POLYMERS

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the priority benefit of U.S. Provisional Application Serial No. 62/851,066, filed May 21, 2019, which is hereby incorporated by reference in its entirety.

FIELD

[0002] The present disclosure relates to methods of preparing microspheres (e.g, single emulsion microspheres) comprising a therapeutic compound or pharmaceutically acceptable salt thereof and a poly(lactic-co-glycolic acid) (PLGA) polymer comprising at least one hydrophobic terminus, as well as microspheres and pharmaceutical compositions related thereto.

BACKGROUND

[0003] Drug formulations such as microspheres are important for modulating the pharmacokinetic properties of the drug, such as release of the active compound. A variety of microspheres and methods of microsphere preparation have been described. See, e.g., U.S. Pat. Nos. 5,639,480 and 8,916,196. Double emulsion, water/oil/water microspheres have often been used for a variety of hydrophilic drug compounds. These are typically formed by emulsifying an aqueous solution of the hydrophilic drug with a solution of a polymer in organic solvent by high shear mixing (e.g, 20,000 rpm), generating an unstable water/oil emulsion. This unstable emulsion is then usually further emulsified in water, leading to a water/oil/water double emulsion that is then hardened by solvent exchange, and lyophilized into dry microspheres.

[0004] However, this high shear, high energy process is difficult to control or reproduce, leading to a heterogeneous preparation of double emulsion microspheres. One example of such a drug product is SANDOSTATIN® LAR depot (octreotide acetate). This drug is formulated in a double emulsion microsphere for a long release, thereby decreasing the frequency of administration. However, SANDOSTATIN® LAR is notoriously difficult to administer by injection; it frequently blocks flow through the needle (e.g, low

syringeability), requiring thicker needles and leading to more painful injections for the patient. SANDOSTATIN® LAR is also characterized by low loading of the octreotide in the microspheres, necessitating a larger volume for injection that also causes significant pain during injection and can potentially lead to formation of painful nodules at the injection site. This further precludes subcutaneous injection in favor of more painful intramuscular injection. Finally, double emulsion microspheres are difficult to

manufacture, requiring costly investment and poor process control.

[0005] As such, a demand exists for improved microsphere formulations suitable for different active pharmaceutical ingredients (APIs) and characterized by improved pharmacokinetic properties, increased loading, a more uniform size distribution of particles, and easier, well-controlled manufacturing.

[0006] All references cited herein, including patent applications, patent publications, and non-patent literature are herein incorporated by reference in their entirety, as if each individual reference were specifically and individually indicated to be incorporated by reference.

SUMMARY

[0007] To meet these and other demands, provided herein are methods of preparing a plurality of single emulsion microspheres. In some embodiments, the methods comprise combining a first solvent and a therapeutic compound or pharmaceutically acceptable salt thereof to form a first mixture; combining a second solvent and a polymer to form a second mixture; combining the first and second mixtures to form a feed; dispersing the combined first and second mixtures into an aqueous continuous phase to form a plurality of droplets; and hardening the plurality of droplets to form the plurality of single emulsion

microspheres. In some embodiments, the polymer comprises poly(lactic-co-glycolic acid) (PLGA) comprising at least one hydrophobic terminus. In some embodiments, the polymer has a molecular weight of at least 7 kD. In some embodiments, at least 90% of the microspheres of the plurality produced are between 20 pm and 42 pm in diameter.

[0008] In some embodiments, the polymer has a molecular weight of less than 54 kD.

In some embodiments, the polymer has a molecular weight between 7 kD and 17 kD. In some embodiments, the polymer has a molecular weight between 38 kD and 54 kD. In some embodiments, the polymer has a ratio of lactide:glycolide of 50:50. In some embodiments, the second solvent is selected from the group consisting of dichloromethane (DCM), DMA, and DMF. In some embodiments, the second solvent comprises DCM. [0009] In some embodiments, the therapeutic compound comprises a glucocorticoid. In some embodiments, the therapeutic compound comprises triamcinolone. In some embodiments, the first solvent comprises dimethylacetamide (DMA). In some

embodiments, the feed produced by combining the first and second mixtures comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least 400mg/mL. In some embodiments, the polymer has a molecular weight between 7 kD and 17 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 22 pm and 34 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 4% by total weight of the microspheres. In some embodiments, the polymer has a molecular weight between 38 kD and 54 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 20 pm and 32 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 6% by total weight of the microspheres.

[0010] In some embodiments, the therapeutic compound comprises an anesthetic. In some embodiments, the therapeutic compound comprises bupivacaine. In some

embodiments, the first solvent comprises benzyl alcohol. In some embodiments, the feed produced by combining the first and second mixtures comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least 400mg/mL. In some embodiments, the polymer has a molecular weight between 7 kD and 17 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 22 pm and 36 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 5% by total weight of the microspheres. In some embodiments, the polymer has a molecular weight between 38 kD and 54 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 24 pm and 42 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 9% by total weight of the microspheres.

[0011] In some embodiments, the methods further comprise adjusting the pH of the aqueous continuous phase into which the feed is dispersed. In some embodiments, the pH of the aqueous continuous phase is greater than 7, greater than 7.5, or about 8.0. In some embodiments, the methods further comprise, e.g., after forming the plurality of droplets and prior to hardening the plurality of droplets to form the plurality of single emulsion microspheres, washing the plurality of microspheres in a second aqueous continuous phase.

[0012] Further provided herein is a plurality of microspheres produced by the method according to any one of the above embodiments. [0013] Further provided herein is a plurality of single emulsion microspheres, wherein each microsphere of the plurality comprises: a therapeutic compound or pharmaceutically acceptable salt thereof; and a polymer comprising poly(lactic-co-glycolic acid) (PLGA) comprising at least one hydrophobic terminus. In some embodiments, the polymer has a molecular weight of at least 38 kD. In some embodiments, at least 90% of the

microspheres of the plurality are between 20 pm and 42 pm in diameter.

[0014] In some embodiments, the polymer has a molecular weight of less than 54 kD.

In some embodiments, the polymer has a molecular weight between 7 kD and 17 kD. In some embodiments, the polymer has a molecular weight between 38 kD and 54 kD. In some embodiments, the polymer has a ratio of lactide:glycolide of 50:50.

[0015] In some embodiments, the therapeutic compound comprises a glucocorticoid. In some embodiments, the therapeutic compound comprises triamcinolone. In some embodiments, the polymer has a molecular weight between 7 kD and 17 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 22 pm and 34 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 4% by total weight of the microspheres. In some embodiments, the polymer has a molecular weight between 38 kD and 54 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 20 pm and 32 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 6% by total weight of the microspheres.

[0016] In some embodiments, the therapeutic compound comprises an anesthetic. In some embodiments, the therapeutic compound comprises bupivacaine. In some

embodiments, the polymer has a molecular weight between 7 kD and 17 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 22 pm and 36 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 5% by total weight of the microspheres. In some embodiments, the polymer has a molecular weight between 38 kD and 54 kD. In some embodiments, at least 90% of the microspheres of the plurality are between 24 pm and 42 pm in diameter. In some embodiments, the therapeutic compound or salt is greater than 9% by total weight of the microspheres.

[0017] Further provided herein are pharmaceutical compositions comprising the single emulsion microspheres or plurality of microspheres according to any one of the above embodiments.

[0018] It is to be understood that one, some, or all of the properties of the various embodiments described herein may be combined to form other embodiments of the present invention. These and other aspects of the invention will become apparent to one of skill in the art. These and other embodiments of the invention are further described by the detailed description that follows.

BRIEF DESCRIPTION OF THE DRAWINGS

[0019] FIG. 1 compares the size distributions of SANDOSTATIN® LAR microspheres (dotted line) with the size distribution of octreotide/PLGA microspheres (dashed line), triamcinolone/PLGA microspheres formulated as described in Table B (see preparation 5), and bupivacaine/PLGA microspheres formulated as described in Table D (see preparation

3)·

[0020] FIG. 2A shows a preparation of triamcinolone/PLGA microspheres imaged using light microscopy.

[0021] FIG. 2B shows a preparation of bupivacaine/PLGA microspheres imaged using light microscopy.

DETAILED DESCRIPTION

[0022] Provided herein are methods of preparing microspheres ( e.g ., single emulsion microspheres) comprising a therapeutic compound or pharmaceutically acceptable salt thereof and a poly(lactic-co-glycolic acid) (PLGA) polymer comprising at least one hydrophobic terminus, as well as microspheres and pharmaceutical compositions related thereto. The present disclosure is based, at least in part, on the finding that making microspheres with PLGA polymer having a hydrophobic or ester-capped terminus allowed for increased loading of compound (e.g., triamcinolone or bupivacaine) in the

microspheres. Moreover, increasing the molecular weight of such PLGA polymers led to further increases in compound loading. These microspheres were also characterized by a more uniform size distribution that other microspheres, e.g. , SANDOSTATIN® LAR depot double emulsion microspheres.

I. Methods of Preparing Microspheres

[0023] Certain aspects of the present disclosure relate to methods of preparing a microsphere of the present disclosure, e.g. , a single emulsion microsphere comprising a therapeutic compound or pharmaceutically acceptable salt thereof and a PLGA polymer comprising at least one hydrophobic terminus. [0024] In some embodiments, the methods of preparing a microsphere of the present disclosure comprise: combining a first solvent and a therapeutic compound or

pharmaceutically acceptable salt thereof to form a first mixture; combining a second solvent and a polymer ( e.g ., PLGA comprising at least one hydrophobic terminus and having a molecular weight of at least 7 kD) to form a second mixture; combining the first and second mixtures to form a feed; dispersing the combined first and second mixtures into an aqueous continuous phase to form a plurality of droplets; and hardening the plurality of droplets to form the plurality of single emulsion microspheres.

[0025] In some embodiments, the second solvent (e.g., mixed with the polymer) is an organic solvent. In some embodiments, the second solvent (e.g, mixed with the polymer) is selected from the group consisting of dichloromethane (DCM), DMA, and DMF.

Suitable polymers (e.g, PLGA polymers) of the present disclosure are discussed in greater detail in section II infra.

[0026] In some embodiments, the dispersed phase comprises the polymer at a concentration of at least 200 mg/mL or about 200 mg/mL. In some embodiments, the dispersed phase comprises an organic acid (e.g, DMA) and an organic solvent (e.g, DCM). In some embodiments, the dispersed phase comprises the organic acid (e.g, DMA) and the organic solvent (e.g, DCM) at a ratio of 1 :9 (organic acid:organic solvent). In some embodiments, the dispersed phase comprises an alcohol (e.g, benzyl alcohol) and an organic solvent (e.g, DCM). In some embodiments, the dispersed phase comprises the alcohol (e.g, benzyl alcohol) and the organic solvent (e.g, DCM) at a ratio of 1 :9

(alcohol: organic solvent).

[0027] In some embodiments, the methods of the present disclosure comprise dispersing the combined first and second mixtures of the present disclosure (e.g, a feed or dispersed phase) into an aqueous continuous phase to form a plurality of droplets.

[0028] In some embodiments, the aqueous continuous phase further comprises an alcohol, e.g, poly(vinyl alcohol) or PVA. In some embodiments, the aqueous continuous phase further comprises a buffer, e.g, an acetate or glycyl -glycine buffer. In some embodiments, the aqueous continuous phase further comprises DCM. In some

embodiments, the aqueous continuous phase is saturated with DCM.

[0029] In some embodiments, the pH of the aqueous continuous phase is greater than 7. In some embodiments, the pH of the aqueous continuous phase is greater than 7.5, or less than 9.0. In some embodiments, the pH of the aqueous continuous phase is about 8.0. In some embodiments, the methods of the present disclosure further include adjusting the pH of the aqueous continuous phase into which the feed is dispersed ( e.g ., so as to achieve a pH of greater than 7.5, less than 9.0, or about 8.0).

[0030] In some embodiments, the methods of the present disclosure further include (e.g., after dispersing the feed and prior to hardening) washing the plurality of microspheres in a second aqueous continuous phase.

[0031] In some embodiments, the methods of the present disclosure comprise hardening a plurality of droplets of the present disclosure to form a plurality of single emulsion microspheres. In some embodiments, hardening the droplet(s) to form single emulsion microsphere(s) of the present disclosure comprises exacervation.

[0032] In some embodiments, the methods of the present disclosure further include (e.g, after initial hardening the droplet to form a microsphere, and/or prior to optional lyophilization or freeze drying) washing the microsphere, e.g, in water.

[0033] In some embodiments, the methods further include (e.g, after hardening the droplet to form a microsphere) lyophilizing or freeze drying the microsphere(s).

[0034] In some embodiments, e.g, when using a glucocorticoid such as triamcinolone as the therapeutic compound, the first solvent comprises an organic acid. In some embodiments, the first solvent (e.g, mixed with the therapeutic compound) comprises dimethylformamide (DMF). In some embodiments, the first solvent (e.g, mixed with the therapeutic compound) comprises DMA.

[0035] In some embodiments, e.g, when using a glucocorticoid such as triamcinolone as the therapeutic compound, the first mixture comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least 400mg/mL. For example, in some embodiments, the first mixture comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least 200mg/mL, at least 250mg/mL, at least 300mg/mL, at least 350mg/mL, or at least 400mg/mL.

[0036] In some embodiments, e.g, when using a glucocorticoid such as triamcinolone as the therapeutic compound, the dispersed phase or feed comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least

20mg/mL or about 20mg/mL.

[0037] In some embodiments, e.g, when using an anesthetic such as bupivacaine as the therapeutic compound, the first solvent comprises an organic acid or alcohol. In some embodiments, the first solvent (e.g, mixed with the therapeutic compound) comprises formic acid, acetic acid, or benzyl alcohol. In some embodiments, the first solvent (e.g, mixed with the therapeutic compound) comprises benzyl alcohol. [0038] In some embodiments, e.g, when using an anesthetic such as bupivacaine as the therapeutic compound, the first mixture comprises the therapeutic compound or

pharmaceutically acceptable salt thereof at a concentration of at least 400mg/mL. For example, in some embodiments, the first mixture comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least 200mg/mL, at least 250mg/mL, at least 300mg/mL, at least 350mg/mL, or at least 400mg/mL.

[0039] In some embodiments, e.g. , when using an anesthetic such as bupivacaine as the therapeutic compound, the dispersed phase or feed comprises the therapeutic compound or pharmaceutically acceptable salt thereof at a concentration of at least 40mg/mL or about 40mg/mL.

II Microspheres

[0040] Certain aspects of the present disclosure relate to microspheres, e.g. , single emulsion microspheres comprising a therapeutic compound or pharmaceutically acceptable salt thereof and a PLGA polymer comprising at least one hydrophobic terminus. In some embodiments, the PLGA polymer has a molecular weight of at least 7 kD. These microspheres can be produced, e.g. , using any of the methods described in section I.

[0041] Further provided herein is a plurality of microspheres produced by any of the methods of the present disclosure, e.g. , as described in section I.

[0042] The term microsphere denotes the encapsulation of the therapeutic

compounds/peptides by the polymer, in some embodiments with the therapeutic

compound/peptide distributed throughout the polymer, which is then a matrix for the therapeutic compound/peptide.

[0043] In some embodiments, a polymer of the present disclosure refers to a PLGA polymer, e.g. , a random or block copolymer made of two types of monomers: glycolyic acid and lactic acid. In some embodiments, the PLGA polymer comprises at least one hydrophobic terminus. In some embodiments, the PLGA polymer comprises more than one hydrophobic terminus. In some embodiments, the PLGA polymer comprises at least one ester-capped terminus. In some embodiments, the PLGA polymer comprises more than one ester-capped terminus. In some embodiments, a PLGA polymer of the present disclosure has a ratio of lactide:glycolide of 50:50. Exemplary PLGA polymers comprising a hydrophobic terminus include, without limitation, RESOMER® RG 502, 503, and 504 (Evonik Industries). Advantageously, the present disclosure demonstrates that using a PLGA polymer comprising a hydrophobic terminus can lead to increased loading of therapeutic compound into the microspheres.

[0044] In some embodiments, a therapeutic compound or salt of the present disclosure comprises a small molecule drug or compound. Exemplary therapeutic compounds are described in greater detail infra.

[0045] In some embodiments, a microsphere of the present disclosure comprises more than one therapeutic compound or pharmaceutically acceptable salt thereof.

Glucocorticoid-containing microspheres

[0046] In some embodiments, the therapeutic compound or salt comprises a

glucocorticoid. In some embodiments, the therapeutic compound or salt comprises triamcinolone.

[0047] In some embodiments, a microsphere of the present disclosure comprises a therapeutic compound or pharmaceutically acceptable salt thereof ( e.g. , a glucocorticoid such as triamcinolone) at a loading level of greater than 6% by total weight of the microsphere. In some embodiments, a microsphere of the present disclosure comprises a therapeutic compound or pharmaceutically acceptable salt thereof (e.g, a glucocorticoid such as triamcinolone) at a loading level of greater than 3%, 4%, 5%, or 6% by total weight of the microsphere. In some embodiments, a microsphere of the present disclosure comprises a therapeutic compound or pharmaceutically acceptable salt thereof (e.g, a glucocorticoid such as triamcinolone) at a loading level of about 6.5% by total weight of the microsphere.

[0048] In some embodiments, a PLGA polymer of the present disclosure has a molecular weight of at least 7 kD. In some embodiments, a molecular weight refers to the average molecular weight of a polymer species. In some embodiments, a molecular weight refers to the minimum or maximum molecular weight of a polymer species. For example, RESOMER® RG 502 (Evonik Industries) has a molecular weight of 7kD-17kD, and RESOMER® RG 503 (Evonik Industries) has a molecular weight of 24kD-38kD. In some embodments, a PLGA polymer of the present disclosure has a molecular weight between 7 kD and 17 kD. In some embodiments, a PLGA polymer of the present disclosure has a molecular weight of at least 7 kD but less than 17 kD (e.g, RESOMER® RG 502; Evonik Industries). In some embodments, a PLGA polymer of the present disclosure has a molecular weight between 38 kD and 54 kD. In some embodiments, a PLGA polymer of the present disclosure has a molecular weight of at least 38 kD but less than 54 kD (e.g, RESOMER® RG 504; Evonik Industries). [0049] Advantageously, the present disclosure provides a plurality of microspheres ( e.g ., as described supra) characterized by a uniform size distribution. For example, in some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 22 and 34pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 90% of the microspheres of the plurality are between 22 and 34pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are within 6pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 90% of the microspheres of the plurality are within 6pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 22 and 34pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and 90%-95% of the microspheres of the plurality are between 22 and 34pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 60% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 60% of the microspheres of the plurality are within 3 pm of a mean diameter.In some embodiments, a plurality of microspheres of the present disclosure is provided in which 60%-70% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and 60%-70% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are between 26 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 20% or at least 30% of the microspheres of the plurality are between 26 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are within 2pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 2pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 20%-30% of the microspheres of the plurality are between 26 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 2pm of a mean diameter. In some

embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 22 and 34pm in diameter, at least 60% of the microspheres of the plurality are between 24 and 30pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 26 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, at least 90% of the microspheres of the plurality are between 22 and 34pm in diameter, at least 60% of the microspheres of the plurality are between 24 and 30pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 26 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 22 and 34pm in diameter, 60%-70% of the microspheres of the plurality are between 24 and 30pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 26 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, 90%-95% of the microspheres of the plurality are between 22 and 34pm in diameter, 60%-70% of the microspheres of the plurality are between 24 and 30pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 26 and 30pm in diameter.

[0050] In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 20 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 90% of the microspheres of the plurality are between 20 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are within 6pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 90% of the

microspheres of the plurality are within 6pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 20 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and 90%- 95% of the microspheres of the plurality are between 20 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are between 22 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 60% of the microspheres of the plurality are between 22 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are within 4pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 60% of the microspheres of the plurality are within 4pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 60%-70% of the microspheres of the plurality are between 22 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and 60%-70% of the microspheres of the plurality are between 22 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 20% or at least 30% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 20%-30% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 20 and 32pm in diameter, at least 60% of the microspheres of the plurality are between 22 and 30pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, at least 90% of the microspheres of the plurality are between 20 and 32pm in diameter, at least 60% of the microspheres of the plurality are between 22 and 30pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 20 and 32pm in diameter, 60%-70% of the microspheres of the plurality are between 22 and 30pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 24 and 30pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, 90%-95% of the microspheres of the plurality are between 20 and 32pm in diameter, 60%-70% of the microspheres of the plurality are between 22 and 30pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 24 and 30pm in diameter.

Anesthetic-containing microspheres

[0051] In some embodiments, the therapeutic compound or salt comprises an anesthetic. In some embodiments, the therapeutic compound or salt comprises bupivacaine.

[0052] In some embodiments, a PLGA polymer of the present disclosure has a molecular weight of at least 7 kD. In some embodiments, a molecular weight refers to the average molecular weight of a polymer species. In some embodiments, a molecular weight refers to the minimum or maximum molecular weight of a polymer species. For example, RESOMER® RG 502 (Evonik Industries) has a molecular weight of 7kD-17kD, and RESOMER® RG 503 (Evonik Industries) has a molecular weight of 24kD-38kD. In some embodments, a PLGA polymer of the present disclosure has a molecular weight between 7 kD and 17 kD. In some embodiments, a PLGA polymer of the present disclosure has a molecular weight of at least 7 kD but less than 17 kD ( e.g. , RESOMER® RG 502; Evonik Industries). In some embodiments, a PLGA polymer of the present disclosure has a molecular weight between 38 kD and 54 kD. In some embodiments, a PLGA polymer of the present disclosure has a molecular weight of at least 38 kD but less than 54 kD (e.g, RESOMER® RG 504; Evonik Industries).

[0053] In some embodiments, a microsphere of the present disclosure comprises a therapeutic compound or pharmaceutically acceptable salt thereof (e.g, an anesthetic such as bupivacaine) at a loading level of greater than 9% by total weight of the microsphere. In some embodiments, a microsphere of the present disclosure comprises a therapeutic compound or pharmaceutically acceptable salt thereof ( e.g ., an anesthetic such as bupivacaine) at a loading level of greater than 5%, 6%, 7%, 8%, or 9% by total weight of the microsphere. In some embodiments, a microsphere of the present disclosure comprises a therapeutic compound or pharmaceutically acceptable salt thereof (e.g., an anesthetic such as bupivacaine) at a loading level of about 9.5% by total weight of the microsphere.

[0054] Advantageously, the present disclosure provides a plurality of microspheres (e.g, as described supra) characterized by a uniform size distribution. For example, in some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 22 and 36pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 90% of the microspheres of the plurality are between 22 and 36pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are within 7pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 90% of the microspheres of the plurality are within 7pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 22 and 36pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and 90%-95% of the microspheres of the plurality are between 22 and 36pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 60% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 60% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 60%-70% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and 60%-70% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are between 28 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 20% or at least 30% of the microspheres of the plurality are between 28 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are within 2pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 2pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 20%-30% of the microspheres of the plurality are between 28 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 2pm of a mean diameter. In some

embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 22 and 36pm in diameter, at least 60% of the microspheres of the plurality are between 26 and 32pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 28 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, at least 90% of the microspheres of the plurality are between 22 and 36pm in diameter, at least 60% of the microspheres of the plurality are between 26 and 32pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 28 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 22 and 36pm in diameter, 60%-70% of the microspheres of the plurality are between 26 and 32pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 28 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 7 kD but less than 17 kD, 90%-95% of the microspheres of the plurality are between 22 and 36pm in diameter, 60%-70% of the microspheres of the plurality are between 26 and 32pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 28 and 32pm in diameter.

[0055] In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 24 and 42pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 90% of the microspheres of the plurality are between 24 and 42pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are within 4pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 90% of the

microspheres of the plurality are within 4pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 24 and 42pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and 90%- 95% of the microspheres of the plurality are between 24 and 42pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are between 26 and 34pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 60% of the microspheres of the plurality are between 26 and 34pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 60% of the microspheres of the plurality are within 4pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 60% of the microspheres of the plurality are within 4pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 60%-70% of the microspheres of the plurality are between 26 and 34pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and 60%-70% of the microspheres of the plurality are between 26 and 34pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 20% or at least 30% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 20% or at least 30% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 3 pm of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 20%-30% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, and at least 20% or at least 30% of the microspheres of the plurality are within 3mih of a mean diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which at least 90% of the microspheres of the plurality are between 24 and 42pm in diameter, at least 60% of the microspheres of the plurality are between 26 and 34pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, at least 90% of the microspheres of the plurality are between 24 and 42pm in diameter, at least 60% of the microspheres of the plurality are between 26 and 34pm in diameter, and/or at least 20% or at least 30% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, a plurality of microspheres of the present disclosure is provided in which 90%-95% of the microspheres of the plurality are between 24 and 42pm in diameter, 60%-70% of the microspheres of the plurality are between 26 and 34pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 26 and 32pm in diameter. In some embodiments, the PGLA polymer has a molecular weight of at least 38 kD but less than 54 kD, 90%-95% of the microspheres of the plurality are between 24 and 42pm in diameter, 60%-70% of the microspheres of the plurality are between 26 and 34pm in diameter, and/or 20%-30% of the microspheres of the plurality are between 26 and 32pm in diameter.

III. Methods of Using Microspheres

[0056] Certain aspects of the present disclosure relate to methods of treating a condition by administering to an individual ( e.g ., in need thereof) a therapeutically effective amount of a microsphere (see sections I and II) or pharmaceutical composition (see section IV) of the present disclosure.

[0057] In some embodiments, e.g., when using a glucocorticoid such as triamcinolone as the therapeutic compound, the condition is an inflammatory condition, including but not limited to gout, arthritis, allergy/allergic reaction, and skin conditions.

[0058] In some embodiments, e.g, when using an anesthetic such as bupivacaine as the therapeutic compound, the condition is pain. In some embodiments, the microsphere or composition is used when analgesia/anesthetic is required, such as for a medical procedure like surgery. In some embodiments, the microsphere or composition is used to effect local anesthesia, caudal nerve block, epidural nerve block, peripheral nerve block, retrotubular anesthesia, or spinal anesthesia. [0059] In some embodiments, the microsphere or composition is administered by injection, e.g ., subcutaneous or intramuscular injection. In some embodiments, the microsphere or composition allows for use of a smaller needle size and/or less frequent administration, e.g. , as compared to a reference microsphere (e.g, a double emulsion microsphere) comprising the same active pharmaceutical ingredient.

[0060] In some embodiments, an“individual” is a mammal. Mammals include, but are not limited to, domesticated animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain embodiments, the individual is a human.

IV. Pharmaceutical Compositions, Kits, and Articles of Manufacture

[0061] Certain aspects of the present disclosure relate to pharmaceutical compositions comprising a microsphere of the present disclosure (e.g, as described in section II above, or prepared by a method described in section I above). The pharmaceutical compositions may find use, e.g, in any of the methods described in section III above.

[0062] The term“pharmaceutical composition” or "pharmaceutical formulation" refers to a preparation which is in such form as to permit the biological activity of an active ingredient contained therein to be effective, and which contains no additional components which are unacceptably toxic to a subject to which the formulation would be administered.

[0063] In some embodiments, a pharmaceutical composition of the present disclosure comprises a microsphere of the present disclosure and a pharmaceutically acceptable carrier. A“pharmaceutically acceptable carrier” refers to an ingredient in a pharmaceutical formulation, other than an active ingredient, which is nontoxic to a subject. A

pharmaceutically acceptable carrier includes, but is not limited to, a buffer, excipient, stabilizer, diluent, or preservative.

[0064] As is well known in the art, pharmaceutically acceptable excipients are relatively inert substances that facilitate administration of a pharmacologically effective substance and can be supplied as liquid solutions or suspensions, as emulsions, or as solid forms suitable for dissolution or suspension in liquid prior to use. In some embodiments, the excipient is external to the microsphere. For example, an excipient can give form or consistency, or act as a diluent. Suitable excipients include but are not limited to stabilizing agents, wetting and emulsifying agents, salts for varying osmolarity, encapsulating agents, pH buffering substances, and buffers. Such excipients include any pharmaceutical agent suitable for injection which may be administered without undue toxicity. Pharmaceutically acceptable excipients include, but are not limited to, sorbitol, any of the various TWEEN compounds, and liquids such as water, saline, glycerol and ethanol. Pharmaceutically acceptable salts can be included therein, for example, mineral acid salts such as

hydrochlorides, hydrobromides, phosphates, sulfates, and the like; and the salts of organic acids such as acetates, propionates, malonates, benzoates, and the like. A thorough discussion of pharmaceutically acceptable excipients is available in REMINGTON’S PHARMACEUTICAL SCIENCES (Mack Pub. Co., N.J. 1991). In certain embodiments, the excipient is selected from acetate, citrate, lactate, polyols ( e.g ., mannitol or glycerol), carboxy-methyl cellulose and hydroxy-prolyl cellulose, and glycine.

[0065] Formulations described herein may be utilized in depot form, e.g. injectable microspheres or implants.

[0066] In some embodiments, pharmaceutically acceptable excipients may include pharmaceutically acceptable carriers. Such pharmaceutically acceptable carriers can be sterile liquids, such as water and oil, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, and the like. Saline solutions and aqueous dextrose, polyethylene glycol (PEG) and glycerol solutions can also be employed as liquid carriers, particularly for injectable solutions. Additional ingredients may also be used, for example preservatives, buffers, tonicity agents, antioxidants and stabilizers, nonionic wetting or clarifying agents, viscosity-increasing agents (e.g., carboxymethylcellulose or a poloxamer), and the like. The kits described herein can be packaged in single unit dosages or in multidosage forms.

[0067] In some embodiments, the kits or articles further comprise a package insert with instructions for using the microspheres or pharmaceutical compositions related thereto, e.g, in any of the methods described in section III above.

[0068] The use of the terms“a” and“an” and“the” and similar referents (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. The terms “comprising,”“having,”“including,” and“containing” are to be construed as open-ended terms (i.e., meaning“including, but not limited to,”) unless otherwise noted. Wherever an open-ended term is used to describe a feature or element, it is specifically contemplated that a closed-ended term can be used in place of the open-ended term without departing from the spirit and scope of the disclosure. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g.,“such as”) provided herein, is intended merely to better illuminate the description and does not pose a limitation on the scope of the description unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the compositions, methods, and kits disclosed herein.

[0069] Preferred embodiments are described herein. Variations of those preferred embodiments may become apparent to those working in the art upon reading the foregoing description. It is expected that skilled artisans will be able to employ such variations as appropriate, and the practice of the compositions, methods, and kits described herein otherwise than as specifically described herein. Accordingly, the compositions, methods, and kits described herein include all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the description unless otherwise indicated herein or otherwise clearly contradicted by context.

EXAMPLES

[0070] The present disclosure will be more fully understood by reference to the following examples. The examples should not, however, be construed as limiting the scope of the present disclosure. It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims.

Example 1: Preparation of single emulsion microspheres using end-capped polymers

Methods

Formulations

[0071] Triamcinolone:PLGA and bupivacaine:PLGA microspheres were generated according to the formulations shown in Tables B and D, respectively. Concentrations refer to the mixtures prior to extrusion.

737732000640

Table A. Solvent study on triamcinolone

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737732000640

Table B. Formulations for triamcinolone:PLGA microspheres

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737732000640

Table C. Solvent study on bupivacaine

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Table D. Formulations for bupivacaine:PLGA microspheres

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Microsphere production

[0072] Briefly, microspheres were generated as follows. Triamcinolone acetonide was dissolved in DMA, and PLGA (502, 502H, or 504) was dissolved in DCM, as indicated in Table B. Bupivacaine hydrochloride was dissolved in benzyl alcohol, and PLGA (502, 502H, or 504) was dissolved in DCM, as indicated in Table D. For octreotide

microspheres, octreotide was dissolved in ethanol, and PLGA was dissolved in methylene chloride.

[0073] All mixtures were then combined and extruded through calibrated pore-size membranes into a flowing continuous phase (components as listed in Table B for triamcinolone and as listed in Table D for bupivacaine) that strips the droplets from the surface of the membrane. The droplets were then hardened into microspheres by solvent exchange with the continuous phase without methylene chloride, washed with water, and dried by lyophilization.

Scanning electron microscopy (SEM)

[0074] For surface images, samples were mounted to an aluminum stub using a carbon tab followed by gold sputter coating and SEM imaging.

[0075] For cross-sections, samples were mixed with Loctite epoxy and allowed to cure overnight. The samples were then frozen using liquid nitrogen and cracked using a mortar and pestle. Cracked portions were then mounted to an aluminum stub using carbon tape and colloidal graphite and sputter coated with gold.

Size distribution

[0076] Size distribution of microsphere populations was quantitated using automated image analysis. Cell Profiler (cellprofiler.org) software was used to find and measure microsphere diameter based on light microscopy images. The identifyPrimaryObjects module was used and calibrated with a micrometric scale. Starting with light microscopic images of microspheres, images were analyzed to automatically detect microsphere shape, fill in microsphere shapes, and measure microsphere dimensions.

Results

[0077] To test microsphere formulations, the following solvents were tested for ability to solubilize triamcinolone acetonide: ethanol, methanol, acetone, dichloromethane (DCM), formic acid, ethyl acetate, dimethylsulfoxide (DMSO), dimethylacetamide (DMA), dimethylformamide (DMF), water, benzyl alcohol, and ethyl formate. The results are shown in Table A. On the basis of these studies, microspheres were generated using the dispersed and continuous phases described in Table B.

[0078] The following solvents were tested for ability to solubilize bupivacaine hydrochloride: ethanol, methanol, acetone, dichloromethane (DCM), formic acid, ethyl acetate, dimethylsulfoxide (DMSO), dimethylacetamide (DMA), dimethylformamide (DMF), water, benzyl alcohol, acetic acid, acetonitrile, trimethylamine, and ethyl formate. The results are shown in Table C. On the basis of these studies, microspheres were generated using the dispersed and continuous phases described in Table D.

[0079] Microspheres with up to 6.45% loading of triamcinolone and 9.41% loading of bupivacaine were generated. Table E summarizes the conditions that led to maximum loading levels observed in microspheres with triamcinolone, bupivacaine, or octreotide.

Table E.

[0080] As shown in Table E, optimal conditions for microsphere formation and loading differed among octreotide, triamcinolone, and bupivacaine. In particular, while octreotide loading was increased using smaller PLGA polymers ( e.g ., 502H or 503H), larger, end- capped PLGA polymers (e.g., 504H) led to greater loading of triamcinolone or bupivacaine.

[0081] The effects of different PLGA polymers on microsphere loading were tested.

As shown in Table F, for both triamcinolone and bupivacaine, increasing the molecular weight of the end-capped PLGA species led to increased loading of API, with highest loading observed when using a 504 PLGA species (MW: 38-54kD).

Table F.

[0082] Other microsphere properties were investigated. Compared to Sandostatin LAR microspheres, the triamcinolone- or bupivacaine-containing microspheres produced as described above were characterized by a more uniform size distribution (FIG. 1). 90-95% of triamcinolone or bupivacaine microspheres had a diameter from 16-32 pm, with 60-70% of the microspheres having a diameter from 22-26 pm, and 20-30% of the microspheres having a diameter from 22-24 pm.

[0083] Triamcinolone- and bupivacaine-containing microspheres were also examined by light microscopy. A relatively homogeneous distribution of spherical microspheres was observed for microspheres containing triamcinolone (FIG. 2A) or bupivacaine (FIG. 2B).

[0084] Although the foregoing disclosure has been described in some detail by way of illustration and example for purposes of clarity of understanding, the descriptions and examples should not be construed as limiting the scope of the present disclosure. The disclosures of all patent and scientific literature cited herein are expressly incorporated in their entirety by reference.