consisting of sucrose, trehalose, lactose and combinations thereof. If desired, the wetting agent can also include a monosaccharide, such as glucose, fructose, galactose, xylose, ribose, and combinations thereof. When the formulation contains a lyoprotectant that comprises a cyclic oligosaccharide and also contains a wetting agent, the ratio of cyclic oligosaccharide to the wetting agent (w/w) can be about 0.5:1.5 to about 1.5:0.5 and/or the ratio of cyclic oligosaccharide plus wetting agent to polymer (w/w) can be about 1:1 to about 10:1. Sucrose is a preferred wetting agent.

In some embodiments, a therapeutic agent and/or potentiating agent is bonded to the polymer through a linker. In such embodiments, the linker can be, for example, an alkanoate linker, a PEG-based linker, a linker that comprises a disulfide bond, a self-immolative linker, an amino acid, a peptide, a glutamic acid, a branched glutamic acid, a polyglutamic acid, any derivative thereof, and any combination thereof. In preferred aspects, the therapeutic agent and potentiating agent are bonded to the polymer, and the particulate construct is a dual conjugate nanoparticle or dual conjugate microparticle.

The therapeutic agent comprises a drug, such as a cancer drug, an anti-inflammatory drug, a thrombolytic drug, a cardiovascular drug, an anti-anginal drug, a diuretic drug, a antihistamine drug, a hormone-containing drug, an antibiotic drug, a pain reliever drug, an anti-diarrheal drug, an antiemetic, an anti-metabolite drug, an immunomodulator drug, a radiation drug, a chemotherapy drug, an any derivative thereof and any combination thereof. In particular embodiments, the therapeutic agent is a taxane drug. The therapeutic agent is generally present in a range of about 5% to about 20% (w/w) of the particulate construct. In some embodiments, the ratio of therapeutic agent to polymer (e.g., biodegradable block copolymer) is in the range of from about 85:15 to about 55:45 (w/w).

The potentiating agent generally comprises a hydrophilic polymer, such as a poly(alkylene glycol), any derivative thereof, or any combination thereof. Particular examples of hydrophilic polymers include polyethylene glycol, polypropylene glycol, polybutylene glycol, derivatives thereof, and combinations thereof.

In some aspects, the particulate construct further comprises a stabilizing polymer such as a poly(vinyl alcohol), a poly(vinyl pyrrolidone), a poly(vinyl acetate), a crown ether, any derivative thereof, and any combination thereof.

Liquid formulations of the invention, such as a resuspended lyophilized preparation, comprise a solvent, that is preferably physiologically acceptable, such as an aqueous solvent, an organic solvent, and any combination thereof. In some embodiments, the organic solvent comprises an organic compound selected from the group consisting of: acetone, methanol, ethanol, tert-butylmethyl ether, heptane, dichloromethane, dimethylformamide, acetylnitrile, 1-propanol, 1-butanol, formic acid, acetic acid, formamide, tetrahydrofuran, methyl ethyl ketone, ethyl acetate, acetonitrile, dimethylsufoxide, hexane, benzene, methyl ether, carbon tetrachloride, methylene chloride, butyl acetate, propyl acetate, any derivative thereof, and any combination thereof. When the solvent is an aqueous-organic solvent mixture the water:organic solvent ratio can be from about 1:1 to about 1:10 (v/v). In preferred aspects, the liquid formulation comprises a polymer concentration of at least about 30 mg/mL, more preferably at least about 70 mg/mL. When the liquid formulation is a resuspended lyophilized preparation, it is preferred that particles in the liquid formulation have a Z-average diameter, poly-dispersity index, and Dv₉₀that differ from the Z-average diameter, poly-dispersity index, and Dv₉₀of the particles in the formulation that was lyophilized to produce said lyophilized preparation by no more than about 20%.

In more particular aspects, the invention is a liquid (e.g., pre-lyophilization, or resuspended after lyophilization) or lyophilized formulation that comprises a particulate construct (e.g., nanoparticle or microparticle), and a lyoprotectant that comprises a cyclodextrin, any derivative thereof, and any combination thereof, and a wetting agent selected from the group consisting of a monosaccharide, a disaccharide, a surfactant, an amino acid, any derivative thereof, and any combination thereof. In some embodiments, the lyoprotectant is 2-hydroxypropyl-β-cyclodextrin. In these embodiments or in alternative embodiments, the wetting agent is a disaccharide selected from the group consisting of sucrose, trehalose, lactose and combinations thereof. Preferably, the wetting agent is sucrose. The ratio of lyoprotectant to wetting agent (w/w) is preferably about 0.7:1.3 to about 1.3:0.7. The ratio of lyoprotectant plus wetting agent to polymer (w/w) is preferably about 1:1 to about 3:1.

One embodiment provides a liquid formulation comprising a lyoprotectant that comprises a cyclic oligosaccharide; and a particulate construct, the particulate construct comprising a polymer composition; a therapeutic agent; and a potentiating agent, wherein the therapeutic agent and the potentiating agent are associated with the polymer composition.

In some embodiments the particulate construct is a dual conjugate nanoparticle.

In some embodiments the particulate construct comprises an additional additive selected from the group consisting of: a stabilizing polymer, an excipient, a surfactant, a derivative thereof, and any combination thereof.

In some embodiments the stabilizing polymer comprises a polymer selected from the group consisting of: a poly(vinyl alcohol), a poly(vinyl pyrrolidone), a poly(vinyl acetate), a crown ether, any derivative thereof, and any combination thereof.

In some embodiments the particulate construct comprises a polymer selected from the group consisting of: a polyester, a polysaccharide, a polyamide, a polyether, a polycarbonate, a polyacrylate, a polylactic acid, a polyglycolic acid, a polydioxanone, a poly-(lactide-co-glycolide), a polyethylenimine, a copolymer of methoxy-poly(ethylene glycol)-block-poly(lactide-co-glycolide), a chitin, a chitosan, a poly(glycolide), a poly(ε-caprolactone), a poly(hydroxy ester ether), a poly(hydroxybutyrate), a poly(anhydride), a poly(orthoester), a poly(amino acids), a poly(ethylene oxide), a poly(phosphazene), a poly etherester, a polyester amide, a polyamide, derivatives, copolymers, blends, and other combinations thereof.

In some embodiments the particulate construct comprises a biodegradable block copolymer with a weight in the range of from about 1 kDa to about 21 kDa.

In some embodiments the therapeutic agent to biodegradable block copolymer ratio is in the range of from about 85:15 to about 55:45 by weight.

In some embodiments the particulate construct has a Dv₉₀of less than about 200 nm.

In some embodiments the particulate construct has a weight in the range of from about 7 kDa to about 30 kDa.

In some embodiments the particulate construct is a nanoparticle or a microparticle.

In some embodiments the particulate construct comprises a targeting ligand.

In some embodiments the potentiating agent is covalently bonded to the polymer composition.

In some embodiments the potentiating agent comprises a hydrophilic polymer.

In some embodiments the hydrophilic polymer comprises a polymer selected from the group consisting of: a polyethylene glycol, a poly(alkylene glycol), a poly(propylene glycol), a poly(butylene glycol), any derivative thereof, and any combination thereof.

In some embodiments the potentiating agent comprises a compound selected from the group consisting of: a polysorbate, an enzyme, a peptide, a lecithin, a polysaccharide, a phospholipid analog, a poly(phosphazene), a poloxamer, a poly(oxyethylene ester), a poly(vinylpyrrolidone), a poly(vinyl alcohol), a glycolipid, a polyol, a salt, a crown ether, a specific nanoparticle-aptamer bioconjugate, any derivative thereof and any combination thereof.

In some embodiments the potentiating agent protects the particulate construct under biological conditions and/or increases the solubility of the particulate construct.

In some embodiments the cyclic oligosaccharide comprises a cyclodextrin moiety selected from the group consisting of: an α-cyclodextrin, a β-cyclodextrin, a 2-hydroxypropyl-β-cyclodextrin, a β-cyclodextrin sulfobutylethers sodium, any derivative thereof, and any combination thereof.

In some embodiments the lyoprotectant further comprises a polymer.

In some embodiments an oligosaccharide is incorporated within a backbone of the polymer.

In some embodiments at least one occurrence of the cyclic oligosaccharide is oxidized.

In some embodiments the lyoprotectant is added in a ratio from about 0.75:1 to about 3:1 by weight of the lyoprotectant:liquid formulation.

In some embodiments the polymer composition to lyoprotectant ratio is in the range of from about 0.75:1 to about 15:1 by weight.

In some embodiments at least one therapeutic agent is covalently bonded to the polymer composition.

In some embodiments the therapeutic agent is attached to the polymer composition through a linker.

In some embodiments the linker comprises a linking compound selected from the group consisting of: an alkanoate linker, a PEG-based linker, a linker that comprises a disulfide bond, a self-immolative linker, an amino acid, a peptide, a glutamic acid, a branched glutamic acid, a polyglutamic acid, any derivative thereof, and any combination thereof.

In some embodiments the therapeutic agent comprises a taxane drug.

In some embodiments the therapeutic agent comprises a drug selected from the group consisting of: a cancer drug, an anti-inflammatory drug, a thrombolytic drug, a cardiovascular drug, an anti-anginal drug, a diuretic drug, an antihistamine drug, a hormone-containing drug, an antibiotic drug, a pain reliever drug, an anti-diarrheal drug, an antiemetic, an anti-metabolite drug, an immunomodulator drug, a radiation drug, a chemotherapy drug, any derivative thereof, and any combination thereof.

In some embodiments the therapeutic agent is included in a range of about 5% to about 20% by weight of the particulate construct.

In some embodiments the liquid formulation comprises a physiologically acceptable liquid selected from the group consisting of: an aqueous liquid, an organic liquid, and any combination thereof.

In some embodiments the organic liquid comprises an organic solvent selected from the group consisting of: acetone, methanol, ethanol, tert-butylmethyl ether, heptane, dichloromethane, dimethylformamide, acetylnitrile, 1-propanol, 1-butanol, formic acid, acetic acid, formamide, tetrahydrofuran, methyl ethyl ketone, ethyl acetate, acetonitrile, dimethylsufoxide, hexane, benzene, methyl ether, carbon tetrachloride, methylene chloride, butyl acetate, propyl acetate, any derivative thereof, and any combination thereof.

In some embodiments the liquid formulation has an organic solvent:water ratio in the range of from about 1:1 to about 1:10 by volume.

In some embodiments the liquid formulation comprises a reconstitution reagent.

One embodiment provides a lyophilized preparation that comprises a particulate construct, the particulate construct comprising a polymer composition, a therapeutic agent, and a potentiating agent; wherein the therapeutic agent and the potentiating agent are associated with the polymer composition; and lyoprotectant that comprises a cyclic oligosaccharide.

Some embodiments further comprise 0.5% water or less by weight of the lyophilized preparation by weight of the lyophilized preparation.

In some embodiments the lyophilized preparation is substantially free of aggregates.

In some embodiments the particulate construct is a dual conjugate nanoparticle.

Some embodiments further comprise a reconstitution reagent.

In some embodiments the particulate construct comprises a polymer selected from the group consisting of: a polyester, a polysaccharide, a polyamide, a polyether, a polycarbonate, a polyacrylate, a polylactic acid, a polyglycolic acid, a polydioxanone, a poly-(lactide-co-glycolide), a polyethylenimine, a copolymer of methoxy-poly(ethylene glycol)-block-poly(lactide-co-glycolide), a chitin, a chitosan, a poly(glycolide), a poly(ε-caprolactone), a poly(hydroxy ester ether), a poly(hydroxybutyrate), a poly(anhydride), a poly(orthoester), a poly(amino acids), a poly(ethylene oxides), a poly(phosphazene), a poly etheresters, a polyester amides, a polyamide, derivatives, copolymers, blends, and other combinations thereof.