EA046729B1 - PHARMACEUTICAL COMPOSITIONS CONTAINING (S)-4-(4-(4-(((2-(2,6-DIOXOPIPERIDIN-3-YL)-1-OXOISOINDOLIN-4-YL)OXY)METHYL)BENZYL)PIPERAZINE-1 -IL)-3-FLUOROBENZONITRILE, AND METHODS OF THEIR APPLICATION - Google Patents

Description

This application claims benefit from U.S. Provisional Application No. 62/790229, filed January 9, 2019, which is incorporated herein by reference in its entirety.

1. Technical field

Presented herein are pharmaceutical compositions containing (S)-4-(4-(4-(((2(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-4-yl)oxy)methyl)benzyl) piperazin-1-yl)-3-fluorobenzonitrile or an enantiomer, mixture of enantiomers, tautomer, isotopologue or pharmaceutically acceptable salt thereof, as well as a carrier or diluent. Methods of using such pharmaceutical compositions for the treatment, prevention and management of various disorders are also provided herein.

2. State of the art

Multiple myeloma (MM) is a cancer of the plasma cells in the bone marrow. Plasma cells typically produce antibodies and play a key role in immune function. However, uncontrolled growth of these cells leads to bone pain and fractures, anemia, infections and other complications. Multiple myeloma is the second most common hematologic malignancy, although the exact causes of multiple myeloma remain unknown. Multiple myeloma causes high levels of proteins in the blood, urine, and organs, including but not limited to M protein and other immunoglobulins (antibodies), albumin, and beta-2 microglobulin, except in some patients (estimated at 1% to 5%), myeloma cells that do not secrete these proteins (so-called nonsecretory myeloma). M protein, short for monoclonal protein, also known as paraprotein, is a particularly abnormal protein produced by myeloma plasma cells and can be found in the blood or urine of almost all patients with multiple myeloma, except patients with nonsecretory myeloma or whose myeloma cells produce immunoglobulin light chains with a heavy chain.

Skeletal symptoms, including bone pain, are among the most clinically significant symptoms of multiple myeloma. Malignant plasma cells secrete osteoclast-stimulating factors (including IL-1, IL-6, and TNF), which cause calcium to be leached from bone, causing lytic lesions; hypercalcemia is another symptom. Osteoclast stimulating factors, also called cytokines, can prevent apoptosis or death of myeloma cells. Fifty percent of patients have radiologically detectable skeletal lesions associated with myeloma at the time of diagnosis. Other common clinical symptoms of multiple myeloma include polyneuropathy, anemia, hyperviscosity, infection, and kidney failure.

Current therapy for multiple myeloma may involve one or more of surgery, stem cell transplant, chemotherapy, immunotherapy, and/or radiation therapy to destroy a patient's multiple myeloma cells. All current therapeutic approaches present significant disadvantages for the patient.

In the last decade, new therapeutic agents, particularly immunomodulatory drugs such as lenalidomide and pomalidomide, have significantly increased response rates and prolonged progression-free survival (PFS) and overall survival (OS) in patients with multiple myeloma. However, persistent levels of residual disease that are below the sensitivity of bone marrow (BM) morphology, immunofixation protein electrophoresis, and light chain quantitation exist in many patients with multiple myeloma even after these patients achieve a complete response (CR), and over time will cause a relapse of the disease. Minimal residual disease (MRD) in myeloma is an independent predictor of progression-free survival (PFS) and is being considered as a surrogate trial endpoint to improve identification of effective treatments, especially for exploratory trials, which currently require 5 to 10 years of follow-up for identifying differences in survival. Thus, minimal residual disease (MRD) monitoring in patients with multiple myeloma provides prognostic value for predicting PFS and OS and making treatment decisions. To detect minimal residual disease (MRD) in myeloma, a threshold of 0.01% (10 ^-4 ) after treatment can be used, i.e. the presence of 10 ^-4 cells or fewer multiple myeloma cells as a percentage of total bone marrow mononuclear cells is considered MRD - negative and having 10 ^-4 cells or higher MRD-positive. The 10 ⁴ MRD threshold was originally based on technical capabilities, but it is now possible to quantify MRD at 10 ^-5 using flow cytometry and 10 ^-6 using high-throughput sequencing. (Rawstron et al., Blood 2015;125(12):1932-1935). Methods for measuring MRD include VDJ DNA sequencing, polymerase chain reaction (PCR) (including allele-specific PCR, ASO-PCR), and multiparameter flow cytometry (MPF). MRD assays, for example based on clonotype profile measurements, are also described in US Pat. No. 8,628,927 to Faham et al., which is incorporated herein by reference.

There is a significant need for safe and effective compounds and methods for the treatment, prevention and management of multiple myeloma, including for patients who have

- 1 046729 multiple myeloma newly diagnosed or that are refractory to standard treatment, while reducing or preventing toxicity and/or side effects associated with conventional treatments.

The variety of possible pharmaceutical compositions (eg, oral dosage forms containing various excipients) creates a potential variety of physical and chemical properties for a given pharmaceutical compound. The discovery and selection of pharmaceutical compositions is of great importance for the development of an effective, stable and marketable pharmaceutical product.

3. Essence of the invention

Provided herein are pharmaceutical compositions (e.g., oral dosage forms) containing Compound 1:

ABOUT

1, or an enantiomer, mixture of enantiomers, tautomer, isotopologue or pharmaceutically acceptable salt and carrier or diluent thereof. Compound 1 has the chemical name (S)-4-(4-(4-(((2-(2,6dioxopiperidin-3-yl)-1-oxoisoindolin-4-yl)oxy)methyl)benzyl)piperazine-1 - yl)-3-fluorobenzonitrile. This document also provides methods for preparing pharmaceutical compositions.

In one embodiment, the carrier or diluent is mannitol. In one embodiment, the carrier or diluent is a mixture of starch and lactose.

The pharmaceutical compositions provided herein are useful compositions for use in animals or humans. Thus, embodiments herein cover the use of these pharmaceutical compositions as a final drug product. Some embodiments provide pharmaceutical compositions useful for preparing final dosage forms with improved properties, e.g., powder flow properties, compaction properties, tabletting properties, stability properties, and excipient compatibility properties, among others, that are needed for manufacturing, processing, formulation, and/or or storage of finished medicinal products.

Also provided are pharmaceutical compositions formulated for administration by suitable route and means containing effective concentrations of Compound 1 provided herein. In one embodiment, the pharmaceutical compositions are oral dosage forms. In one embodiment, the pharmaceutical compositions are immediate release (IR) oral dosage forms.

In one embodiment, the pharmaceutical compositions deliver amounts effective to treat multiple myeloma. In one embodiment, the pharmaceutical compositions deliver amounts effective to prevent multiple myeloma. In one embodiment, the pharmaceutical compositions deliver amounts effective to eliminate multiple myeloma.

In one embodiment, provided herein are methods of treating multiple myeloma, including administering the pharmaceutical compositions provided herein. Also provided herein are combination therapies using the pharmaceutical compositions provided herein in combination with therapy, for example, with another pharmaceutical agent having activity against multiple myeloma or its symptoms. Examples of therapies within the methods include, but are not limited to, surgery, chemotherapy, radiation therapy, biological therapy, stem cell transplantation, cell therapy, and combinations thereof.

Also provided is a pharmaceutical package or kit including one or more containers filled with one or more ingredients of the pharmaceutical compositions. Such container(s) may not necessarily be accompanied by a notice in a form prescribed by a government agency regulating the manufacture, use, or sale of pharmaceutical or biological products, the notice reflecting the agency's approval of the manufacture, use, and sale for administration to humans. The packaging or kit may contain information about the method of administration, the sequence of administration of the drug

- 2 046729 means (for example, separately, sequentially or simultaneously) or the like.

Methods for preparing the pharmaceutical compositions presented herein are also provided. In one embodiment, the method includes coating the active ingredient with hydrophobic silica.

These and other aspects of the subject matter described herein will become apparent upon reference to the following detailed description.

4. Brief description of graphic materials

In fig. Figure 1 shows a typical powder X-ray diffraction (pXRD) pattern of Form K of the free base of compound 1.

In fig. Figure 2 shows a typical pXRD pattern of the K' form of the free base of compound 1.

In fig. Figure 3 shows data on hydrolysis and oxidation impurities in the binary compatibility study.

In fig. Figure 4 shows data on chiral impurities in a binary compatibility study.

In fig. 5A, FIG. 5B, FIG. 5C and FIG. 5D shows the stability profiles of various capsules for degradation products at relative retention times (RRT) of 4.2 min, 7.1 min, 7.6 min, and 11.3 min, respectively.

In fig. Figure 6 shows the chiral impurity profiles of various capsules.

In fig. Figure 7 shows two-stage dissolution of 2 mg formulations at pH 2 (0-90 min) and 6.8 (90-150 min).

In fig. 8 shows comparative dissolution release profiles of Cap-25, Cap-29, Cap33 and Cap-34.

In fig. Figure 9 shows an accelerated evaluation of the stability of a mannitol-starch-lactose capsule formulation (BIC).

In fig. 10 shows a comparative accelerated evaluation of the stability of BIC formulations based on mannitol with fumaric acid and maleic acid.

In fig. 11 shows comparative chemical degradation profiles of BIC formulations based on mannitol with fumaric acid and maleic acid.

In fig. 12 shows comparative oxidative degradation profiles (oxidation-1) of starch-lactose BIC formulations with disintegrants compared to non-disintegrant formulations and mannitol-based BIC formulations.

In fig. 13 shows comparative oxidative degradation (oxidation-2) profiles of starch-lactose BIC formulations with disintegrants compared to non-disintegrant formulations and mannitol-based BIC formulations.

In fig. 14 shows comparative hydrolytic degradation profiles (hydrolytic-1) of starch-lactose BIC formulations with disintegrants compared to non-disintegrant formulations and mannitol-based BIC formulations.

In fig. 15 shows comparative hydrolytic degradation profiles (hydrolytic-2) of starch-lactose BIC formulations with disintegrants compared to non-disintegrant formulations and mannitol-based BIC formulations.

In fig. 16 shows comparative profiles of chiral impurities in starch-lactose BIC formulations with disintegrants compared to non-disintegrant formulations and mannitol-based BIC formulations.

In fig. 17A and FIG. 17B shows SEM images of a crystal of Compound 1 uncoated and coated with 2% Aerosil R-972, respectively.

In fig. 18A, FIG. 18B, FIG. 18C and FIG. 18D shows SEM images of a crystal of Compound 1 coated with 5% Aerosil R-972, coated with 5% Aerosil 300, coated with 5% Aerosil 200, and uncoated with silica, respectively.

In fig. Figure 19 shows the comparative compatibility of the drug and the excipient of various brands of silica.

In fig. 20 shows the effect of different levels of Aerosil 200 Pharma on the release profile when dissolving 2 mg of a starch-lactose BIC formulation.

In fig. 21 shows the powder flow behavior of 0.1 mg, 0.5 mg and 2 mg BIC formulations of mannitol and starch-lactose.

In fig. 22 shows a comparative content uniformity (CU) assessment for 2 mg starch-lactose BIC formulations using 2% Aerosil 200 and 1% Aerosil 200.

In fig. 23 shows comparative dissolution profiles of BIC formulations with 0.1 mg mannitol and starch-lactose BIC formulations.

In fig. 24 shows comparative dissolution profiles of BIC formulations with 0.5 mg mannitol and starch-lactose BIC formulations.

In fig. 25 shows comparative dissolution profiles of BIC formulations with 2 mg mannitol and starch-lactose BIC formulations.

In fig. Figure 26 shows stability data (degradation chemicals) for BIC formulations with 0.1 mg, 0.5 mg, and 2 mg mannitol in an HDPE bottle without desiccant.

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In fig. 27 shows the chiral impurity profiles of BIC formulations with 0.1 mg, 0.5 mg and 2 mg mannitol.

In fig. 28 shows comparative dissolution stability data for BIC formulations with 0.1 mg mannitol and starch-lactose BIC formulations.

In fig. 29 shows comparative dissolution stability data for BIC formulations with 0.5 mg mannitol and starch-lactose BIC formulations.

In fig. 30 shows comparative dissolution stability data for BIC formulations with 2 mg mannitol and starch-lactose BIC formulations.

5. Detailed description

5.1 Definitions

As used herein and in the specification and appended claims, the singular forms include the plural as well as single references unless the context clearly indicates otherwise.

As used herein, the terms containing and including may be used interchangeably. The terms containing and including should be interpreted as indicating the presence of specified features or components, but do not exclude the presence or addition of one or more features or components, or groups thereof. Additionally, the terms containing and including are intended to include examples covered by the term consisting of. Therefore, the term consisting of may be used in place of the terms containing and including to provide more specific embodiments of the invention.

The term consisting of means that an object contains at least 90%, 95%, 97%, 98%, or 99% of the stated functions or components of which it is composed. In another embodiment, the term consisting of excludes from the scope of any subsequent listing any other features or components other than those that are not essential to the technical effect to be achieved.

As used herein, the term or should be interpreted to include or to mean any one or any combination. Therefore, A, B or C means any of the following: A; B; C; A and B; A and C; B and C; A, B and C. An exception to this definition will occur only when the combination of elements, functions, stages or activities are in some way inherently exclusive of each other.

As used herein and unless otherwise indicated, the terms about and approximately, when used in connection with doses, amounts, or weight percentages of the ingredients of a composition or dosage form, mean the dose, amount, or weight percent as recognized by one of ordinary skill in the art for providing a pharmacological effect equivalent to the effect obtained from the specified dose, amount or mass percentage. In some embodiments, the terms about and about when used in this context mean a dose, amount, or percentage by weight of within 30%, within 20%, within 15%, within 10%, or within 5% of said dose, amount, or mass percent.

As used herein and unless otherwise specified, the terms about and approximately when used in connection with a numerical value or range of values that is provided to characterize a particular solid form, such as a specific temperature or range of temperatures, such as, for example, describing temperature melting, dehydration, desolvation or vitrification; a change in mass, such as, for example, a change in mass depending on temperature or humidity; solvent or water content, for example, by weight or percentage; or peak position, such as, for example, in analysis by IR or Raman spectroscopy or pXRD; indicate that the value or range of values may deviate to an extent considered reasonable by one skilled in the art while still describing a particular solid form. For example, in specific embodiments, the terms about and about when used in this context indicate that the numerical value or range of values may vary within the range of 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6% , 5%, 4%, 3%, 2%, 1.5%, 1%, 0.5%, or 0.25% of the specified value or range of values. For example, in some embodiments, the pSAR peak position value may vary by up to ±0.2 degrees 2θ while still describing a particular pSAR peak. In this context, a tilde (i.e. ~) preceding a numeric value or range of values indicates about or approximately.

Unless otherwise noted, the terms x-ray powder diffraction, powder x-ray diffraction, PXRD, and pXRD are used interchangeably in this application.

As used herein and unless otherwise noted, the terms solid form and related terms refer to a physical form that is not primarily in a liquid or gaseous state. As used herein, the terms solid form and solid forms include semi-solids. Solid forms may be crystalline, amorphous, partially crystalline, partially amorphous, or mixtures of forms.

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As used herein and unless otherwise indicated, the term crystalline and related terms used herein, when used to describe a substance, component, product or form, mean that the substance, component, product or form is substantially crystalline, for example, as determined by x-ray diffraction. See, for example, Remington: The Science and Practice of Pharmacy, ^21st edition, Lippincott, Williams and Wilkins, Baltimore, MD (2005); The United States Pharmacopeia, ^23rd edition, 1843-1844 (1995).

As used herein and unless otherwise indicated, the term amorphous, amorphous form and related terms as used herein mean that the substance, component or product in question is not substantially crystalline as determined by x-ray diffraction. In particular, the term amorphous form describes a disordered solid form, rn.e. solid form, devoid of long-range crystalline order. In some embodiments, the amorphous form of the substance may be substantially free of other amorphous forms and/or crystalline forms. In other embodiments, the amorphous form of the substance may contain less than about 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%. 45 or 50% of one or more other amorphous forms and/or crystalline forms, based on weight. In some embodiments, the amorphous form of the substance may be physically and/or chemically pure. In some embodiments, the amorphous form of the substance may be physically and/or chemically pure about 99%, 98%, 97%, 96%, 95%, 94%, 93%, 92%, 91%, or 90%. In some embodiments, the amorphous form of the substance may contain additional components or ingredients (eg, an additive, polymer, or filler that may serve to further stabilize the amorphous form). In some embodiments, the amorphous form may be a solid solution.

As used herein, unless otherwise indicated, the term pharmaceutically acceptable salts refers to salts derived from pharmaceutically acceptable, relatively non-toxic acids, including inorganic acids and organic acids. In some embodiments, suitable acids include, but are not limited to, acetic, benzenesulfonic, benzoic, camphorsulfonic, carbonic, citric, dihydrophosphoric, ethenesulfonic, fumaric, galacturonic, gluconic, glucuronic, glutamic, hydrobromic, hydrochloric, hydroiodic, isobutyric, iseic, lactic, maleic, malic, malonic, mandelic, methanesulfonic, monohydric carbonic, monohydrophosphoric, monohydrosulfuric, mucus, nitric, pamoic, pantothenic, phosphoric, phthalic, propionic, suberic, succinic, sulfuric, tartaric, toluenesulfonic acid, etc. (See, e.g., S. M. Berge et al., J. Pharm. Sci., 66:1-19 (1977); and Handbook of Pharmaceutical Salts: Properties, Selection and Use, P. H. Stahl and C. G. Wermuth, Eds., ( 2002), Wiley, Weinheim). In some embodiments, suitable acids are strong acids (e.g., with a pKa less than about 1), including, but not limited to, hydrochloric, hydrobromic, sulfuric, nitric, methanesulfonic, benzenesulfonic, toluenesulfonic, naphthalene sulfonic, naphthalene disulfonic, pyridine sulfonic, or other substituted sulfonic acids. acids. Also included are salts of other relatively non-toxic compounds that have acidic properties, including amino acids such as aspartic acid and the like, and other compounds such as aspirin, ibuprofen, saccharin and the like. Acid addition salts can be prepared by contacting the neutral form of the compound with a sufficient amount of the desired acid, either pure or in a suitable solvent. As solids, salts can exist in crystalline or amorphous forms or mixtures thereof. Salts can also exist in polymorphic forms.

As used herein, the term multiple myeloma refers to hematological conditions characterized by malignant plasma cells and includes the following disorders: monoclonal gammopathy of undetermined significance (MGUS); low, intermediate and high risk multiple myeloma; newly diagnosed multiple myeloma (including newly diagnosed low-, intermediate-, and high-risk multiple myeloma); transplant eligible and transplant ineligible multiple myeloma; smoldering (sluggish) multiple myeloma (including smoldering multiple myeloma of low, intermediate and high risk); active multiple myeloma; solitary plasmacytoma; extramedullary plasmacytoma; plasma cell leukemia; multiple myeloma of the central nervous system; light chain myeloma; nonsecretory myeloma; Immunoglobulin D myeloma; and immunoglobulin E myeloma; and multiple myeloma, characterized by genetic abnormalities such as cyclin D translocations (eg, t(11;14)(q13;q32); t(6;14)(p21;32); t(12;14)(p13;q32) ); or t (6; 20);); MMSET translocations (eg t(4;14)(p16;q32)); MAF translocations (e.g. t(14;16)(q32;q32); t(20;22); t(16;22)(q11;q13); or t(14;20)(q32;q11)); or other chromosomal factors (eg, deletion of 17p13 or chromosome 13; del(17/17p), non-hyperdiploidy, and amplification (1q)).

As used herein and unless otherwise noted, the terms treat, therapy and treatment refer to the alleviation or reduction in severity of a symptom associated with the disease or condition being treated, such as multiple myeloma.

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The term prevention includes suppression of a symptom of a specific disease or disorder, such as multiple myeloma. In some embodiments, patients with a family history of multiple myeloma are candidates for prophylactic regimens. Typically, the term prevention refers to administering a drug before symptoms appear, especially in patients at risk for multiple myeloma.

As used herein, unless otherwise specified, the term management includes preventing the recurrence of a particular disease or disorder, such as multiple myeloma, in a patient who has suffered from it, prolonging the time a patient has suffered from that disease or disorder to remain in remission, reducing the level of patient mortality and/or maintaining reduction in severity or avoidance of a symptom associated with the disease or condition being treated.

As used herein, a subject or patient means an animal, typically a mammal, including a human, such as a human patient.

The term relapse refers to a situation where patients who have had multiple myeloma in remission after therapy experience a return of myeloma cells and/or a decrease in normal cells in the bone marrow.

The term refractory or resistant refers to circumstances where patients, even after intensive treatment, have residual myeloma cells and/or reduced normal cells in the bone marrow.

As used herein, the term induction therapy refers to the first treatment of a disease or the first treatment to induce complete remission of a disease, such as cancer. Induction therapy itself is considered the best treatment available. If residual cancer is found, patients are treated with another therapy called re-induction. If the patient is in complete remission after induction therapy, additional consolidation and/or maintenance therapy is given to prolong remission or potentially cure the patient.

As used herein, the term consolidation therapy refers to the treatment of a disease after remission has first been achieved. For example, consolidation therapy for cancer is treatment given after the cancer has disappeared after initial therapy. Consolidation therapy may include radiation therapy, a stem cell transplant, or treatment with cancer drugs. Consolidation therapy is also called intensification therapy and post-remission therapy.

As used herein, the term maintenance therapy refers to the treatment of a disease after remission or best response has been achieved to prevent or delay relapse. Maintenance therapy may include chemotherapy, hormonal therapy, or targeted therapy.

Remission as used herein means a reduction or disappearance of the signs and symptoms of a cancer, such as multiple myeloma. In partial remission, some, but not all, signs and symptoms of cancer disappear. In complete remission, all signs and symptoms of cancer have disappeared, although cancer may still be present in the body.

As used herein, the term transplant refers to high-dose therapy for the purpose of stem cell salvage. Hematopoietic (blood) or bone marrow stem cells are used not for treatment, but to save the patient after high-dose therapy, such as chemotherapy and/or high-dose radiation. The transplant includes an autologous stem cell transplant (ASCT), which refers to the use of the patient's own stem cells, which are collected and used as replacement cells. In some embodiments, the graft also includes a tandem graft or multiple grafts.

As used herein, unless otherwise indicated, the terms a therapeutically effective amount and an effective amount of a compound refer to an amount sufficient to provide a therapeutic effect in the treatment, prevention and/or management of a disease, such as multiple myeloma, or to delay or minimize one or more symptoms. associated with a disease or disorder that needs to be treated. The terms therapeutically effective amount and effective amount can encompass an amount that improves overall therapy, reduces or eliminates symptoms or causes of a disease or disorder, or increases the therapeutic effectiveness of another therapeutic agent.

The terms co-administration and in combination with include the administration of one or more therapeutic agents (for example, a compound provided herein and another anti-multiple myeloma agent, anti-cancer agent or supportive care agent) either simultaneously, concurrently or sequentially without any specific deadlines. In one embodiment, the agents are present in the cell or body of the patient at the same time or exert their biological or therapeutic effects at the same time. In one embodiment, the therapeutic agents are in the same composition or unit dosage form. In another embodiment, the therapeutic agents are in separate compositions or

- 6 046729 standard dosage forms.

The term maintenance therapy refers to any substance that treats, prevents, or controls an adverse effect of treatment with Compound 1, or an enantiomer thereof, or a mixture of enantiomers, a tautomer, an isotopologue, or a pharmaceutically acceptable salt.

The term biological therapy refers to the administration of biological therapeutics such as umbilical cord blood, stem cells, growth factors and the like.

In the context of a cancer such as multiple myeloma, inhibition can be assessed by inhibition of disease progression, inhibition of tumor growth, shrinkage of the primary tumor, relief of tumor-associated symptoms, inhibition of factors secreted by the tumor, delay in the appearance of primary or secondary tumors, delayed development of primary or secondary tumors. tumors, reducing the incidence of primary or secondary tumors, slowing or reducing the severity of secondary disease effects, delaying tumor growth and regression, increasing time to progression (TTP), increasing progression-free survival (PFS), increasing overall survival (OS), among others. As used herein, OS refers to the time from the start of treatment to death from any cause. TTP, as used herein, refers to the time from initiation of treatment to tumor progression; The TTP does not include deaths. In one embodiment, PFS means the time from the start of treatment to tumor progression or death. In one embodiment, PFS means the time from the first dose of a compound to the first occurrence of disease progression or death from any cause. In one implementation of PFS, scores will be calculated using Kaplan-Meier estimates. Event-free survival (EFS) refers to the time from the start of treatment to any treatment failure, including disease progression, discontinuation of treatment for any reason, or death. In one embodiment, the overall response rate (ORR) means the percentage of patients who achieve a response. In one embodiment, ORR means the sum of the percentage of patients who achieve a complete response and a partial response. In one embodiment, ORR means the percentage of patients who have best response > partial response (PR), according to the IMWG Unified Response criteria. In one embodiment, duration of response (DoR) is the time from achieving response to disease relapse or progression. In one embodiment, DoR is the time from achieving response > partial response (PR) to disease relapse or progression. In one embodiment, the DoR is the time from the first documentation of a response to the first documentation of progressive disease or death. In one embodiment, the DoR is the time from first documentation of response > partial response (PR) to first documentation of progressive disease or death. In one embodiment, time to response (TTR) means the time from the first dose of a compound to the first confirmation of a response. In one embodiment, TTR means the time from the first dose of a compound to the first confirmation of response > partial response (PR). In extreme cases, complete inhibition is referred to herein as prophylaxis or chemoprevention. As used herein, the term prevention includes either completely preventing the occurrence of clinically obvious cancer or preventing the occurrence of preclinically obvious cancer. This definition is also intended to include preventing transformation into malignant cells or stopping or reversing the development of precancerous cells into malignant cells. This includes preventive treatment for individuals at risk of developing cancer.

In some embodiments, treatment for multiple myeloma may be assessed using the International Uniform Response Criteria for Multiple Myeloma (IURC) (see Durie BGM, Harousseau J-L, Miguel JS, et al. International uniform response criteria for multiple myeloma. Leukemia, 2006; (10 ) 10: 1-7), using the response and endpoint definitions shown below:_______________________

Response subcategory Answer criteria ^a sCR CR as defined below plus Normal FLC and Absence of clonal cells in the bone ^marrow by immunohistochemistry or immunofluorescence ⁰ CR Negative immunofixation in serum and urine and disappearance of any soft tissue plasma cells and <5% of plasma cells in the bone marrow ^b VGPR Serum and urinary M protein detected by immunofixation but not electrophoresis, or a 90% or greater decrease in serum M protein plus urinary M protein <100 mg in 24 hours

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Response subcategory Answer criteria ^a PR A decrease in serum M-protein levels by >50% and a decrease in 24-hour urine M-protein levels by >90% or to <200 mg per day. If serum and urine M protein are not measurable, ^d a >50% reduction in the difference between involved and uninvolved FLC levels is required instead of M protein criteria. If serum and urine M protein are not measurable and the serum-free light assay is also not measurable, a >50% reduction in plasma cells is required instead of M protein, provided that the baseline percentage of bone marrow plasma cells was >30%. In addition to the above criteria, if present at baseline, a >50% reduction in the size of soft tissue plasmacytomas is also required. SD (not recommended for use as an indicator of response; disease stability is best described by indicating time to assess progression) Not meeting criteria for CR, VGPR, PR or progressive disease

Abbreviations: CR - complete answer; FLC, free light chain; PR - partial response; SD stable disease; sCR=strict complete response; VGPR is a very good partial answer.

^a All response categories require two consecutive assessments made at any time before the start of any new therapy; All categories also do not require known evidence of progressive or new bone disease if radiographic studies were performed. Radiographic examinations are not required to satisfy these response requirements.

^b Confirmation by repeat bone marrow biopsy is not required.

^c The presence/absence of clonal cells is based on the κ/λ ratio. An abnormal κ/λ ratio by immunohistochemistry and/or immunofluorescence requires a minimum of 100 plasma cells for analysis. The abnormal ratio, reflecting the presence of an abnormal clone, is κ/λ>4:1 or <1:2.

^d Measurable disease defined by at least one of the following: bone marrow plasma cells >30%; Serum M-protein >1 g/dL (>10 g/L) [10 g/L]; M-protein in urine >200 mg/24 h; Serum FLC assay: involved FLC level >10 mg/dL (>100 mg/L); provided that the serum FLC ratio is abnormal.

In this context, ECOG status refers to the Eastern Cooperative Oncology Group (ECOG) performance status (Oken M, et al Toxicity and response criteria of the Eastern Cooperative Oncology Group. Am J Clin Oncol 1982;5(6):649-655), as shown below:

Rating scale External characteristics 0 Fully active, able to perform all pre-disease activities without restrictions. 1 Limited in physically strenuous activities, but ambulatory and able to perform light or sedentary work, such as light housework, office work. 2 Ambulatory and capable of self-care, but unable to perform any work. Wakeful and more than 50% of the time awake. 3 Capable of only limited self-care and confined to bed or chair more than 50% of waking hours. 4 Completely disabled. Cannot perform self-care. Completely confined to a bed or chair 5 Dead

Unless otherwise indicated, to the extent there is a discrepancy between the depicted chemical structure of a compound presented herein and the chemical name of the compound presented herein, the chemical structure shall control.

5.2 Pharmaceutical compositions containing compound 1

In a specific embodiment, provided herein are pharmaceutical compositions (e.g., oral dosage forms) containing Compound 1:

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1, or an enantiomer, mixture of enantiomers, tautomer, isotopologue or pharmaceutically acceptable salt and carrier or diluent thereof.

In some embodiments, the pharmaceutical compositions provided herein are suitable for oral administration to a patient. In one embodiment, the pharmaceutical compositions provided herein exhibit beneficial physical and/or pharmacological properties. Such properties include, but are not limited to, ease of analysis, uniformity of content, flow properties for production, dissolution and bioavailability, and stability. In one embodiment, the pharmaceutical compositions provided herein have a shelf life of at least about 6 months, at least about 12 months, at least about 18 months, at least about 24 months, at least at least about 30 months, or at least about 36 months, without refrigeration. In some embodiments, no refrigeration refers to a temperature of 20°C or higher. In one embodiment, the pharmaceutical compositions provided herein are stored refrigerated. In one embodiment, the pharmaceutical compositions provided herein have a shelf life of at least about 6 months, at least about 12 months, at least about 18 months, at least about 24 months, at least at least about 30 months, or at least about 36 months if stored refrigerated. In one embodiment, the properties of the pharmaceutical compositions provided herein make them suitable for immediate release (IR).

The pharmaceutical compositions provided herein may be formulated as suitable pharmaceutical formulations such as solutions, suspensions, tablets, dispersible tablets, pills, capsules, powders, sustained release formulations or elixirs, for oral administration or as sterile solutions or suspensions for ophthalmic or parenteral administration, as well as preparations for transdermal patches and dry powder inhalers. Typically, the compounds described above are formulated into pharmaceutical compositions using techniques and procedures well known in the art (see, for example, Ansel Introduction to Pharmaceutical Dosage Forms, Seventh Edition 1999). In one embodiment, the pharmaceutical compositions provided herein are in oral dosage forms. In one embodiment, the oral dosage unit form is a tablet. In one embodiment, the oral dosage unit form is a caplet. In one embodiment, the oral dosage unit form is a capsule. In one embodiment, the pharmaceutical compositions provided herein are immediate release capsules. In one embodiment, the pharmaceutical compositions provided herein are immediate release (IR) capsule capsules (BIC).

Tablets, caplets and capsules typically contain from about 50 mg to about 500 mg of pharmaceutical composition (ie active ingredient and excipient(s)). Capsules can be of any size. Examples of standard sizes include No. 000, No. 00, No. 0, No. 1, No. 2, No. 3, No. 4, and No. 5. See, for example, Remington's Pharmaceutical Sciences,^. 1658-1659 (Alfonso Gennaro ed., Mack Publishing Company, Easton Pennsylvania, 18th ed., 1990), which is incorporated by reference. In some embodiments, capsules provided herein are size #1 or larger, #2 or larger, #3 or larger, or #4 or larger.

In the compositions, effective concentrations of one or more compounds or pharmaceutically acceptable salts are admixed with a suitable pharmaceutical carrier or carrier medium. In some embodiments, the concentrations of the compounds in the compositions are effective to deliver an amount when administered that treats, prevents, or alleviates one or more symptoms and/or progression of multiple myeloma.

Forms of connection 1

Compound 1 has the chemical name ^)-4-(4-(4-(((2-(2,6-dioxopiperidin-3-yl)-1oxoisoindolin-4-yl)oxy)methyl)benzyl)piperazin-1-yl )-3-fluorobenzonitrile. The method for preparing Compound 1 is described in US Application No. 16/030695, which is incorporated herein in its entirety.

- 9 046729 via link.

In one embodiment, Compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof is provided in a solid form in the pharmaceutical composition. In one embodiment, the solid form is amorphous. In one embodiment, the solid form is crystalline. In one embodiment, the solid form is a hydrate. In one embodiment, the solid form is anhydrous. In one embodiment, the solid form is a solvate. In one embodiment, the solid form is non-solvable.

Solid forms can be characterized using a number of techniques known to one of ordinary skill in the art, including, but not limited to, single crystal X-ray diffraction, powder X-ray diffraction (PXRD), microscopy (e.g., optical microscopy, scanning electron microscopy (SEM)) , thermal analysis (e.g. differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and hot stage microscopy), dynamic vapor sorption (DVS), spectroscopy (e.g. infrared, Raman and nuclear magnetic resonance), high performance liquid chromatography (HPLC ). The particle size and particle size distribution of the solid form presented herein can be determined by conventional methods such as laser light scattering.

In one embodiment, the pharmaceutical composition contains the free base of Compound 1. In one embodiment, the free base of Compound 1 is amorphous. In one embodiment, the free base of compound 1 is crystalline. In one embodiment, the free base of compound 1 is a mixture of one or more amorphous and crystalline forms.

In one embodiment, the pharmaceutical composition contains a salt of Compound 1. In one embodiment, the salt is a hydrochloride salt, a mesylate salt, a hydrobromide salt, a besylate salt, a glycolate salt, an L-malate salt, a napadisylate salt, a sulfate salt, a tosylate salt, an oxalate salt, isethionate salt, maleate salt, phosphate salt, malonate salt, gentisate salt, L-tartrate salt, fumarate salt, citrate salt, R-mandelate salt, L-ascorbate salt, succinate salt, nitrate salt, salicylate salt, edisylate salt, cyclamate salt, esylate salt salt, D-glucuronate salt, 4-aminosalicylate salt, caproate salt, cinnamon salt, caprylate salt, camphor salt, D-aspartate salt or D-glutamate salt. In one embodiment, the salt of Compound 1 is amorphous. In one embodiment, the salt of Compound 1 is crystalline. In one embodiment, the salt of Compound 1 is a mixture of one or more amorphous and crystalline forms.

In one embodiment, the pharmaceutical composition contains a hydrochloride salt of Compound 1. In one embodiment, the pharmaceutical composition contains a mesylate salt of Compound 1. In one embodiment, the pharmaceutical composition contains a hydrobromide salt of Compound 1. In one embodiment, the pharmaceutical composition contains a besylate salt of Compound 1. In one embodiment, the pharmaceutical composition contains a hydrochloride salt of Compound 1. in an embodiment, the pharmaceutical composition contains a glycolate salt of Compound 1. In one embodiment, the pharmaceutical composition contains an L-malate salt of Compound 1.

In one embodiment, the pharmaceutical composition contains Form K of the free base of Compound 1, Form K' of the free base of Compound 1, or an intermediate form between Form K and Form K', or a mixture thereof.

In one embodiment, Form K is a channel hydrate of the free base of Compound 1. In one embodiment, Form K is a monohydrate of the free base of Compound 1. In one embodiment, Form K' is a dehydrated hydrate of Form K. In one embodiment, without limitation According to any particular theory, the K' form is converted to the K form as humidity increases, and the K form is converted to the K' form as humidity decreases. Accordingly, intermediate forms between form K and form K' exist depending on the degree of moisture. In one embodiment, form K converts to form K' when the water activity does not exceed about 0.11. In one embodiment, Form K' is converted to Form K when the water activity is at least about 0.17.

In one embodiment, the pharmaceutical composition provided herein contains Form K, Form K', or an intermediate form between Form K and Form K', or a mixture thereof, of the free base of Compound 1, characterized by an XRD pattern containing peaks at about 14.6 , 18.2 and 18.3° 2θ. In one embodiment, the XRD pattern further contains peaks at about 22.3 and 23.1° 2θ. In one embodiment, the XRD pattern further contains peaks at about 20.5 and 20.9° 2θ. In one embodiment, the XRD pattern contains peaks at about 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 20.5, 20.9, 22.3, and 23.1° 2θ. In one embodiment, the pharmaceutical composition provided herein contains

- 10 046729 Form K of the free base of compound 1, characterized by an XRD pattern further containing at least a peak at about 14.2, 18.6 or 20.3° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base form K' of Compound 1, characterized by a pXRD pattern further comprising at least a peak at about 18.0 or 18.8° 2θ.

A typical X-ray diffraction pattern of form K is shown in Fig. 1.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 , 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or all peaks located around the following positions: 8.6, 10.8, 14.2, 14.3, 14, 6, 16.6, 17.3, 17.5, 18.2, 18.3, 18.6, 20.3, 20.5, 20.9, 21.8, 22.3, 22.5, 23.1, 24.5, 25.1, 25.7, 26.0, 27.4, 27.9 and 31.4° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 , 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or all peaks located around the following positions: 8.59, 10.78, 14.21, 14.32, 14, 60, 16.55, 17.26, 17.45, 18.21, 18.34, 18.62, 20.25, 20.47, 20.87, 21.79, 22.28, 22.45, 23.05, 24.54, 25.05, 25.67, 26.01, 27.43, 27.89 and 31.44° 2θ. In one embodiment, the solid form has 3 peaks. In one embodiment, the solid form has 5 peaks. In one embodiment, the solid form has 7 peaks. In one embodiment, the solid form has 9 peaks. In one embodiment, the solid form has 11 peaks. In one embodiment, the solid form is characterized by all peaks.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at about 14.2, 14.6, 18.2 and 18.3° 2θ. In one embodiment, the pXRD pattern further contains peaks at about 22.3, 23.1, and 24.5° 2θ. In one embodiment, the pXRD pattern further contains peaks at about 20.5 and 20.9° 2θ. In one embodiment, the xXRD pattern contains peaks at about 8.6, 14.2, 14.3, 14.6, 16.6, 18.2, 18.3, 20.5, 20.9, 22.3, 23.1, 24.5 and 26.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.8° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern including peaks at 14.2, 14.6, 18.2 and 18.3° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.6, 14.2, 14.3, 14.6, 16.6, 18.2, 18.3, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ ± 0.04° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.21, 14.60, 18.21 and 18.34° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.21, 14.32, 14.60, 16.55, 18.21, 18.34, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ ± 0.04° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern including peaks at 14.2, 14.6, 18.2 and 18.3° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ ± 0.02° 2θ. In one embodiment, the dXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ ± 0.02° 2θ. In one embodiment, the pSAR pattern contains peaks at 8.6, 14.2, 14.3, 14.6, 16.6, 18.2, 18.3, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern does not contain a peak at 18.8° 2θ ± 0.02° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern including peaks at 14.21, 14.60, 18.21 and 18.34° 2θ ± 0 .02° 2θ. In one

- 11 046729 embodiment, the pXRD diffraction pattern additionally contains peaks at 22.28, 23.05 and 24.54° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.21, 14.32, 14.60, 16.55, 18.21, 18.34, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ ± 0.02° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.2, 14.6, 18.2 and 18.3° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ. In one embodiment, the XRD pattern contains peaks at 8.6, 14.2, 14.3, 14.6, 16.6, 18.2, 18.3, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.21, 14.60, 18.21 and 18.34° 2θ. In one embodiment, the xXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.21, 14.32, 14.60, 16.55, 18.21, 18.34, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at approximately 14.6, 18.2, 18.3 and 18.6° 2θ. In one embodiment, the pXRD pattern further contains peaks at about 22.3, 23.1, and 24.5° 2θ. In one embodiment, the XRD pattern further contains peaks at about 20.5 and 20.9° 2θ. In one embodiment, the XRD pattern contains peaks at about 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 18.6, 20.5, 20.9, 22.3, 23.1, 24.5 and 26.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.8° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.2, 18.3 and 18.6° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 18.6, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ ± 0.04° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.60, 18.21, 18.34 and 18.62° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.32, 14.60, 16.55, 18.21, 18.34, 18.62, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ ± 0.04° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.2, 18.3 and 18.6° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ ± 0.02° 2θ. In one embodiment, the dXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ ± 0.02° 2θ. In one embodiment, the pSAR pattern contains peaks at 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 18.6, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ ± 0.02° 2θ. In one version

- 12 046729 implementation of the pharmaceutical composition presented in this document contains the free base of compound 1, which is a solid form characterized by an XRD diffraction pattern containing peaks at 14.60, 18.21, 18.34 and 18.62° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.32, 14.60, 16.55, 18.21, 18.34, 18.62, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ ± 0.02° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.2, 18.3 and 18.6° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ. In one embodiment, the XRD pattern contains peaks at 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 18.6, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.60, 18.21, 18.34 and 18.62° 2θ. In one embodiment, the xXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.32, 14.60, 16.55, 18.21, 18.34, 18.62, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at about 14.6, 18.2, 18.3 and 20.3° 2θ. In one embodiment, the pXRD diffraction pattern further contains peaks at about 22.3, 23.1, and 24.5° 2θ. In one embodiment, the XRD pattern further contains peaks at about 20.5 and 20.9° 2θ. In one embodiment, the xXRD pattern contains peaks at about 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 20.3, 20.5, 20.9, 22.3, 23.1, 24.5 and 26.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.8° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.2, 18.3 and 20.3° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 20.3, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ ± 0.04° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.60, 18.21, 18.34 and 20.25° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.32, 14.60, 16.55, 18.21, 18.34, 20.25, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ ± 0.04° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.2, 18.3 and 20.3° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ ± 0.02° 2θ. In one embodiment, the dXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ ± 0.02° 2θ. In one embodiment, the pSAR pattern contains peaks at 8.6.

- 13 046729

14.3, 14.6, 16.6, 18.2, 18.3, 20.3, 20.5, 20.9, 22.3, 23.1, 24.5 and 26.0° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern does not contain a peak at 18.8° 2θ ± 0.02° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.60, 18.21, 18.34 and 20.25° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.47 and 20.87° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.32, 14.60, 16.55, 18.21, 18.34, 20.25, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ ± 0.02° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.2, 18.3 and 20.3° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.3, 23.1, and 24.5° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.5 and 20.9° 2θ. In one embodiment, the XRD pattern contains peaks at 8.6, 14.3, 14.6, 16.6, 18.2, 18.3, 20.3, 20.5, 20.9, 22.3, 23 ,1, 24.5 and 26.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.0° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.8° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.60, 18.21, 18.34 and 20.25° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 22.28, 23.05, and 24.54° 2θ. In one embodiment, the pXRD diffraction pattern additionally contains peaks at 20.47 and 20.87° 2θ. In one embodiment, the XRD pattern contains peaks at 8.59, 14.32, 14.60, 16.55, 18.21, 18.34, 20.25, 20.47, 20.87, 22.28, 23 .05, 24.54 and 26.01° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.75° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of Compound 1, which is a solid form exhibiting an xXRD pattern that matches the xXRD pattern shown in FIG. 1.

A typical X-ray diffraction pattern of form K' is shown in Fig. 2.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 , 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or all peaks located near the following positions: 8.7, 10.8, 14.4, 14.6, 16 ,6, 17.4, 17.5, 18.0, 18.3, 18.4, 18.8, 20.5, 20.9, 21.8, 22.4, 22.6, 23.2 , 24.7, 25.2, 25.8, 26.2, 26.4, 27.5, 28.1, 31.7 and 38.4° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 , 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or all peaks located near the following positions: 8.65, 10.79, 14.36, 14.63, 16 .55, 17.35, 17.53, 18.02, 18.25, 18.40, 18.75, 20.52, 20.92, 21.81, 22.36, 22.64, 23.19 , 24.68, 25.20, 25.82, 26.17, 26.39, 27.54, 28.08, 31.69 and 38.41° 2θ. In one embodiment, the solid form has 3 peaks. In one embodiment, the solid form has 5 peaks. In one embodiment, the solid form has 7 peaks. In one embodiment, the solid form has 9 peaks. In one embodiment, the solid form has 11 peaks. In one embodiment, the solid form is characterized by all peaks.

In one embodiment, the pharmaceutical composition provided herein contains the free base of Compound 1, which is a solid form characterized by an XRD pattern containing peaks at about 14.6, 18.0, 18.3, and 18.4° 2θ. In one embodiment, the pXRD diffraction pattern further contains peaks at about 20.9, 22.4, and 23.2° 2θ. In one embodiment, the pXRD pattern further contains peaks at about 16.6 and 20.5° 2θ. In one embodiment, the XRD pattern contains peaks at about 8.7, 14.4, 14.6, 16.6, 18.0, 18.3, 18.4, 20.5, 20.9, 22.4, 23.2 and 24.7° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 14.2° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.6° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 20.3° 2θ.

In one embodiment, the pharmaceutical composition provided herein is

- 14 046729 contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.0, 18.3 and 18.4° 2θ ± 0.04° 2θ.

In one embodiment, the xXRD pattern additionally contains peaks at 20.9, 22.4, and

23.2° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.6 and 20.5° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at

8.7, 14.4, 14.6, 16.6, 18.0, 18.3, 18.4, 20.5, 20.9, 22.4, 23.2 and 24.7° 2θ ± 0.04° implementations implementations implementations base pXRD diffraction pattern does not contain a peak at 14.2°2θ pXRD diffractogram does not contain a peak at 18.6°2θ pXRD diffractogram does not contain a peak at 20.3°2θ pharmaceutical composition presented in this

0.04°

0.04°

0.04°

2θ.

2θ.

2θ.

2θ.

IN

IN

IN

In one embodiment, the document contains free compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.63, 18.02, 18.25 and 18.40° 2θ ± 0.04 °2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.92, 22.36, and 23.19° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.55 and 20.52° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.65, 14.36, 14.63, 16.55, 18.02, 18.25, 18.40, 20.52, 20.92, 22.36, 23 .19 and 24.68° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.21° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.62° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.25° 2θ ± 0.04° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.0, 18.3 and 18.4° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.9, 22.4, and 23.2° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.6 and 20.5° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at

8.7, 14.4, 14.6, 16.6, 18.0, 18.3, 18.4, 20.5, 20.9, 22.4, 23.2 and 24.7° 2θ ± 0.02° implementations implementations implementations base X-ray diffraction pattern does not contain a peak at 14.2°2θ X-ray diffraction pattern does not contain a peak at 18.6°2θ X-ray diffraction pattern does not contain a peak at 20.3°2θ pharmaceutical composition presented in this

0.02°

0.02°

0.02°

2θ.

2θ.

2θ.

2θ.

IN

IN

IN

In one embodiment, the document contains free compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.63, 18.02, 18.25 and 18.40° 2θ ± 0.02 °2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.92, 22.36, and 23.19° 2θ ± 0.02° 2θ. In one embodiment, the pXRD diffraction pattern additionally contains peaks at 16.55 and

20.52° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at 8.65, 14.36, 14.63, 16.55, 18.02, 18.25, 18.40, 20.52, 20.92, 22.36, 23 .19 and 24.68° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.21° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.62° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.25° 2θ ± 0.02° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.0, 18.3 and 18.4° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.9, 22.4, and 23.2° 2θ. In one embodiment, the pXRD diffraction pattern additionally contains peaks at 16.6 and 20.5° 2θ. In one embodiment, the XRD pattern contains peaks at 8.7, 14.4, 14.6, 16.6, 18.0, 18.3, 18.4, 20.5, 20.9, 22.4, 23 .2 and 24.7° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.2° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.6° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.3° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.63, 18.02, 18.25 and 18.40° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.92, 22.36, and 23.19° 2θ. In one embodiment, the pXRD diffraction pattern additionally contains peaks at 16.55 and 20.52° 2θ. In one embodiment, the XRD pattern contains peaks at 8.65, 14.36, 14.63, 16.55, 18.02, 18.25, 18.40, 20.52, 20.92, 22.36, 23 .19 and 24.68° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.21° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.62° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.25° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of Compound 1, which is a solid form,

- 15 046729 characterized by a XRD pattern containing peaks at about 14.6, 18.3, 18.4 and 18.8° 2θ. In one embodiment, the pXRD diffraction pattern further contains peaks at about 20.9, 22.4, and 23.2° 2θ. In one embodiment, the pXRD pattern further contains peaks at about 16.6 and 20.5° 2θ. In one embodiment, the XRD pattern contains peaks at about 8.7, 14.4, 14.6, 16.6, 18.3, 18.4, 18.8, 20.5, 20.9, 22.4, 23.2 and 24.7° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 14.2° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 18.6° 2θ. In one embodiment, the XRD pattern does not contain a peak at about 20.3° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.3, 18.4 and 18.8° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.9, 22.4, and 23.2° 2θ ± 0.04° 2θ. In one embodiment, the dXRD pattern additionally contains peaks at 16.6 and 20.5° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.7, 14.4, 14.6, 16.6, 18.3, 18.4, 18.8, 20.5, 20.9, 22.4, 23 .2 and 24.7° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.2° 2θ ± 0.04° 2θ. In one embodiment, the pSAR pattern does not contain a peak at 18.6° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.3° 2θ ± 0.04° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.63, 18.25, 18.40 and 18.75° 2θ ± 0 .04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.92, 22.36, and 23.19° 2θ ± 0.04° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.55 and 20.52° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern contains peaks at 8.65, 14.36, 14.63, 16.55, 18.25, 18.40, 18.75, 20.52, 20.92, 22.36, 23 .19 and 24.68° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.21° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.62° 2θ ± 0.04° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.25° 2θ ± 0.04° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.3, 18.4 and 18.8° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.9, 22.4, and 23.2° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.6 and 20.5° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at

8.7, 14.4, 14.6, 16.6, 18.3, 18.4, 18.8, 20.5, 20.9, 22.4, 23.2 and 24.7° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.2° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.6° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.3° 2θ ± 0.02° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.63, 18.25, 18.40 and 18.75° 2θ ± 0 .02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.92, 22.36, and 23.19° 2θ ± 0.02° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.55 and 20.52° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern contains peaks at 8.65, 14.36, 14.63, 16.55, 18.25, 18.40, 18.75, 20.52, 20.92, 22.36, 23 .19 and 24.68° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.21° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.62° 2θ ± 0.02° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.25° 2θ ± 0.02° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of compound 1, which is a solid form characterized by an XRD pattern containing peaks at 14.6, 18.3, 18.4 and 18.8° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.9, 22.4, and 23.2° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.6 and 20.5° 2θ. In one embodiment, the XRD pattern contains peaks at 8.7, 14.4, 14.6, 16.6, 18.3, 18.4, 18.8, 20.5, 20.9, 22.4, 23 .2 and 24.7° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.2° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.6° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.3° 2θ. In one embodiment, the pharmaceutical composition provided herein contains the free base of Compound 1, which is a solid form characterized by

- 16 046729 XRD pattern containing peaks at 14.63, 18.25, 18.40 and 18.75° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 20.92, 22.36, and 23.19° 2θ. In one embodiment, the pXRD pattern additionally contains peaks at 16.55 and 20.52° 2θ. In one embodiment, the XRD pattern contains peaks at 8.65, 14.36, 14.63, 16.55, 18.25, 18.40, 18.75, 20.52, 20.92, 22.36, 23 .19 and 24.68° 2θ. In one embodiment, the XRD pattern does not contain a peak at 14.21° 2θ. In one embodiment, the XRD pattern does not contain a peak at 18.62° 2θ. In one embodiment, the XRD pattern does not contain a peak at 20.25° 2θ.

In one embodiment, the pharmaceutical composition provided herein contains the free base of Compound 1, which is a solid form exhibiting an xXRD pattern that matches the xXRD pattern shown in FIG. 2.

In one embodiment, and without being limited to any particular theory, compared to Form K, the pXRD peaks of Form K' are slightly shifted toward higher °2θ values, suggesting that Form K' has a slightly compressed lattice

In one embodiment, XRD patterns are obtained using Cu Kα radiation.

Pharmaceutical composition based on mannitol

In one embodiment, the carrier or diluent in the pharmaceutical composition provided herein is mannitol.

In one embodiment, the pharmaceutical composition further comprises a glidant, an acidifier, a lubricant, or a mixture thereof.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof in an amount of from about 0.05 to about 3% w/w; 2) mannitol in an amount of from about 80 to about 98% w/w; 3) a glidant in an amount of from about 0 to about 10% w/w; 4) an acidifier in an amount of from about 0 to about 6% w/w; and 5) a lubricant in an amount of from about 0 to about 8% wt./wt.

In one embodiment, Compound 1, or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof, is the free base of Compound 1. In one embodiment, the free base of Compound 1 is the crystalline free base of Compound 1. In one embodiment, the free base of Compound 1 1 is characterized by a pSAR pattern containing peaks at approximately 14.6, 18.2 and 18.3° 2θ.

In one embodiment, the amount of Compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof is from about 0.05 to about 3% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount is from about 0.05 to about 2% w/w. In one embodiment, the amount is from about 0.1 to about 1.5% w/w. In one embodiment, the amount is from about 0.13 to about 1% w/w. In one embodiment, the amount is from about 0.13 to about 0.5% w/w. In one embodiment, the amount is from about 0.5 to about 1% w/w.

In one embodiment, the amount of Compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof is about 0.05, about 0.06, about 0.07, about 0.08, about 0.09, about 0. 1, about 0.11, about 0.12, about 0.13, about 0.14, about 0.15, about 0.16, about 0.17, about 0.18, about 0.19, about 0, 2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, about 0.55, about 0.6, about 0.65, about 0, 7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1, about 1.1, about 1.2, about 1.3, about 1.4, about 1.5, about 1.6, about 1.7, about 1.8, about 1.9, about 2, about 2.1, about 2.2, about 2.3, about 2.4, about 2 .5, about 2.6, about 2.7, about 2.8, about 2.9 or about 3% w/w. In one embodiment, the amount is about 0.13% w/w. In one embodiment, the amount is about 0.5% w/w. In one embodiment, the amount is about 1% w/w.

In one embodiment, the mannitol is spray dried. In one embodiment, mannitol has a crystalline grade of 200 SD (eg, perlitol 200 SD). In one embodiment, mannitol has a crystalline grade of 100 SD (eg, perlitol 100 SD).

In one embodiment, the amount of mannitol is from about 80 to about 98% wt./wt. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of mannitol is from about 85 to about 95% wt./wt. In one embodiment, the amount of mannitol is from about 90 to about 93% w/w.

In one embodiment, the amount of mannitol is about 80, about 81, about 82, about 83, about 84, about 85, about 86, about 87, about 88, about 88.5, about 89, about 89.5, about 90, about 90.5, about 91, about 91.5, about 92, about 92.5, about 93, about 94, about 95, about 96, about 97 or about 98% w/w. In one embodiment, the amount of mannitol is about 90% w/w. In one embodiment, the amount of mannitol is about 91.5% w/w. In one version

- 17 046729 sales the amount of mannitol is approximately 92.4% wt./wt.

In one embodiment, the glidant is silica dimethyl silylate or fumed silica. In one embodiment, the glidant is silica dimethyl silylate. In one embodiment, the glidant is colloidal silica.

In one embodiment, the glidant is a hydrophobic glidant. In one embodiment, the glidant is Aerosil R972.

In one embodiment, the glidant is a hydrophilic glidant. In one embodiment, the glidant has a surface area of about 200 m ² /g. In one embodiment, the glidant has a surface area of about 300 m ² /g. In one embodiment, the glidant is Aerosil 200. In one embodiment, the glidant is Aerosil 300.

In one embodiment, the amount of glidant is from about 0 to about 10% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of glidant is from about 0 to about 4% w/w. In one embodiment, the amount of glidant is from about 0.25 to about 3% w/w. In one embodiment, the amount of glidant is from about 0.5 to about 2% w/w.

In one embodiment, the amount of glidant is about 0, about 0.1, about 0.2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.9, about 1, about 1.1, about 1, 2, about 1.3, about 1.4, about 1.5, about 1.75, about 2, about 2.25, about 2.5, about 2.75, about 3, about 3.25, about 3 .5, about 3.75, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about 9 , about 9.5 or about 10% w/w. In one embodiment, the amount of glidant is about 0.5% w/w. In one embodiment, the amount of glidant is about 1% w/w. In one embodiment, the amount of glidant is about 2% w/w. In one embodiment, the amount of glidant is from about 0.5 to about 1% w/w.

In one embodiment, the glidant is silica dimethyl silylate in an amount of about 0.5% w/w. In one embodiment, the glidant is colloidal silica in an amount of about 1% w/w. In one embodiment, the glidant is colloidal silica in an amount of about 2% w/w.

In one embodiment, the pharmaceutical compositions provided herein contain an acidifying agent in an amount sufficient to maintain the pH of the pharmaceutical composition no higher than 5. In one embodiment, without being limited by theory, Compound 1 may degrade at a pH higher than 5.

In one embodiment, the pharmaceutical compositions provided herein contain an acidifying agent in an amount sufficient to maintain the pH of the pharmaceutical composition no greater than 4. In one embodiment, the pharmaceutical compositions provided herein contain an acidifying agent in an amount sufficient to maintain the pH of the pharmaceutical composition from about 2 to about 3.

In one embodiment, the acidulant is fumaric acid. In one embodiment, the acidulant is maleic acid. In one embodiment, the acidulant is succinic acid.

In one embodiment, the amount of acidifier is from about 0 to about 6% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of acidifier is from about 2 to about 5% w/w. In one embodiment, the amount of acidifier is from about 2 to about 4% w/w.

In one embodiment, the amount of acidifier is about 0, about 0.5, about 1, about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 5.5 or about 6% w/w. In one embodiment, the amount of acidifier is about 3% w/w.

In one embodiment, the acidulant is fumaric acid in an amount of about 3% w/w.

In one embodiment, the lubricant is stearic acid. In one embodiment, the lubricant is magnesium stearate.

In one embodiment, the amount of lubricant is from about 0 to about 8% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of lubricant is from about 2 to about 6% w/w.

- 18 046729

In one embodiment, the amount of lubricant is about 0, about 0.5, about 1, about

1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about

7, about 7.5 or about 8% w/w. In one embodiment, the amount of lubricant is about 4% w/w.

In one embodiment, the lubricant is stearic acid in an amount of about 4% w/w.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 (eg, Form K) in an amount of about 0.1 to about 0.2% w/w; 2) mannitol in an amount of from about 91 to about 93% w/w; 3) silica dimethyl silylate in an amount of from about 0.4 to about 0.6% w/w; 4) fumaric acid in an amount of from about 2 to about 4% w/w; and 5) stearic acid in an amount of from about 3 to about 5% w/w. In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 (eg, Form K) in an amount of about 0.13% w/w; 2) mannitol in an amount of about 92.37% w/w; 3) silica dimethyl silylate in an amount of about 0.5% w/w; 4) fumaric acid in an amount of about 3% w/w; and 5) stearic acid in an amount of about 4% wt./wt. In one embodiment, the pharmaceutical composition has a total weight of about 75 mg. In one embodiment, the pharmaceutical composition is contained in a size 4 capsule.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 (eg, Form K) in an amount of about 0.4 to about 0.6% w/w; 2) mannitol in an amount of from about 90.5 to about 92.5% w/w; 3) colloidal silica in an amount of from about 0.8 to about 1.2% w/w; 4) fumaric acid in an amount of from about 2 to about 4% w/w; and 5) stearic acid in an amount of from about 3 to about 5% w/w. In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 (eg, Form K) in an amount of about 0.5% w/w; 2) mannitol in an amount of about 91.5% w/w; 3) colloidal silicon dioxide in an amount of about 1% w/w; 4) fumaric acid in an amount of about 3% w/w; and 5) stearic acid in an amount of about 4% w/w; In one embodiment, the pharmaceutical composition has a total weight of about 100 mg. In one embodiment, the pharmaceutical composition is contained in a size 3 capsule.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 (eg, Form K) in an amount of from about 0.8 to about 1.2% w/w; 2) mannitol in an amount of from about 89 to about 91% w/w; 3) colloidal silica in an amount of from about 1.8 to about 2.2% w/w; 4) fumaric acid in an amount of from about 2 to about 4% w/w; and 5) stearic acid in an amount of from about 3 to about 5% w/w. In one embodiment, provided herein is a pharmaceutical composition comprising: 1) Compound 1 (eg, Form K) in an amount of about 1% w/w; 2) mannitol in an amount of about 90% w/w; 3) colloidal silicon dioxide in an amount of about 2% w/w; 4) fumaric acid in an amount of about 3% w/w; and 5) stearic acid in an amount of about 4% wt./wt. In one embodiment, the pharmaceutical composition has a total weight of about 200 mg. In one embodiment, the pharmaceutical composition is contained in a size 2 capsule.

Pharmaceutical composition based on starch and lactose

In one embodiment, the carrier or diluent in the pharmaceutical composition provided herein is a mixture of starch and lactose.

In one embodiment, the pharmaceutical composition further comprises a glidant, an acidifier, a lubricant, a disintegrant, or a mixture thereof.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof in an amount of from about 0.05 to about 3% w/w; 2) a mixture of starch and lactose in an amount of from about 70 to about 90% w/w; 3) a glidant in an amount of from about 0 to about 10% w/w; 4) an acidifier in an amount of from about 0 to about 8% w/w; 5) lubricant in an amount of from about 0 to about 8% w/w; and 6) baking powder in an amount of from about 0 to about 20% w/w.

In one embodiment, Compound 1, or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof, is the free base of Compound 1. In one embodiment, the free base of Compound 1 is the crystalline free base of Compound 1. In one embodiment, the free base of Compound 1 1 is characterized by a pSAR pattern containing peaks at approximately 14.6, 18.2 and 18.3° 2θ.

In one embodiment, the amount of Compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof is from about 0.05 to about 3% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount is from about 0.05 to about 2% w/w. In one embodiment, the amount is from about 0.1 to about 1.5% w/w. In one embodiment, the amount is from about 0.13 to about 1% w/w. In one embodiment, the amount is from about 0.13 to about 0.5%

- 19 046729 wt./wt. In one embodiment, the amount is from about 0.5 to about 1% w/w.

In one embodiment, the amount of Compound 1 or an enantiomer, mixture of enantiomers, tautomer, isotopologue, or pharmaceutically acceptable salt thereof is about 0.05, about 0.06, about 0.07, about 0.08, about 0.09, about 0. 1, about 0.11, about 0.12, about 0.13, about 0.14, about 0.15, about 0.16, about 0.17, about 0.18, about 0.19, about 0, 2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, about 0.55, about 0.6, about 0.65, about 0, 7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1, about 1.1, about 1.2, about 1.3, about 1.4, about 1.5, about 1.6, about 1.7, about 1.8, about 1.9, about 2, about 2.1, about 2.2, about 2.3, about 2.4, about 2 .5, about 2.6, about 2.7, about 2.8, about 2.9 or about 3% w/w. In one embodiment, the amount is about 0.13% w/w. In one embodiment, the amount is about 0.5% w/w. In one embodiment, the amount is about 1% w/w.

In one embodiment, the starch is partially pregelatinized starch. In one embodiment, the starch is pregelatinized starch.

In one embodiment, the lactose is lactose anhydrous. In one embodiment, the lactose is lactose monohydrate. In one embodiment, the lactose is spray dried.

In one embodiment, the amount of the mixture of starch and lactose is from about 70 to about 90% wt./wt. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of the starch and lactose mixture is from about 75 to about 85% w/w. In one embodiment, the amount of the starch and lactose mixture is from about 80 to about 82% w/w.

In one embodiment, the amount of starch and lactose mixture is about 70, about 71, about 72, about 73, about 74, about 75, about 76, about 77, about 78, about 78.5, about 79, about 79.5, about 80, about 80.5, about 81, about 81.5, about 82, about 82.5, about 83, about 83.5, about 84, about 85, about 86, about 87, about 88, about 89 or about 90% w/w In one embodiment, the amount of starch and lactose mixture is about 80% w/w. In one embodiment, the amount of starch and lactose mixture is about 80.5% w/w. In one embodiment, the amount of starch and lactose mixture is about 81.4% w/w.

In one embodiment, the amount of starch is from about 17 to about 22% w/w, and the amount of lactose is from about 53 to about 68% w/w. In one embodiment, the amount of starch is from about 18 to about 21% w/w, and the amount of lactose is from about 57 to about 64% w/w. In one embodiment, the amount of starch is from about 19 to about 20% w/w, and the amount of lactose is from about 61 to about 62% w/w.

In one embodiment, the amount of starch is about 17, about 17.5, about 18, about 18.5, about 19, about 19.5, about 20, about 20.5, about 21, about 21.5, or about 22% wt./wt. In one embodiment, the amount of starch is about 19% w/w. In one embodiment, the amount of starch is about 19.5% w/w. In one embodiment, the amount of starch is about 20% w/w.

In one embodiment, the amount of lactose is about 53, about 54, about 55, about 56, about 57, about 58, about 58.5, about 59, about 59.5, about 60, about 60.5, about 61, about 61.5, about 62, about 62.5, about 63, about 64, about 65, about 66, about 67 or about 68% w/w. In one embodiment, the amount of lactose is about 61% w/w. In one embodiment, the amount of lactose is about 61.4% w/w.

In one embodiment, the weight ratio of starch to lactose is from about 1:2 to about 1:4. In one embodiment, the weight ratio of starch to lactose is from about 1:2.5 to about 1:3.5. In one embodiment, the starch to lactose weight ratio is from about 1:3 to about 1:3.3. In one embodiment, the weight ratio of starch to lactose is about 1:3. In one embodiment, the weight ratio of starch to lactose is about 1:3.1. In one embodiment, the weight ratio of starch to lactose is about 1:3.2. In one embodiment, the weight ratio of starch to lactose is about 1:3.3.

In one embodiment, the glidant is silica dimethyl silylate or colloidal silica. In one embodiment, the glidant is silica dimethyl silylate. In one embodiment, the glidant is colloidal silica.

In one embodiment, the glidant is a hydrophobic glidant. In one embodiment, the glidant is Aerosil R972.

In one embodiment, the glidant is a hydrophilic glidant. In one embodiment, the glidant has a surface area of about 200 m ² /g. In one embodiment, the glidant has a surface area of about 300 m ² /g. In one embodiment, the glidant is Aerosil 200. In one embodiment, the glidant is Aerosil 300.

- 20 046729

In one embodiment, the amount of glidant is from about 0 to about 10% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of glidant is from about 0 to about 4% w/w. In one embodiment, the amount of glidant is from about 0.25 to about 3% w/w. In one embodiment, the amount of glidant is from about 0.5 to about 2% w/w. In one embodiment, the amount of glidant is from about 0.5 to about 1% w/w.

In one embodiment, the amount of glidant is about 0, about 0.1, about 0.2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.9, about 1, about 1.1, about 1, 2, about 1.3, about 1.4, about 1.5, about 1.75, about 2, about 2.25, about 2.5, about 2.75, about 3, about 3.25, about 3 .5, about 3.75, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about 9 , about 9.5 or about 10% w/w. In one embodiment, the amount of glidant is about 0.5% w/w. In one embodiment, the amount of glidant is about 1% w/w. In one embodiment, the amount of glidant is about 2% w/w.

In one embodiment, the glidant is silica dimethyl silylate in an amount of about 0.5% w/w. In one embodiment, the glidant is colloidal silica in an amount of about 1% w/w. In one embodiment, the glidant is colloidal silica in an amount of about 2% w/w.

In one embodiment, the pharmaceutical compositions provided herein contain an acidifying agent in an amount sufficient to maintain the pH of the pharmaceutical composition no higher than 5. In one embodiment, without being limited to any particular theory, Compound 1 may degrade at a pH higher than 5.

In one embodiment, the pharmaceutical compositions provided herein contain an acidifying agent in an amount sufficient to maintain the pH of the pharmaceutical composition no greater than 4. In one embodiment, the pharmaceutical compositions provided herein contain an acidifying agent in an amount sufficient to maintain the pH of the pharmaceutical composition from about 2 to about 3.

In one embodiment, the acidulant is fumaric acid. In one embodiment, the acidulant is maleic acid. In one embodiment, the acidulant is succinic acid.

In one embodiment, the amount of acidifier is from about 0 to about 8% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of acidifier is from about 1 to about 7% w/w. In one embodiment, the amount of acidifier is from about 2 to about 6% w/w. In one embodiment, the amount of acidifier is from about 3 to about 5% w/w.

In one embodiment, the amount of acidifier is about 0, about 0.5, about 1, about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5 or about 8% w/w. In one embodiment, the amount of acidifier is about 4% w/w.

In one embodiment, the acidulant is fumaric acid in an amount of about 4% w/w.

In one embodiment, the lubricant is stearic acid. In one embodiment, the lubricant is magnesium stearate.

In one embodiment, the amount of lubricant is from about 0 to about 8% w/w. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of lubricant is from about 1 to about 7% w/w. In one embodiment, the amount of lubricant is from about 2 to about 6% w/w. In one embodiment, the amount of lubricant is from about 3 to about 5% w/w.

In one embodiment, the amount of lubricant is about 0, about 0.5, about 1, about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5 or about 8% w/w. In one embodiment, the amount of lubricant is about 4% w/w.

In one embodiment, the lubricant is stearic acid in an amount of about 4% w/w.

In one embodiment, the leavening agent is sodium starch glycolate. In one embodiment, the leavening agent is croscarmellose sodium. In one embodiment, the disintegrant is crospovidone.

In one embodiment, the amount of leavening agent is from about 0 to about 20%

- 21 046729 wt./wt. (based on the total weight of the pharmaceutical composition). In one embodiment, the amount of disintegrant is from about 5 to about 15% w/w. In one embodiment, the amount of disintegrant is from about 8 to about 12% w/w.

In one embodiment, the amount of leavening agent is about 0, about 2, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 18 or about 20% w/w. In one embodiment, the amount of leavening agent is about 10% w/w.

In one embodiment, the leavening agent is sodium starch glycolate in an amount of about 10% w/w. In one embodiment, the leavening agent is croscarmellose sodium in an amount of about 10% w/w. In one embodiment, the disintegrant is crospovidone in an amount of about 10% w/w.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) compound 1 (eg, Form K) in an amount of about 0.1 to about 0.2% w/w; 2) partially pregelatinized starch in an amount of from about 19 to about 21% wt./wt. and anhydrous lactose in an amount of from about 60 to about 62% w/w; 3) silica dimethyl silylate in an amount of from about 0.4 to about 0.6% w/w; 4) fumaric acid in an amount of from about 3 to about 5% w/w; 5) stearic acid in an amount of from about 3 to about 5% w/w; and 6) sodium starch glycolate in an amount of from about 9 to about 11% wt./wt. In one embodiment, provided herein is a pharmaceutical composition comprising: 1) Compound 1 (eg, Form K) in an amount of about 0.13% w/w; 2) partially pregelatinized starch in an amount of about 20% wt./wt. and anhydrous lactose in an amount of about 61.4% w/w; 3) silica dimethyl silylate in an amount of about 0.5% w/w; 4) fumaric acid in an amount of about 4% w/w; 5) stearic acid in an amount of about 4% w/w; and 6) sodium starch glycolate in an amount of about 10% w/w. In one embodiment, the pharmaceutical composition has a total weight of about 75 mg. In one embodiment, the pharmaceutical composition is contained in a size 4 capsule.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) Compound 1 (eg, Form K) in an amount of about 0.4 to about 0.6% w/w; 2) partially pregelatinized starch in an amount of from about 19 to about 20% wt./wt. and anhydrous lactose in an amount of from about 60 to about 62% w/w; 3) colloidal silica in an amount of from about 0.8 to about 1.2% w/w; 4) fumaric acid in an amount of from about 3 to about 5% w/w; 5) stearic acid in an amount of from about 3 to about 5% w/w; and 6) sodium starch glycolate in an amount of from about 9 to about 11% wt./wt. In one embodiment, provided herein is a pharmaceutical composition comprising: 1) Compound 1 (eg, Form K) in an amount of about 0.5% w/w; 2) partially pregelatinized starch in an amount of about 19.5% wt./wt. and anhydrous lactose in an amount of about 61% w/w; 3) colloidal silicon dioxide in an amount of about 1% w/w; 4) fumaric acid in an amount of about 4% w/w; 5) stearic acid in an amount of about 4% w/w; and 6) sodium starch glycolate in an amount of about 10% w/w. In one embodiment, the pharmaceutical composition has a total weight of about 100 mg. In one embodiment, the pharmaceutical composition is contained in a size 3 capsule.

In one embodiment, provided herein is a pharmaceutical composition comprising: 1) Compound 1 (eg, Form K) in an amount of about 0.8 to about 1.2% w/w; 2) partially pregelatinized starch in an amount of from about 18 to about 20% wt./wt. and anhydrous lactose in an amount of from about 60 to about 62% w/w; 3) colloidal silica in an amount of from about 0.8 to about 1.2% w/w; 4) fumaric acid in an amount of from about 3 to about 5% w/w; 5) stearic acid in an amount of from about 3 to about 5% w/w; and 6) sodium starch glycolate in an amount of from about 9 to about 11% wt./wt. In one embodiment, provided herein is a pharmaceutical composition comprising: 1) Compound 1 (eg, Form K) in an amount of about 1% w/w; 2) partially pregelatinized starch in an amount of about 19% wt./wt. and anhydrous lactose in an amount of about 61% w/w; 3) colloidal silicon dioxide in an amount of about 1% w/w; 4) fumaric acid in an amount of about 4% w/w; 5) stearic acid in an amount of about 4% w/w; and 6) sodium starch glycolate in an amount of about 10% w/w. In one embodiment, the pharmaceutical composition has a total weight of about 200 mg. In one embodiment, the pharmaceutical composition is contained in a size 2 capsule.

Additional embodiments of pharmaceutical compositions

In one embodiment, the pharmaceutical compositions provided herein may optionally further contain one or more additional excipients. Additional excipients include, but are not limited to, a wetting agent, a solubilizer, a crystallization stabilizer, a release agent, and a precipitation inhibitor.

In one embodiment, the pharmaceutical compositions provided herein optionally further contain one or more of the following components: Tween 80, Poloxamer 188, sodium lauryl sulfate (SLS), HPBCD, VitE-TPGS, HPMCAS-LF, HPMC E3, PVP VA64, PVP K30, HPC EXF and talc.

- 22 046729

In one embodiment, the pharmaceutical compositions provided herein are formulated in capsule form. In one embodiment, the capsule is an HPMC capsule.

In one embodiment, the capsule is a gelatin capsule.

Typically the compositions are formulated for single dose administration. To formulate the composition, a mass fraction of the compound is dissolved, suspended, dispersed or otherwise mixed in a selected vehicle at an effective concentration such that the condition being treated is alleviated or improved. Pharmaceutical carriers or carrier media suitable for administration of the compounds presented herein include any such carriers known to those skilled in the art as suitable for the particular route of administration.

In addition, the compounds may be formulated as the sole pharmaceutically active ingredient in a composition or may be combined with other active ingredients. Liposome suspensions, including tissue-targeted liposomes, such as tumor-targeted liposomes, may also be suitable as pharmaceutically acceptable carriers. They can be prepared by methods known to those skilled in the art. For example, liposome formulations can be prepared as known in the art. Briefly, liposomes such as multilayer vesicles (MLVs) can be formed by drying egg phosphatidylcholine and brain phosphatidylserine (7:3 molar ratio) inside a flask. Add a solution of the compound provided herein in divalent cation-free phosphate buffered saline (PBS) and shake the flask until the lipid film is dispersed. The resulting vesicles are washed to remove unencapsulated compound, pelleted by centrifugation and then resuspended in PBS.

The active compound is included in a pharmaceutically acceptable carrier in an amount sufficient to provide a therapeutically beneficial effect without causing undesirable side effects in the patient being treated. The therapeutically effective concentration can be determined empirically by testing compounds in the in vitro and in vivo systems described herein and then extrapolated to human dosages.

The concentration of the active compound in the pharmaceutical composition will depend on the absorption, tissue distribution, inactivation, metabolism and excretion of the active compound, the physicochemical characteristics of the compound, the dosage regimen and amount administered, as well as other factors known to those skilled in the art. For example, the amount delivered is sufficient to relieve one or more symptoms of cancer, including solid tumors and blood-borne tumors.

Solutions or suspensions used for parenteral, intradermal, subcutaneous or topical use may include any of the following: a sterile diluent such as water for injection, saline, fatty vegetable oil, polyethylene glycol, glycerin, propylene glycol, dimethyl acetamide or other synthetic solvent; antimicrobial agents such as benzyl alcohol and methylparabens; antioxidants such as ascorbic acid and sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid (EDTA); buffers such as acetates, citrates and phosphates; and tone-regulating agents such as sodium chloride or dextrose. Formulations for parenteral administration may be contained in ampoules, pens, disposable syringes, or single-dose or multiple-dose vials made of glass, plastic, or other suitable material.

In cases where compounds exhibit insufficient solubility, methods for solubilizing the compounds can be used. Such methods are known to those skilled in the art and include, but are not limited to, the use of cosolvents such as dimethyl sulfoxide (DMSO), the use of surfactants such as TWEEN®, or dissolution in aqueous sodium bicarbonate.

When the compound(s) are mixed or added, the resulting mixture may be a solution, suspension, emulsion, or the like. The form of the resulting mixture depends on a number of factors, including the intended route of administration and the solubility of the compound in the selected vehicle or carrier medium. The effective concentration is sufficient to relieve the symptoms of the disease, disorder or condition being treated and can be determined empirically.

The pharmaceutical compositions are intended for administration to humans and animals in unit dosage forms such as tablets, capsules, pills, powders, granules, sterile parenteral solutions or suspensions, and oral solutions or suspensions, as well as oil-in-water emulsions, containing suitable amounts of the compounds or their pharmaceutically acceptable salts. Pharmaceutically therapeutically active compounds and their salts are formulated and administered in single dosage forms or multiple dosage forms. As used herein, unit dosage forms refer to physically discrete units suitable for use in humans and animals and individually packaged as is known in the art. Each unit dose contains a predetermined amount of a therapeutically active compound sufficient to produce the desired therapeutic effect, in combination with the necessary

- 23 046729 pharmaceutical carrier, carrier medium or diluent. Examples of unit dosage forms include ampoules and syringes, as well as individually packaged tablets or capsules. Unit dose forms can be administered in fractional or multiple doses. A multiple dose form is a plurality of identical unit dosage forms packaged in a single container for administration in separate unit dosage forms. Examples of multiple dose forms include vials, tablet or capsule bottles, or pint or gallon bottles. Therefore, a multiple dose form is a plurality of unit doses that are not separated within the package.

Dosage forms or compositions can be prepared containing the active ingredient in the range of 0.005% to 100%, with the balance being a non-toxic carrier. For oral administration, a pharmaceutically acceptable non-toxic composition is prepared by including any of the commonly used excipients, such as, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, talc, cellulose derivatives, crosscarmellose sodium, glucose, sucrose, magnesium carbonate or sodium saccharin . Such compositions include solutions, suspensions, tablets, capsules, powders and sustained release formulations such as, but not limited to, implants and microencapsulated delivery systems, as well as biodegradable, biocompatible polymers such as collagen, ethylene vinyl acetate, polyanhydrides, polyglycolic acid, polyorthoesters, polylactic acid and others. Methods for preparing these compositions are known to those skilled in the art.

The active compounds or pharmaceutically acceptable salts can be formulated with carriers that protect the compound from rapid elimination from the body, such as sustained release formulations or coatings.

The compositions may include other active compounds to provide the desired combinations of properties. The compounds provided herein, or pharmaceutically acceptable salts thereof, as described herein, may also advantageously be administered for therapeutic or prophylactic purposes together with another pharmacological agent known in the art to be valuable in the treatment of one or more diseases or medical conditions. mentioned above, such as diseases associated with oxidative stress. It should be understood that such combination therapy constitutes an additional aspect of the compositions and methods of treatment presented herein.

Manufacturing process of dosage forms

The pharmaceutical compositions (dosage forms) provided herein can be prepared by any of the pharmaceutical methods, but all methods include the step of combining the active ingredient with an excipient that constitutes one or more of the required ingredients. Typically, compositions are prepared by uniformly mixing (eg, direct mixing) the active ingredient with liquid excipients or finely divided solid excipients or both, and then, if necessary, shaping the product into the desired form (eg, compaction such as a roller compaction). If desired, tablets can be coated using standard aqueous or non-aqueous methods.

The dosage form provided herein may be prepared by compression or molding, optionally with one or more accessory ingredients. Compressed tablets can be prepared by compressing in a suitable device the active ingredient in bulk form such as powder or granules, optionally mixed with an excipient as defined above and/or a surfactant or dispersing agent. Molded tablets can be prepared by molding in a suitable machine a mixture of a powdered compound moistened with an inert liquid diluent. Encapsulation of the dosage forms presented herein can be accomplished using hydroxypropyl methylcellulose, calcium alginate, or gelatin capsules.

In some embodiments, the active ingredients and excipients are directly mixed and loaded, for example, into a capsule or compressed directly into tablets. A direct compound dosage form may be more advantageous than a compressed (e.g., roller-compressed) dosage form in certain cases because direct compound may reduce or eliminate the adverse health effects that may be caused by airborne ingredient particles during manufacturing, using compaction process. In some embodiments, direct mixing also helps minimize degradation of the active ingredient.

Direct mixing can minimize the formation of airborne particles or dust, while roller compaction processes can produce dust. In the roller compaction process, the compacted material is often crushed into smaller particles for further processing. The grinding operation can produce significant amounts of airborne particulates since the purpose of this production step is to reduce the particle size of the material. The crushed material is then mixed with other ingredients before making

- 24 046729 final dosage form.

For some active ingredients, particularly compounds with low solubility, the particle size of the active ingredient is reduced to a fine powder to help increase the rate of solubilization of the active ingredient. Increasing the rate of solubilization is often necessary for efficient absorption of the active ingredient in the gastrointestinal tract. However, for fine powders that must be directly mixed and loaded into capsules, excipients should preferably provide certain characteristics that make the ingredients suitable for the direct mixing process. Examples of such characteristics include, but are not limited to, acceptable flow characteristics. Thus, in one embodiment, provided herein are the uses and compositions containing excipients that can provide characteristics that make the resulting mixture suitable for a direct mixing process, such as good flow characteristics.

In some embodiments, the proposed dosage form or pharmaceutical composition is prepared through a process including one or more mixing and residue removal steps, followed by an encapsulation step. In some embodiments, the method further includes preparing a pretreatment mixture from one or more excipients and processing equipment used in the process with the pretreatment mixture. In one embodiment, the pretreatment mixture is prepared from one or more diluents and lubricants (eg, a mixture of mannitol and stearic acid or a mixture of lactose anhydrous and stearic acid). In one embodiment, the use of a pretreatment mixture helps minimize adhesion of the active ingredient to equipment surfaces during the manufacturing process.

In some embodiments, the proposed dosage form or pharmaceutical composition is prepared by a process comprising coating the active ingredient (eg, Compound 1) with hydrophobic silica. In one embodiment, the coating is applied by mixing the active ingredient and hydrophobic silica to form a binary mixture (before mixing with additional excipients). In one embodiment, the coating is applied by dispersing the active ingredient with hydrophobic silica. In one embodiment, the hydrophobic silica is dimethyl silica. In one embodiment, the hydrophobic silica is Aerosil R-972. In one embodiment, the dosage form or pharmaceutical composition has a relatively low loading of active ingredient, for example, from about 0.1 to about 0.2% w/w. of the total weight, or about 0.1 mg of active ingredient per capsule for about 75 mg.

5.3 Methods of use

In one embodiment, provided herein is a method of treating multiple myeloma, which includes administering to a patient a pharmaceutical composition provided herein. In one embodiment, provided herein is a pharmaceutical composition provided herein for use in a method of treating multiple myeloma, wherein the method includes administering said pharmaceutical composition to a patient.

In one embodiment, provided herein is a method of preventing multiple myeloma, which includes administering to a patient a pharmaceutical composition provided herein. In one embodiment, provided herein is a pharmaceutical composition provided herein for use in a method for preventing multiple myeloma, wherein said method includes said compound for a patient.

In one embodiment, provided herein is a method of treating multiple myeloma, which includes administering to a patient a pharmaceutical composition provided herein. In one embodiment, provided herein is a pharmaceutical composition provided herein for use in a method of treating multiple myeloma, wherein the method includes administering the compound to a patient.

In one embodiment, this document also provides methods for inducing a therapeutic response assessed using the International Uniform Response Criteria for Multiple Myeloma (IURC) (see Durie BGM, Harousseau J-L, Miguel JS, et al. International uniform response criteria for multiple myeloma. Leukemia, 2006; (10) 10: 1-7) to a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to a patient suffering from multiple myeloma. In another embodiment, provided herein are methods of achieving a strict complete response, a complete response, or a very good partial response, as defined by the International Unified Response Criteria for Multiple Myeloma (IURC) in a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to the patient, suffering from multiple myeloma. In another embodiment, this document provides methods

- 25 046729 achieving an increase in overall survival, progression-free survival, event-free survival, time to progression or disease-free survival in a patient, including the administration of an effective amount of a pharmaceutical composition presented herein. patient with multiple myeloma. In another embodiment, provided herein are methods for improving overall survival of a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to a patient suffering from multiple myeloma. In another embodiment, provided herein are methods of increasing progression-free survival in a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to a patient suffering from multiple myeloma. In another embodiment, provided herein are methods of achieving increased event-free survival in a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to a patient suffering from multiple myeloma. In another embodiment, provided herein are methods of achieving an increase in time to progression in a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to a patient suffering from multiple myeloma. In another embodiment, provided herein are methods of increasing disease-free survival of a patient, comprising administering an effective amount of a pharmaceutical composition provided herein to a patient suffering from multiple myeloma.

This document also provides treatments for patients who have previously been treated for multiple myeloma but do not respond to standard treatments, as well as for patients who have not previously been treated. It also covers treatments for patients who have undergone surgery to attempt treatment for multiple myeloma, as well as those who have not. This document also presents treatments for patients who have previously undergone transplant therapy, as well as those who have not.

The methods provided herein include the treatment of multiple myeloma that is relapsed, refractory or refractory. Methods provided herein include the prevention of multiple myeloma that is relapsed, refractory or resistant. The methods provided herein include the treatment of multiple myeloma that is relapsed, refractory or refractory. In some such embodiments, the myeloma is primary, secondary, tertiary, quadruple, or quintuple multiple myeloma. In one embodiment, the methods presented herein reduce, maintain, or eliminate minimal residual disease (MRD). In one embodiment, the methods provided herein include the treatment, prevention, or management of various types of multiple myeloma, such as monoclonal gammopathy of undetermined significance (MGUS), low-, intermediate-, and high-risk multiple myeloma, newly diagnosed multiple myeloma (including newly diagnosed multiple myeloma). low-intermediate and high-risk myeloma), transplant-eligible and transplant-ineligible multiple myeloma, smoldering (smoldering) multiple myeloma (including low-, intermediate- and high-risk smoldering multiple myeloma), active multiple myeloma, solitary plasmacytoma, extramedullary plasmacytoma, plasma cell leukemia , multiple myeloma of the central nervous system, light chain myeloma, non-secretory myeloma, immunoglobulin D myeloma and immunoglobulin E myeloma by administering a therapeutically effective amount of a pharmaceutical composition provided herein. In another embodiment, the methods provided herein include the treatment, prevention, or management of multiple myeloma characterized by genetic abnormalities such as cyclin D translocations (e.g., t(11;14)(q13;q32); t(6;14) (p21; 32); t (12; 14) (p13; q32); or t (6; 20);); MMSET translocations (eg t(4;14)(p16;q32)); MAF translocations (e.g. t(14;16)(q32;q32); t(20;22); t(16;22)(q11;q13); or t(14;20)(q32;q11)); or other chromosomal factors (eg, deletion of 17p13 or chromosome 13; del (17/17p), non-hyperdiploidy and amplification (1q)) by administering a therapeutically effective amount of a pharmaceutical composition provided herein.

In some embodiments, the methods include administering a therapeutically effective amount of a pharmaceutical composition provided herein as induction therapy. In some embodiments, the methods include administering a therapeutically effective amount of a pharmaceutical composition provided herein as consolidation therapy. In some embodiments, the methods include administering a therapeutically effective amount of a pharmaceutical composition provided herein as maintenance therapy.

In one specific embodiment of the methods described herein, multiple myeloma is a plasma cell leukemia.

In one embodiment of the methods described herein, multiple myeloma

- 26 046729 is a high-risk multiple myeloma. In some such embodiments, the high-risk multiple myeloma is relapsed or refractory. In one embodiment, high-risk multiple myeloma is multiple myeloma that recurs within 12 months of initial treatment. In yet another embodiment, high-risk multiple myeloma is multiple myeloma that is characterized by genetic abnormalities, such as one or more of del(17/17p) and t(14;16)(q32;q32). In some such embodiments, the high-risk multiple myeloma has relapsed or is resistant to one, two, or three previous courses of treatment.

In one embodiment, multiple myeloma is characterized by a p53 mutation. In one embodiment, the p53 mutation is a Q331 mutation. In one embodiment, the p53 mutation is the R273H mutation. In one embodiment, the p53 mutation is a K132 mutation. In one embodiment, the p53 mutation is a K132N mutation. In one embodiment, the p53 mutation is an R337 mutation. In one embodiment, the p53 mutation is the R337L mutation. In one embodiment, the p53 mutation is a W146 mutation. In one embodiment, the p53 mutation is an S261 mutation. In one embodiment, the p53 mutation is an S261T mutation. In one embodiment, the p53 mutation is an E286 mutation. In one embodiment, the p53 mutation is an E286K mutation. In one embodiment, the p53 mutation is an R175 mutation. In one embodiment, the p53 mutation is the R175H mutation. In one embodiment, the p53 mutation is an E258 mutation. In one embodiment, the p53 mutation is an E258K mutation. In one embodiment, the p53 mutation is an A161 mutation. In one embodiment, the p53 mutation is an A161T mutation.

In one embodiment, multiple myeloma is characterized by a homozygous deletion of p53. In one embodiment, multiple myeloma is characterized by a homozygous deletion of wild-type p53.

In one embodiment, multiple myeloma is characterized by wild-type p53.

In one embodiment, multiple myeloma is characterized by activation of one or more oncogenic factors. In one embodiment, the one or more oncogenic drivers are selected from the group consisting of C-MAF, MAFB, FGFR3, MMset, Cyclin D1, and Cyclin D. In one embodiment, multiple myeloma is characterized by activation of C-MAF. In one embodiment, multiple myeloma is characterized by activation of MAFB. In one embodiment, multiple myeloma is characterized by activation of FGFR3 and MMset. In one embodiment, multiple myeloma is characterized by activation of C-MAF, FGFR3, and MMset. In one embodiment, multiple myeloma is characterized by activation of cyclin D1. In one embodiment, multiple myeloma is characterized by activation of MAFB and cyclin D1. In one embodiment, multiple myeloma is characterized by activation of cyclin D.

In one embodiment, multiple myeloma is characterized by one or more chromosomal translocations. In one embodiment, the chromosomal translocation is t(14;16). In one embodiment, the chromosomal translocation is t(14;20). In one embodiment, the chromosomal translocation is t(4;14). In one embodiment, the chromosomal translocations are t(4;14) and t(14;16). In one embodiment, the chromosomal translocation is t(11;14). In one embodiment, the chromosomal translocation is t(6;20). In one embodiment, the chromosomal translocation is t(20;22). In one embodiment, the chromosomal translocations are t(6;20) and t(20;22). In one embodiment, the chromosomal translocation is t(16;22). In one embodiment, the chromosomal translocations are t(14;16) and t(16;22). In one embodiment, the chromosomal translocations are t(14;20) and t(11;14).

In one embodiment, multiple myeloma is characterized by a Q331 p53 mutation, C-MAF activation, and a chromosomal translocation at t(14;16). In one embodiment, multiple myeloma is characterized by a homozygous p53 deletion, C-MAF activation, and a chromosomal translocation at t(14;16); 16). In one embodiment, multiple myeloma is characterized by a p53 K132N mutation, MAFB activation, and a chromosomal translocation at t(14;20). In one embodiment, multiple myeloma is characterized by wild-type p53, FGFR3 and MMset activation, and a chromosomal translocation at t(4;14). In one embodiment, multiple myeloma is characterized by wild-type p53, C-MAF activation, and a chromosomal translocation at t(14;16). In one embodiment, multiple myeloma is characterized by homozygous deletion of p53, activation of FGFR3, MMset and C-MAF, and chromosomal translocations at t(4;14) and t(14;16). In one embodiment, multiple myeloma is characterized by a homozygous deletion of p53, activation of cyclin D1, and a chromosomal translocation at t(11;14). In one embodiment, multiple myeloma is characterized by

- 27 046729 p53 mutation R337L, activation of cyclin D1 and chromosomal translocation at t(11;14). In one embodiment, multiple myeloma is characterized by a p53 W146 mutation, activation of FGFR3 and MMset, and a chromosomal translocation at t(4;14). In one embodiment, multiple myeloma is characterized by the S261T p53 mutation, MAFB activation, and chromosomal translocations at t(6;20) and t(20;22). In one embodiment, multiple myeloma is characterized by a p53 E286K mutation, activation of FGFR3 and MMset, and a chromosomal translocation at t(4;14). In one embodiment, multiple myeloma is characterized by a p53 R175H mutation, activation of FGFR3 and MMset, and a chromosomal translocation at t(4;14). In one embodiment, multiple myeloma is characterized by a p53 E258K mutation, C-MAF activation, and chromosomal translocations at t(14;16) and t(16;22). In one embodiment, multiple myeloma is characterized by wild-type p53, activation of MAFB and cyclin D1, and chromosomal translocations at t(14;20) and t(11;14). In one embodiment, multiple myeloma is characterized by the A161T p53 mutation, cyclin D activation, and a chromosomal translocation at t(11;14).

In some embodiments of the methods described herein, the multiple myeloma is a newly diagnosed multiple myeloma suitable for transplantation. In another embodiment, the multiple myeloma is a newly diagnosed multiple myeloma ineligible for transplantation.

In yet other embodiments, multiple myeloma is characterized by early progression (eg, less than 12 months) after initial treatment. In other embodiments, multiple myeloma is characterized by early progression (eg, less than 12 months) after autologous stem cell transplantation. In another embodiment, the multiple myeloma is resistant to lenalidomide. In another embodiment, multiple myeloma is refractory to pomalidomide. In some such embodiments, the multiple myeloma is assumed to be resistant to pomalidomide (eg, based on molecular characteristics). In another embodiment, the multiple myeloma is recurrent or resistant to 3 or more treatments and has been exposed to a proteasome inhibitor (eg, bortezomib, carfilzomib, ixazomib, oprosomib, or marizomib) and an immunomodulatory compound (eg, thalidomide, lenalidomide, pomalidomide, iberdomide, or avadomide), or doubly resistant to a proteasome inhibitor and an immunomodulatory compound. In still other embodiments, the multiple myeloma is relapsed or refractory to 3 or more prior treatments, including, for example, a CD38 monoclonal antibody (CD38 mAb, e.g., daratumumab or isatuximab), a proteasome inhibitor (e.g., bortezomib, carfilzomib, ixazomib, or marizomib) and an immunomodulatory compound (eg, thalidomide, lenalidomide, pomalidomide, iberdomide, or avadomide) or dual resistance to a proteasome inhibitor or immunomodulatory compound and a CD38 mAb. In other embodiments, the multiple myeloma is triple refractory, e.g., the multiple myeloma is resistant to a proteasome inhibitor (e.g., bortezomib, carfilzomib, ixazomib, oprosomib, or marizomib), an immunomodulatory compound (e.g., thalidomide, lenalidomide, pomalidomide, iberdomide, or avadomide), and another active agent as described herein.

In certain embodiments, provided herein are methods of treating, preventing, and/or managing multiple myeloma, including relapsed/refractory multiple myeloma in patients with renal impairment or symptom thereof, comprising administering a therapeutically effective amount of a pharmaceutical composition provided herein to a patient with relapsed/refractory multiple myeloma with impaired renal function.

In certain embodiments, provided herein are methods of treating, preventing, and/or managing multiple myeloma, including relapsed or refractory multiple myeloma in a debilitated patient or symptoms thereof, comprising administering a therapeutically effective amount of a pharmaceutical composition provided herein to a debilitated patient with multiple myeloma . In some such embodiments, the frail patient is characterized by being unsuitable for induction therapy or intolerant to dexamethasone treatment. In some such embodiments, the frail patient is elderly, such as over 65 years of age.

In certain embodiments, provided herein are methods of treating, preventing, or managing multiple myeloma, comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition provided herein, wherein the multiple myeloma is fourth-line relapsed/refractory multiple myeloma.

In certain embodiments, provided herein are methods of treating, preventing, or managing multiple myeloma, comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition provided herein, in

- 28 046729 as induction therapy, where multiple myeloma is newly diagnosed multiple myeloma suitable for transplantation.

In certain embodiments, provided herein are methods of treating, preventing, or managing multiple myeloma, comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition provided herein as maintenance therapy following other therapy or transplantation where multiple myeloma is newly diagnosed, multiple myeloma , suitable for transplantation, before other therapy or transplantation.

In certain embodiments, provided herein are methods of treating, preventing, or managing multiple myeloma, comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition provided herein as maintenance therapy following other therapy or transplantation. In some embodiments, multiple myeloma is newly diagnosed multiple myeloma eligible for transplantation, prior to other therapy and/or transplantation. In some embodiments, the other pre-transplant therapy is treatment with chemotherapy or Compound 1.

In certain embodiments, provided herein are methods of treating, preventing, or managing multiple myeloma, comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition provided herein, wherein multiple myeloma is high-risk multiple myeloma that recurs or is refractory to one, two , or three previous treatments.

In certain embodiments, provided herein are methods of treating, preventing, or managing multiple myeloma, comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition provided herein, wherein the multiple myeloma is newly diagnosed multiple myeloma not suitable for transplantation.

In certain embodiments, the therapeutically or prophylactically effective amount of the compound is from about 0.01 to about 25 mg per day, from about 0.01 to about 10 mg per day, from about 0.01 to about 5 mg per day, from about 0 .01 to about 2 mg per day, about 0.01 to about 1 mg per day, about 0.01 to about 0.5 mg per day, about 0.01 to about 0.25 mg per day, from about 0.1 to about 25 mg per day, about 0.1 to about 10 mg per day, about 0.1 to about 5 mg per day, about 0.1 to about 2 mg per day, about 0 1 to about 1 mg per day, about 0.1 to about 0.5 mg per day, about 0.1 to about 0.25 mg per day, about 0.5 to about 25 mg per day, from about 0.5 to about 10 mg per day, about 0.5 to about 5 mg per day, about 0.5 to about 2 mg per day, about 0.5 to about 1 mg per day, about 1 up to about 25 mg per day, about 1 to about 10 mg per day, about 1 to about 5 mg per day, about 1 to about 2.5 mg per day, or about 1 to about 2 mg per day. In one embodiment, a therapeutically or prophylactically effective amount of Compound 1 is from about 0.1 mg per day to about 0.4 mg per day.

In some embodiments, the therapeutically or prophylactically effective amount is about 0.1, about 0.2, about 0.3, about 0.4, about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 15, about 20, or about 25 mg per day. In some such embodiments, the therapeutically or prophylactically effective amount is about 0.1, about 0.2, about 0.3, about 0.4, about 0.5, about 0.6, or about 0.7 mg per day.

In one embodiment, the recommended daily dosage range for Compound 1 for the conditions described herein is from about 0.1 mg to about 25 mg per day, preferably administered as a single dose once daily or in divided doses over throughout the day. In other embodiments, the dosage is from about 0.1 to about 10 mg per day. Specific daily doses include 0.1, 0.2, 0.3, 0.4, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 mg per day. More specific daily doses include 0.1, 0.2, 0.3, 0.4, or 0.5 mg per day.

In a particular embodiment, the recommended starting dose may be 0.1, 0.2, 0.3, 0.4, 0.5, 1, 2, 3, 4, 5, 10, 15, 20, or 25 mg per day. In another embodiment, the recommended starting dose may be 0.1, 0.2, 0.3, 0.4, or 0.5 mg per day. The dose can be increased to 1, 2, 3, 4 or 5 mg per day.

In some embodiments, the therapeutically or prophylactically effective amount is from about 0.001 to about 5 mg/kg/day, from about 0.001 to about 4 mg/kg/day, from about 0.001 to about 3 mg/kg/day, from about 0.001 to about 2 mg/kg/day, about 0.001 to about 1 mg/kg/day, about 0.001 to about 0.05 mg/kg/day, about 0.001 to about 0.04 mg/kg/day, about 0.001 to about 0.03 mg/kg/day, about 0.001 to about 0.02 mg/kg/day, about 0.001 to about 0.01 mg/kg/day, or about 0.001 to about 0.005 mg/kg/day day.

The dose administered may also be expressed in units other than mg/kg/day. For example, doses

- 29 046729 for parenteral administration can be expressed in mg/m ² /day. One skilled in the art will readily recognize how to convert doses from mg/kg/day to mg/m ² /day based on the subject's height or weight, or both (see www.fda.gov/cder/cancer/animalframe .htm). For example, a dose of 1 mg/kg/day for a 65 kg person is approximately 38 mg/ ^m2 /day.

In some embodiments, the patient to be treated with one of the methods presented herein has not been treated with multiple myeloma therapy prior to administration of the pharmaceutical composition provided herein. In some embodiments, a patient to be treated with one of the methods presented herein has been treated with multiple myeloma therapy prior to administration of the pharmaceutical composition provided herein. In some embodiments, a patient being treated with one of the methods presented herein has developed drug resistance to therapy against multiple myeloma. In some such embodiments, the patient has developed resistance to one, two, or three anti-multiple myeloma drugs, wherein the treatments are selected from a CD38 monoclonal antibody (CD38 mAb, e.g., daratumumab or isatuximab), a proteasome inhibitor (for example, bortezomib, carfilzomib, ixazomib or marizomib) and an immunomodulatory compound (eg thalidomide, lenalidomide, pomalidomide, iberdomide or avadomide).

The methods presented herein cover treatment of the patient regardless of the patient's age. In some embodiments, the subject is 18 years of age or older. In other embodiments, the subject is over 18, 25, 35, 40, 45, 50, 55, 60, 65, or 70 years of age. In other embodiments, the subject is under 65 years of age. In other embodiments, the subject is over 65 years of age. In one embodiment, the subject is an elderly multiple myeloma subject, such as a subject over 65 years of age. In one embodiment, the subject is an elderly multiple myeloma subject, such as a subject over 75 years of age.

Depending on the disease state being treated and the condition of the subject, the pharmaceutical composition provided herein may be administered orally, parenterally (e.g., intramuscular, intraperitoneal, intravenous, CIV, intracellular injection or infusion, subcutaneous injection or implantation), inhalation, nasal, vaginal, rectal, sublingual, or external (eg, transdermal or topical) route of administration. The pharmaceutical composition provided herein may be formulated alone or together in a suitable dosage unit with pharmaceutically acceptable excipients, carriers, adjuvants and carrier vehicles suitable for each route of administration.

In one embodiment, the pharmaceutical composition provided herein is administered orally. In another embodiment, the pharmaceutical composition provided herein is administered parenterally. In yet another embodiment, the pharmaceutical composition provided herein is administered intravenously.

The pharmaceutical composition provided herein may be delivered in a single dose, such as, for example, a single bolus injection or oral tablets or pills; or over time, such as, for example, continuous infusion over time or divided bolus doses over time. The compounds described herein may be re-administered as necessary, for example, until the patient exhibits stable disease or regression, or until the patient experiences disease progression or unacceptable toxicity. Stable disease or absence thereof is determined by methods known in the art, such as assessment of the patient's symptoms, physical examination, imaging of the tumor, which has been visualized by X-ray, computed tomography, PET or MRI, and other conventional assessment methods.

The pharmaceutical composition provided herein may be administered once daily (QD or qd) or divided into multiple daily doses such as twice daily (BID or bid), three times daily (TID or tid) and four times per day (QID or qid). In addition, administration can be continuous (eg daily for consecutive days or every day), intermittent, such as cyclically (eg including days, weeks or months of rest without drug). As used herein, the term daily is intended to mean that the therapeutic compound is administered one or more times each day, for example, over a period of time. The term continuous is intended to mean that the therapeutic compound is administered daily for a continuous period of at least 7 days to 52 weeks. The term intermittent or intermittent as used in this document is intended to mean stopping and starting at regular or irregular intervals. For example, intermittent administration of a pharmaceutical composition provided herein is administration for one to six days per week, administration in cycles (e.g., daily administration for two to eight consecutive weeks, then a rest period without administration for up to one week), or introduction every other day. The term cycle as used in this document is intended to

- 30 046729 indicating that the therapeutic compound is administered daily or continuously, but with a rest period. In some such embodiments, administration is once daily for two to six days, followed by a rest period without administration for five to seven days.

In some embodiments, the frequency of administration ranges from about a daily dose to about a monthly dose. In some embodiments, administration is once daily, twice daily, three times daily, four times daily, once every other day, twice weekly, once weekly, once every two weeks, once every three weeks or once every four weeks. In one embodiment, the pharmaceutical composition provided herein is administered once daily. In another embodiment, the pharmaceutical composition provided herein is administered twice daily. In yet another embodiment, the pharmaceutical composition provided herein is administered three times daily. In yet another embodiment, the pharmaceutical composition provided herein is administered four times daily.

In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 20 days followed by a rest period. In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 15 days followed by a rest period. In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 10 days followed by a rest period. In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 7 days followed by a rest period. In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 5 days followed by a rest period. In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 4 days followed by a rest period. In one embodiment, a therapeutically effective amount of a pharmaceutical composition provided herein is administered in a treatment cycle that includes an administration period of up to 3 days followed by a rest period.

In one embodiment, the treatment cycle includes an administration period of up to 14 days followed by a rest period. In one embodiment, the treatment cycle includes an administration period of up to 10 days followed by a rest period. In one embodiment, the treatment cycle includes an administration period of up to 7 days followed by a rest period. In one embodiment, the treatment cycle includes an administration period of up to 5 days followed by a rest period. In one embodiment, the treatment cycle includes an administration period of up to 4 days followed by a rest period. In one embodiment, the treatment cycle includes an administration period of up to 3 days followed by a rest period.

In one embodiment, the rest period ranges from about 2 days to about 11 days. In one embodiment, the rest period ranges from about 2 days to about 10 days. In one embodiment, the rest period is about 2 days. In one embodiment, the rest period is about 3 days. In one embodiment, the rest period is about 4 days. In one embodiment, the rest period is about 5 days. In one embodiment, the rest period is about 6 days. In another embodiment, the rest period is about 7 days. In another embodiment, the rest period is about 8 days. In another embodiment, the rest period is about 9 days. In another embodiment, the rest period is about 10 days. In another embodiment, the rest period is about 11 days.

In one embodiment, the treatment cycle includes an administration period of up to 15 days, followed by a rest period of about 2 days to about 10 days. In one embodiment, the treatment cycle includes an administration period of up to 10 days, followed by a rest period of about 2 days to about 10 days. In one embodiment, the treatment cycle includes an administration period of up to 7 days, followed by a rest period of about 2 days to about 10 days. In one embodiment, the treatment cycle includes an administration period of up to 5 days, followed by a rest period of about 2 days to about 10 days. In one embodiment, the treatment cycle includes an administration period of up to 3 days, followed by a rest period of about 10 days to about 15 days. In one embodiment, the treatment cycle includes an administration period of up to 3 days, followed by a rest period of about 3 days to about 15 days.

In one embodiment, the treatment cycle includes an administration period of up to 15 days followed by a rest period of 7 days. In one embodiment, the treatment cycle includes an administration period of up to 10 days followed by a rest period of 5 days. In one embodiment, the treatment cycle includes an administration period of up to 10 days followed by a rest period of 4 days. In one embodiment, the treatment cycle includes an administration period of up to 10 days followed by a rest period of 3 days. In one embodiment, the treatment cycle includes

- 31 046729 administration period up to 10 days followed by a rest period of 2 days. In one embodiment, the treatment cycle includes an administration period of up to 7 days followed by a rest period of 7 days. In one embodiment, the treatment cycle includes an administration period of up to 5 days followed by a rest period of 5 days. In one embodiment, the treatment cycle includes an administration period of up to 3 days followed by a rest period of 11 days. In another embodiment, the treatment cycle includes an administration period of up to 5 days followed by a rest period of 9 days. In another embodiment, the treatment cycle includes an administration period of up to 5 days followed by a rest period of 2 days. In another embodiment, the treatment cycle includes an administration period of up to 3 days followed by a rest period of 4 days.

In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 5 of a 28-day cycle. In another embodiment, a treatment cycle includes administering a pharmaceutical composition provided herein on days 1 to 10 of a 28-day cycle. In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 21 of a 28-day cycle. In another embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 5 of a 7-day cycle. In another embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 7 of a 7-day cycle. In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 10 and days 15 to 24 of a 28-day cycle (herein referred to as a 20/28 dosing cycle). In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 3 and days 15 to 18 of a 28-day cycle. In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 7 and days 15 to 21 of a 28-day cycle (herein referred to as a 14/28 dosing cycle). In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 5 and days 15 to 19 of a 28-day cycle (herein referred to as a 10/28 dosing cycle). In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 3 and days 15 to 17 of a 28-day cycle (herein referred to as a 6/28 dosing cycle).

In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 14 of a 21-day cycle. In another embodiment, a treatment cycle includes administering a pharmaceutical composition provided herein on days 1 to 4 and 8 to 11 of a 21-day cycle. In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 through 5 and 8 through 12 of a 21-day cycle. In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 through 5 and 11 through 15 of a 21-day cycle. In another embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 5, 8 to 12, and 15 to 19 of a 21-day cycle. In another embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 through 4, 8 through 11, and 15 through 18 of a 21-day cycle. In another embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 4, 8 to 10, and 15 to 17 of a 21-day cycle. In another embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 3 and 8 to 11 of a 21-day cycle. In one embodiment, a treatment cycle includes administering a therapeutically effective amount of a pharmaceutical composition provided herein on days 1 to 3 and 11 to 13 of a 21-day cycle.

Any treatment cycle described herein may be repeated for at least 2, 3, 4, 5, 6, 7, 8 or more cycles. In some cases, a treatment cycle as described herein comprises from 1 to about 24 cycles, from about 2 to about 16 cycles, or from about 2 to about 4 cycles. In some cases, a treatment cycle as described herein comprises from 1 to about 4 cycles. In some embodiments, cycles 1 through 4 are all 28-day cycles. In some embodiments, a therapeutically effective amount of a pharmaceutical composition provided herein is administered over 1 to 13 cycles of 28 days (eg, about 1 year). In some cases, cyclic therapy is not limited to the number of cycles, and therapy continues until the disease progresses. Cycles may in some cases

- 32 046729 include changes in the duration of the administration periods and/or rest periods described in this document.

In one embodiment, the treatment cycle includes administration of a pharmaceutical composition provided herein at a dosage of about 0.1 mg/day, 0.2 mg/day, 0.3 mg/day, 0.4 mg/day, 0.5 mg/day, 0.6 mg/day, 0.7 mg/day, 0.8 mg/day, 0.9 mg/day, 1.0 mg/day, 5.0 mg/day, or 10 mg/day , administered once daily. In one embodiment, the treatment cycle includes administration of a pharmaceutical composition provided herein at a dosage of about 0.1 mg/day, 0.2 mg/day, 0.3 mg/day, 0.4 mg/day, 0.5 mg/day, 0.6 mg/day, 0.7 mg/day, or 0.8 mg/day administered once daily. In some such embodiments, the treatment cycle includes administering a pharmaceutical composition provided herein once daily at a dosage of about 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, or 0.5 mg per day from 1 to 10 of the 28-day cycle. In some such embodiments, the treatment cycle includes administering a pharmaceutical composition provided herein once daily at a dosage of about 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, or 0.5 mg per day from 1 to 10 and from 15 to 24 of the 28-day cycle. In some such embodiments, a treatment cycle includes administering a pharmaceutical composition provided herein once daily at a dosage of about 0.1 mg on days 1 to 10 and 15 to 24 of a 28-day cycle. In other embodiments, the treatment cycle includes administering a pharmaceutical composition provided herein twice daily at a dosage of about 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, or 0.5 mg on days 1 -3 28-day cycle. In other embodiments, the treatment cycle includes administering a pharmaceutical composition provided herein twice daily at a dosage of about 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, or 0.5 mg on days 1 to 3 and 15 to 19 of the 28-day cycle. In other embodiments, the treatment cycle includes administering a pharmaceutical composition provided herein twice daily at a dosage of about 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, or 0.5 mg on days 1 to 3 and 15 to 17 of the 28-day cycle. In other embodiments, the treatment cycle includes administering a pharmaceutical composition provided herein twice daily at a dosage of about 0.2 mg on days 1 to 3 and 15 to 17 of a 28-day cycle. In one such embodiment, the pharmaceutical composition is administered on days 1-3 (morning and evening), day 14 (evening only), days 15 and 16 (morning and evening), and day 17 (morning only) of a 28-day cycle, for example, in cycle 1.

For clarity, it should be noted that, unless otherwise stated, the doses of Compound 1 mentioned herein refer to the amount of Compound 1 in its free base form. In the event that, for example, a pharmaceutically acceptable salt of Compound 1 is used, the amounts indicated above will need to be adapted accordingly.

5.4 Combination therapy with a second active agent

The pharmaceutical composition provided herein may also be combined with or used in combination with (e.g., before, during, or after) conventional therapy, including, but not limited to, surgery, biological therapy (including immunotherapy, e.g., with checkpoint inhibitors). points), radiation therapy, chemotherapy, stem cell transplantation, cell therapy, or other nondrug therapy currently used to treat, prevent, or manage multiple myeloma. The combined use of a compound presented herein and conventional therapy may provide a unique treatment regimen that is unexpectedly effective for some patients. Without being limited by theory, it is believed that the pharmaceutical composition provided herein may provide additive or synergistic effects when used concomitantly with conventional therapy.

As discussed elsewhere herein, included within the context of this document is a method of reducing, treating and/or preventing adverse or undesirable effects associated with conventional therapy, including, but not limited to, surgery, chemotherapy, radiation therapy, biological therapy and immunotherapy. The pharmaceutical composition provided herein and the other active ingredient may be administered to a patient before, during, or after the occurrence of a side effect associated with conventional therapy.

The pharmaceutical composition provided herein may also be combined or used in combination with other therapeutic agents useful for the treatment and/or prevention of multiple myeloma described herein.

In one embodiment, provided herein is a method of treating, preventing, or managing multiple myeloma, comprising administering to a patient a pharmaceutical composition provided herein in combination with one or more second active agents and, optionally, in combination with radiation therapy, blood transfusions, or surgery.

As used herein, the term combination includes the use of more than one therapy (eg, one or more prophylactic and/or therapeutic agents). However, use of the term in combination does not limit the order in which therapies (eg, prophylactic and/or therapeutic agents) are administered to a patient with a disease or disorder. First therapy (eg, prophylactic or therapeutic agent such as

- 33 046729 the pharmaceutical composition presented herein can be administered before (for example, 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours , 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks or 12 weeks before, simultaneously or after (for example, 5 min, 15 min, 30 min, 45 min , 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks or 12 weeks after) administration of a second therapy (eg, prophylactic or therapeutic agent) to the subject. This document also discusses triple therapy as well as quadruple therapy. In one embodiment, the second therapy is dexamethasone.

Administration of the pharmaceutical composition provided herein and one or more second active agents to a patient may occur simultaneously or sequentially by the same or different routes of administration. The suitability of the particular route of administration used for a particular active agent will depend on the active agent itself (eg, whether it can be administered orally without degradation before entering the bloodstream).

The route of administration of the pharmaceutical composition presented herein is independent of the route of administration of the second therapy. In one embodiment, the pharmaceutical composition provided herein is administered orally. In another embodiment, the pharmaceutical composition provided herein is administered intravenously. Thus, in accordance with these embodiments, the pharmaceutical composition provided herein is administered orally or intravenously, and the second therapy can be administered orally, parenterally, intraperitoneally, intravenously, intraarterially, transdermally, sublingually, intramuscularly, rectally, buccally, intranasally, liposomally , by inhalation, vaginally, intraocularly, by local delivery by catheter or stent, subcutaneously, intra-adiposally, intra-articularly, intrathecally or in a slow-release dosage form. In one embodiment, the pharmaceutical composition provided herein and the second therapy are administered by the same route of administration, orally or intravenously. In another embodiment, the pharmaceutical composition provided herein is administered by one route, for example, intravenously, while the second agent (an anti-multiple myeloma agent) is administered by another route, for example, orally.

In one embodiment, the second active agent is administered intravenously or subcutaneously and once or twice daily in an amount of from about 1 to about 1000 mg, from about 5 to about 500 mg, from about 10 to about 350 mg, or from about 50 mg to about 200 mg. The specific amount of the second active agent will depend on the specific agent used, the type of multiple myeloma being treated or controlled, the severity and stage of the disease, as well as the amount of the pharmaceutical composition provided herein and any optional additional active agents administered concurrently to the patient.

One or more second active ingredients or agents can be used in conjunction with the pharmaceutical composition provided herein in the methods and compositions provided herein. The second active agents may be large molecules (eg, proteins), small molecules (eg, synthetic inorganic, organometallic, or organic molecules), or cellular therapeutics (eg, CAR cells).

Examples of second active agents that may be used in the methods and compositions described herein include one or more of the following: melphalan, vincristine, cyclophosphamide, etoposide, doxorubicin, bendamustine, obinutuzmab, a proteasome inhibitor (e.g., bortezomib, carfilzomib, ixazomib, or oprosomib marizomib), histone deacetylase inhibitor (eg panobinostat, ACY241), BET inhibitor (eg GSK525762A, OTX015, BMS-986158, TEN-010, CPI-0610, INCB54329, BAY1238097, FT-1101, ABBV-075, BI 89499 9, GS-5829, GSK1210151A (I-BET-151), CPI-203, RVX-208, XD46, MS436, PFI-1, RVX2135, ZEN3365, XD14, ARV-771, MZ-1, PLX5117, 4-[2- (cyclopropylmethoxy)-5-(methanesulfonyl)phenyl]-2-methylisoquinolin-1(2H)-one, EP11313 and EP11336), BCL2 inhibitor (eg, venetoclax or navitoclax), MCL-1 inhibitor (eg, AZD5991, AMG176, MIK665 , S64315 or S63845), LSD-1 inhibitor (e.g. ORY-1001, ORY-2001, INCB-59872, IMG-7289, TAK-418, GSK-2879552, 4-[2-(4-aminopiperidin-1-yl )-5-(3-fluoro-4-methoxyphenyl)-1-methyl-6-oxo-1,6-dihydropyrimidin-4-yl]-2-fluorobenzonitrile or its salt, corticosteroid (eg prednisone), dexamethasone; an antibody (eg, a CS1 antibody such as elotuzumab; an anti-CD38 antibody such as daratumumab or isatuximab; or a BCMA antibody or an antibody conjugate such as GSK2857916 or BI 836909), a checkpoint inhibitor (as described herein), or cells CAR (as described in this document).

In one embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is dexamethasone.

In some embodiments, dexamethasone is administered at a dose of 4 mg on days 1 and 8 of a 21-day cycle. In some other embodiments, dexamethasone is administered at a dose of 4 mg on days 1, 4, 8, and 11 of a 21-day cycle. In some embodiments, dexamethasone is administered at a dose of 4 mg on days 1, 8, and 15 of a 28-day period.

- 34 046729 cycles. In some other embodiments, dexamethasone is administered at a dose of 4 mg on days 1, 4, 8, 11, 15, and 18 of a 28-day cycle. In some embodiments, dexamethasone is administered at a dose of 4 mg on days 1, 8, 15, and 22 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 4 mg on days 1, 10, 15, and 22 of cycle 1. In some embodiments, dexamethasone is administered at a dose of 4 mg on days 1, 3, 15, and 17 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 4 mg on days 1, 3, 14, and 17 of cycle 1.

In some other embodiments, dexamethasone is administered at a dose of 8 mg on days 1 and 8 of a 21-day cycle. In some other embodiments, dexamethasone is administered at a dose of 8 mg on days 1, 4, 8, and 11 of a 21-day cycle. In some embodiments, dexamethasone is administered at a dose of 8 mg on days 1, 8, and 15 of a 28-day cycle. In some other embodiments, dexamethasone is administered at a dose of 8 mg on days 1, 4, 8, 11, 15, and 18 of a 28-day cycle. In some embodiments, dexamethasone is administered at a dose of 8 mg on days 1, 8, 15, and 22 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 8 mg on days 1, 10, 15, and 22 of cycle 1. In some embodiments, dexamethasone is administered at a dose of 8 mg on days 1, 3, 15, and 17 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 8 mg on days 1, 3, 14, and 17 of cycle 1.

In some embodiments, dexamethasone is administered at a dose of 10 mg on days 1 and 8 of a 21-day cycle. In some other embodiments, dexamethasone is administered at a dose of 10 mg on days 1, 4, 8, and 11 of a 21-day cycle. In some embodiments, dexamethasone is administered at a dose of 10 mg on days 1, 8, and 15 of a 28-day cycle. In some other embodiments, dexamethasone is administered at a dose of 10 mg on days 1, 4, 8, 11, 15, and 18 of a 28-day cycle. In some embodiments, dexamethasone is administered at a dose of 10 mg on days 1, 8, 15, and 22 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 10 mg on days 1, 10, 15, and 22 of cycle 1. In some embodiments, dexamethasone is administered at a dose of 10 mg on days 1, 3, 15, and 17 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 10 mg on days 1, 3, 14, and 17 of cycle 1.

In some embodiments, dexamethasone is administered at a dose of 20 mg on days 1 and 8 of a 21-day cycle. In some other embodiments, dexamethasone is administered at a dose of 20 mg on days 1, 4, 8, and 11 of a 21-day cycle. In some embodiments, dexamethasone is administered at a dose of 20 mg on days 1, 8, and 15 of a 28-day cycle. In some other embodiments, dexamethasone is administered at a dose of 20 mg on days 1, 4, 8, 11, 15, and 18 of a 28-day cycle. In some embodiments, dexamethasone is administered at a dose of 20 mg on days 1, 8, 15, and 22 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 20 mg on days 1, 10, 15, and 22 of cycle 1. In some embodiments, dexamethasone is administered at a dose of 20 mg on days 1, 3, 15, and 17 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 20 mg on days 1, 3, 14, and 17 of cycle 1.

In some embodiments, dexamethasone is administered at a dose of 40 mg on days 1 and 8 of a 21-day cycle. In some other embodiments, dexamethasone is administered at a dose of 40 mg on days 1, 4, 8, and 11 of a 21-day cycle. In some embodiments, dexamethasone is administered at a dose of 40 mg on days 1, 8, and 15 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 40 mg on days 1, 10, 15, and 22 of cycle 1. In certain other embodiments, dexamethasone is administered at a dose of 40 mg on days 1, 4, 8, 11, 15, and 18. daily cycle. In other such embodiments, dexamethasone is administered at a dose of 40 mg on days 1, 8, 15, and 22 of a 28-day cycle. In other such embodiments, dexamethasone is administered at a dose of 40 mg on days 1, 3, 15, and 17 of a 28-day cycle. In one such embodiment, dexamethasone is administered at a dose of 40 mg on days 1, 3, 14, and 17 of cycle 1.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided in the methods and compositions described herein is bortezomib. In yet another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided in the methods and compositions described herein is daratumumab. In some such embodiments, the methods further comprise administering dexamethasone. In some embodiments, the methods include administering a pharmaceutical composition provided herein with a proteasome inhibitor as described herein, a CD38 inhibitor as described herein, and a corticosteroid as described herein.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is panobinostat. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is ACY241. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is vincristine. In some such embodiments, methods

- 35 046729 additionally include the administration of dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is cyclophosphamide. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is etoposide. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is doxorubicin. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is venetoclax. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is AMG176. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is MIK665. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is GSK525762A. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is OTX015. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is 4-[2-(cyclopropylmethoxy)-5-(methanesulfonyl)phenyl]-2-methylisochemical nolin-1(2H)-one. In some such embodiments, the methods further comprise administering dexamethasone.

In another embodiment, the second active agent used in conjunction with the pharmaceutical composition provided herein in the methods and compositions described herein is 4-[2-(4-aminopiperidin-1-yl)-5-(3- fluoro-4-methoxyphenyl)-1-methyl-6oxo-1,6-dihydropyrimidin-4-yl]-2-fluorobenzonitrile or a salt thereof (eg besilate salt). In some such embodiments, the methods further comprise administering dexamethasone.

In some embodiments, a pharmaceutical composition provided herein is administered in combination with checkpoint inhibitors. In one embodiment, one checkpoint inhibitor is used in combination with a pharmaceutical composition provided herein in connection with the methods presented herein. In another embodiment, two checkpoint inhibitors are used in combination with a pharmaceutical composition provided herein in connection with the methods presented herein. In yet another embodiment, three or more checkpoint inhibitors are used in combination with a pharmaceutical composition provided herein in connection with the methods presented herein.

As used herein, the term immune checkpoint inhibitor or checkpoint inhibitor refers to molecules that completely or partially reduce, inhibit, interfere with, or modulate one or more checkpoint proteins. Without being limited to any particular theory, checkpoint proteins regulate T cell activation or function. Numerous checkpoint proteins are known, such as CTLA-4 and its ligands CD80 and CD86; and PD-1 with its ligands PD-L1 and PD-L2 (Pardoll, Nature Reviews Cancer, 2012, 12, 252-264). These proteins appear to be responsible for costimulatory or inhibitory interactions of T cell responses. Immune checkpoint proteins appear to regulate and maintain self-tolerance and the duration and amplitude of physiological immune responses. Immune checkpoint inhibitors include antibodies or are derived from antibodies.

- 36 046729

In one embodiment, the checkpoint inhibitor is a CTLA4 inhibitor. In one embodiment, the CTLA-4 inhibitor is an anti-CTLA-4 antibody. Examples of anti-CTLA-4 antibodies include, but are not limited to, the antibodies described in US Pat. Nos. 5,811,097; 5811097; 5855887; 6051227; 6207157; 6682736; 6984720; and 7605238, all of which are incorporated herein in their entirety. In one embodiment, the anti-CTLA-4 antibody is tremelimumab (also known as ticilimumab or CP-675,206). In another embodiment, the anti-CTLA-4 antibody is ipilimumab (also known as MDX-010 or MDX-101). Ipilimumab is a fully human IgG monoclonal antibody that binds to CTLA-4. Ipilimumab is sold under the trade name Yervoy™.

In one embodiment, the checkpoint inhibitor is a PD1/PD-L1 inhibitor. Examples of PD-1/PD-L1 inhibitors include, but are not limited to, those described in US Pat. No. 7,488,802; 7,943,743; 8,008,449; 8,168,757; 8,217,149 and publications of PCT patent applications WO2003042402, WO2008156712, WO2010089411, WO2010036959, WO2011066342, WO2011159877, WO2011082400 and WO2011161699, all of which are fully included in this document.

In one embodiment, the checkpoint inhibitor is a PD-1 inhibitor. In one embodiment, the PD-1 inhibitor is an anti-PD-1 antibody. In one embodiment, the anti-PD-1 antibody is BGB-A317, nivolumab (also known as ONO-4538, BMS-936558, or MDX1106), or pembrolizumab (also known as MK-3475, SCH 900475, or lambrolizumab). In one embodiment, the anti-PD-1 antibody is nivolumab. Nivolumab is a human IgG4 monoclonal antibody against PD-1 and is marketed under the brand name Opdivo™. In another embodiment, the anti-PD-1 antibody is pembrolizumab. Pembrolizumab is a humanized IgG4 monoclonal antibody marketed under the brand name Keytruda™. In yet another embodiment, the anti-PD-1 antibody is a humanized antibody CT-011. CT-011, administered alone, showed no response in the treatment of relapsed acute myeloid leukemia (AML). In yet another embodiment, the anti-PD-1 antibody is an AMP-224 fusion protein. In another embodiment, the PD-1 antibody is BGB-A317. BGB-A317 is a monoclonal antibody that is specifically engineered to bind Fc gamma receptor I and has a unique PD-1 binding signature with high affinity and excellent target specificity.

In one embodiment, the checkpoint inhibitor is a PD-L1 inhibitor. In one embodiment, the PD-L1 inhibitor is an anti-PD-L1 antibody. In one embodiment, the anti-PD-L1 antibody is MEDI4736 (durvalumab). In another embodiment, the anti-PD-L1 antibody is BMS-936559 (also known as MDX-1105-01). In yet another embodiment, the PD-L1 inhibitor is atezolizumab (also known as MPDL3280A and Tecentriq®).

In one embodiment, the checkpoint inhibitor is a PD-L2 inhibitor. In one embodiment, the PD-L2 inhibitor is an anti-PD-L2 antibody. In one embodiment, the anti-PD-L2 antibody is rHIgM12B7A.

In one embodiment, the checkpoint inhibitor is a lymphocyte activation gene-3 (LAG-3) inhibitor. In one embodiment, the LAG-3 inhibitor is IMP321, a soluble Ig fusion protein (Brignone et al., J. Immunol., 2007, 179, 4202-4211). In another embodiment, the LAG-3 inhibitor is BMS-986016.

In one embodiment, the checkpoint inhibitors are a B7 inhibitor. In one embodiment, the B7 inhibitor is a B7-H3 inhibitor or a B7-H4 inhibitor. In one embodiment, the B7-H3 inhibitor is MGA271, an anti-B7-H3 antibody (Loo et al., Clin. Cancer Res., 2012, 3834).

In one embodiment, the checkpoint inhibitors are a TIM3 (T cell immunoglobulin domain and mucin domain 3) inhibitor (Fourcade et al., J. Exp. Med., 2010, 207, 2175-86; Sakuishi et al., J. Exp. Med., 2010, 207, 2187-94).

In one embodiment, the checkpoint inhibitor is an OX40 (CD134) agonist. In one embodiment, the checkpoint inhibitor is an anti-OX40 antibody. In one embodiment, the anti-OX40 antibody is anti-OX-40. In another embodiment, the anti-OX40 antibody is MEDI6469.

In one embodiment, the checkpoint inhibitor is a GITR agonist. In one embodiment, the checkpoint inhibitor is an anti-GITR antibody. In one embodiment, the anti-GITR antibody is TRX518.

In one embodiment, the checkpoint inhibitor is a CD137 agonist. In one embodiment, the checkpoint inhibitor is an anti-CD137 antibody. In one embodiment, the anti-CD137 antibody is urelumab. In another version

- 37 046729 implementation of the anti-CD137 antibody is PF-05082566.

In one embodiment, the checkpoint inhibitor is a CD40 agonist. In one embodiment, the checkpoint inhibitor is an anti-CD40 antibody. In one embodiment, the anti-CD40 antibody is CF-870,893.

In one embodiment, the checkpoint inhibitor is recombinant human interleukin-15 (rhIL-15).

In one embodiment, the checkpoint inhibitor is an IDO inhibitor. In one embodiment, the IDO inhibitor is INCB024360. In another embodiment, the IDO inhibitor is indoximod.

In some embodiments, the combination therapies provided herein include two or more checkpoint inhibitors described herein (including checkpoint inhibitors of the same or a different class). Moreover, the combination therapies described herein can be used in combination with one or more second active agents as described herein, where appropriate for the treatment of diseases described herein and understood in the art.

In some embodiments, a pharmaceutical composition provided herein may be used in combination with one or more immune cells expressing one or more chimeric antigen receptors (CARs) on their surface (eg, a modified immune cell). Typically, CARs include an extracellular domain of a first protein (eg, antigen binding protein), a transmembrane domain, and an intracellular signaling domain. In some embodiments, once the extracellular domain binds to a target protein, such as tumor-associated antigen (TAA) or tumor-specific antigen (TSA), a signal is generated through the intracellular signaling domain that activates the immune cell, for example, to target and kill the cell expressing the target protein.

Extracellular Domains: The extracellular domains of a CAR bind to the antigen of interest. In some embodiments, the extracellular domain of the CAR includes a receptor or portion of a receptor that binds to the antigen. In some embodiments, the extracellular domain includes or is an antibody or an antigen-binding portion thereof. In particular embodiments, the extracellular domain comprises or is a single chain Fv (scFv) domain. A single chain Fv domain may include, for example, a _VL linked to a VH by a flexible linker, wherein said VL and VH are derived from an antibody that binds the antigen.

In some embodiments, the antigen recognized by the extracellular domain of a polypeptide described herein is a tumor-associated antigen (TAA) or a tumor-specific antigen (TSA). In various specific embodiments, the tumor associated antigen or tumor specific antigen is, without limitation, Her2, prostate stem cell antigen (PSCA), alpha fetoprotein (AFP), carcinoembryonic antigen (CEA), cancer antigen-125 (CA-125), CA19 -9, calretinin, MUC-1, B cell maturation antigen (BCMA), epithelial membrane protein (EMA), epithelial tumor antigen (ETA), tyrosinase, melanoma-associated antigen-24 (MAGE), CD19, CD22, CD27, Cd30 , CD34, CD45, CD70, CD99, CD117, EGFRvIII (epidermal growth factor III variant), mesothelin, PAP (prostate acid phosphatase), prostein, TARP (T-cell receptor gamma alternative reading frame protein), Trpp8, STEAPI (six transmembrane epithelial prostate antigen 1), chromogranin, cytokeratin, desmin, glial fibrillary acidic protein (GFAP), liquid macrogenic cystic disease protein (GCDFP-15), HMB-45 antigen, melan-A protein (melanoma antigen is recognized by T lymphocytes; MART-I), myo-DI, muscle-specific actin (MSA), neurofilament, neuron-specific enolase (N SE), placental alkaline phosphatase, synaptophysis, thyroglobulin, thyroid transcription factor-1, dimeric form of pyruvate kinase type M2 isoenzyme (M2 tumor -PK), abnormal ras protein or abnormal p53 protein. In some other embodiments, the TAA or TSA recognized by the extracellular domain of the CAR is ave3 integrin (CD61), galactin, or Ral-B.

In some embodiments, the TAA or TSA recognized by the extracellular domain of the CAR is a cancer/testis (CT) antigen, e.g., BAGE, CAGE, CTAGE, FATE, GAGE, HCA661, HOM-TES-85, MAGEA, MAGEB, MAGEC, NA88 , NY-ES0-1, NY-SAR-35, OY-TES-1, SPANXBI, SPA17, SSX, SYCPI or TPTE.

In some other embodiments, the TAA or TSA recognized by the extracellular domain of the CAR is a carbohydrate or ganglioside, for example, fuc-GMI, GM2 (oncofetal antigen-immunogenic-1; OFA-I-1); GD2 (OFA-I-2), GM3, GD3, etc.

In some other embodiments, the TAA or TSA recognized by the extracellular domain of the CAR is alpha-actinin-4, Bage-1, BCR-ABL, Bcr-Abl fusion protein, beta-catenin, CA 125, CA 15-3 (CA 27.29 \BCAA), CA 195, CA 242, CA-50, CAM43, Casp-8, cdc27, cdk4, cdkn2a, CEA, coa-l, dek-can fusion protein, EBNA, EF2, Epstein-Barr virus antigens, ETV6- AML1 fusion protein, HLA-38 046729

A2, HLA-All, hsp70-2, KIAA0205, Mart2, Mum-1, 2 and 3, neo-PAP, class I myosin, OS-9, pml-RARa fusion protein, PTPRK, K-ras, N-ras, triosephosphate isomerase, Gage 3,4,5,6,7, GnTV, Herv-K-mel, Lage-1, NA88, NY-Eso-1/Lage-2, SP17, SSX-2, TRP2-Int2, gp100 (Pmel17), tyrosinase, TRP-1, TRP-2, MAGE-1, MAGE-3, RAGE, GAGE-1, GAGE-2, p15 (58), RAGE, SCP-1, Hom/Mel-40, PRAME , p53, HRas, HER-2/neu, E2A-PRL, H4-RET, IGH-IGK, MYL-RAR, human papillomavirus (HPV) antigens E6 and E7, TSP-180, MAGE-4, MAGE-5, MAGE-6, p185erbB2, p180erbB-3, c-met, nm-23H1, PSA, TAG-72-4, CA 19-9, CA 72-4, CAM 17.1, NuMa, K-ras, 13-Catenin, Mum -1, p16, TAGE, P SMA, CT7, telomerase, 43-9F, 5T4, 791Tgp72, 13HCG, BCA225, BTAA, CD68\KP1, C0-029, FGF-5, G250, Ga733 (EpCAM), HTgp-175 , M344, MA-50, MG7-Ag, MOV18, NB\70K, NY-C0-1, RCAS1, SDCCAG16, TA-90, TAAL6, TAG72, TLP or TPS.

In various specific embodiments, the tumor antigen or tumor-specific antigen are tumor antigens associated with AML, as described in S. Anguille et al, Leukemia (2012), 26, 2186-2196.

Other tumor-associated and tumor-specific antigens are known to those skilled in the art.

Receptors, antibodies and scFvs that bind to TSA and TAA useful for constructing chimeric antigen receptors are known in the art, as are the nucleotide sequences that encode them.

In some specific embodiments, the antigen recognized by the extracellular domain of the chimeric antigen receptor is an antigen that is not typically considered a TSA or TAA, but that is nonetheless associated with tumor cells or tumor-induced damage. In some embodiments, for example, the antigen is, for example, a growth factor, cytokine, or interleukin, such as a growth factor, cytokine, or interleukin associated with angiogenesis or vasculogenesis. Such growth factors, cytokines or interleukins may include, for example, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF), hepatocyte growth factor (HGF), insulin-like growth factor. (IGF) or interleukin-8 (IL-8). Tumors can also create a local hypoxic environment relative to the tumor. Thus, in other specific embodiments, the antigen is a hypoxia-associated factor, such as HIF-1a, HIF-1e, HIF-2a, HIF-2e, HIF-3a, or HIF-3e. Tumors can also cause local damage to normal tissue by causing the release of molecules known as damage-associated molecular pattern molecules (DAMPs; also known as alarmins - damage-associated molecular moieties). Therefore, in some other specific embodiments, the antigen is a DAMP, e.g., heat shock protein, chromatin-associated protein high mobility group block 1 (HMGB 1), S100A8 (MRP8, calgranulin A), S100A9 (MRP14, calgranulin B), serum amyloid A (SAA), or may be deoxyribonucleic acid, adenosine triphosphate, uric acid or heparin sulfate.

Transmembrane Domain: In some embodiments, the extracellular domain of the CAR is linked to the transmembrane domain of the polypeptide via a linker, spacer, or hinge polypeptide sequence, such as a CD28 sequence or a CTLA4 sequence. The transmembrane domain may be derived from or derived from the transmembrane domain of any transmembrane protein and may include all or part of such transmembrane domain. In specific embodiments, the transmembrane domain may be derived from, for example, CD8, CD16, cytokine receptor and interleukin receptor, or growth factor receptor or the like.

Intracellular Signaling Domains: In some embodiments, a CAR intracellular domain is or includes an intracellular domain or protein motif that is expressed on the surface of T cells and triggers activation and/or proliferation of said T cells. Such a domain or motif is capable of transmitting the primary antigen-binding signal, which is necessary for T-lymphocyte activation in response to antigen binding to the extracellular part of the CAR. Typically this domain or motif includes or is an ITAM (immunoreceptor tyrosine-based activation motif). ITAM-containing polypeptides suitable for CAR include, for example, CD3 zeta chain (CD3Z) or ITAM-containing portions thereof. In a specific embodiment, the intracellular domain is a CD3Z intracellular signaling domain. In other specific embodiments, the intracellular domain is derived from a lymphocyte receptor chain, a TCR/CD3 complex protein, an Fe receptor subunit, or an IL2 receptor subunit. In some embodiments, the CAR further comprises one or more co-stimulatory domains or motifs, for example, as part of an intracellular domain of a polypeptide. One or more costimulatory domains or motifs may be or may contain one or more CD27 costimulatory polypeptide sequences, a CD28 costimulatory polypeptide sequence, an OX40 costimulatory polypeptide sequence (CD134), a co-stimulatory 4-1BB polypeptide sequence (CD137), or

- 39 046729 costimulatory inducible T cell costimulatory sequence (ICOS) polypeptide sequence, or other costimulatory domain or motif, or any combination thereof.

CAR may also contain a T cell survival motif. A T cell survival motif can be any polypeptide sequence or motif that promotes the survival of a T lymphocyte following antigen stimulation. In some embodiments, the T cell survival motif is or is derived from CD3, CD28, IL7 receptor (IL-7R) intracellular signaling domain, IL-12 receptor intracellular signaling domain, IL-15 receptor intracellular signaling domain, IL-15 receptor intracellular signaling domain -21 or the intracellular signaling domain of the transforming growth factor β receptor (TGFe).

Modified immune cells expressing CARs may be, for example, T lymphocytes (T cells, e.g. CD4+ T cells or CD8+ T cells), cytotoxic lymphocytes (CTL) or natural killer (NK) cells. T lymphocytes used in the compositions and methods presented herein can be naïve T lymphocytes or MHC-restricted T lymphocytes. In some embodiments, the T lymphocytes are tumor infiltrating lymphocytes (TILs). In some embodiments, T lymphocytes were isolated from a tumor biopsy or were expanded from T lymphocytes isolated from a tumor biopsy. In some other embodiments, T cells have been isolated or expanded from T lymphocytes isolated from peripheral blood, umbilical cord blood, or lymph. Immune cells to be used to generate modified CAR-expressing immune cells can be isolated using conventional techniques known in the art, such as blood collection followed by apheresis and possibly antibody-mediated cell isolation or sorting.

The modified immune cells are preferably autologous to the individual to whom the modified immune cells are to be administered. In some other embodiments, the modified immune cells are allogeneic to the individual to whom the modified immune cells are to be administered. If allogeneic T lymphocytes or NK cells are used to produce engineered T lymphocytes, it is preferable to select T lymphocytes or NK cells that will reduce the likelihood of graft-versus-host disease (GVHD) in the individual. For example, in some embodiments, virus-specific T lymphocytes are selected to produce modified T lymphocytes; such lymphocytes are expected to have a significantly reduced natural ability to bind to and thus be activated by any recipient antigens. In some embodiments, recipient-mediated rejection of allogeneic T lymphocytes can be reduced by coadministration of one or more immunosuppressive drugs, such as cyclosporine, tacrolimus, sirolimus, cyclophosphamide, or the like, to the host.

T lymphocytes, for example, unmodified T lymphocytes or T lymphocytes expressing CD3 and CD28, or containing a polypeptide containing a CD3Z signaling domain and a CD28 co-stimulatory domain, can be expanded using antibodies to CD3 and CD28, for example, antibodies attached to balls; see, for example, US patent No. 5948893; 6534055; 6352694; 6692964; 6887466; and 6905681.

Modified immune cells, such as modified T lymphocytes, may optionally contain a suicide gene or safety switch that allows virtually all modified immune cells to be killed if desired. For example, the modified T lymphocytes, in some embodiments, may contain the HSV thymidine kinase (HSV-TK) gene, which causes the modified T lymphocytes to die when exposed to ganciclovir. In another embodiment, the modified T lymphocytes contain an inducible caspase, for example, inducible caspase 9 (icaspase9), for example, a fusion protein between caspase 9 and the human binding protein FK506, allowing dimerization using a specific small molecule pharmaceutical. See Straathof et al., Blood 1 05 (11): 4247-4254 (2005).

In some embodiments, a pharmaceutical composition provided herein is administered to patients with various types or stages of multiple myeloma in combination with chimeric antigen receptor (CAR) T cells. In some embodiments, the CAR T cell in combination targets B-cell maturation antigen (BCMA), and in more specific embodiments, the CAR T cell is bb2121 or bb21217. In some embodiments, the CAR T cell is JCARH125.

6. Examples

Some embodiments of the invention are illustrated by the following non-limiting examples.

6.1 Synthesis of ^)-4-(4-(4-(((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-4-yl)oxy)methyl)benzyl)

- 40 046729 piperazin-1-yl)-3-fluorobenzonitrile (compound 1)

tert-Butyl (4S)-5-amino-4-(benzyloxycarbonylamino)-5-oxopentanoate. Di-tert-butyl dicarbonate (155 g, 711 mmol), pyridine (70.3 g, 889 mmol) and ammonium bicarbonate (105 g, 1.33 mol). The reaction mixture was stirred at 18°C for 16 hours and then evaporated. The residue was dissolved in ethyl acetate (5.0 L) and water (5.0 L), the organic layer was separated and washed with HCl (3.0 ml, 1 N), saturated sodium bicarbonate (3.0 L), brine (3 0 L), dried over anhydrous sodium sulfate, filtered and evaporated to give crude tert-butyl (4S)-5-amino-4-(benzyloxycarbonylamino)-5-oxopentanoate (450 g, crude) as a white solid, which was used in next stage without further purification. ¹ H NMR 400 MHz DMSO^ ₆ δ: 7.35-7.30 (m, 5H), 7.02 (s, 1H), 5.01 (d, J=3.2 Hz, 1H), 3, 93-3.90 (m, 1H), 2.20 (t, J=8.0 Hz, 2H), 1.88-1.84 (m, 1H), 1.72-1.69 (m, 1H), 1.35 (s, 9H).

tert-Butyl (4S)-4,5-diαmino-5-oxo-pentanoate. To a solution of tert-butyl (4S)-5-amino-4(benzyloxycarbonylamino)-5-oxopentanoate (112 g, 333 mmol) in methanol (1.0 L) was added 10% palladium on carbon (15 g) under a nitrogen atmosphere. The suspension was degassed under vacuum and purged with hydrogen several times. The mixture was stirred under an atmosphere of hydrogen gas (40 psi) at 30°C for 16 hours. The reaction mixture was filtered and the filtrate was evaporated to give crude tert-butyl (4S)-4,5-diamino-5-oxopentanoate as colorless oil. 1H NMR 400 MHz DMSO-de δ: 7.30 (s, 1H), 6.95 (s, 1H), 3.10-3.07 (m, 1H), 2.27-2.23 (m, 2H), 1.69-1.78 (m, 1H), 1.59-1.55 (m, 1H), 1.38 (s, 9H).

Methyl 3-hydroxy-2-methylbenzoate. Four batches (200 g each) were processed in parallel. To a solution of 3-hydroxy-2-methylbenzoic acid (200 g, 1.31 mol) in methanol (4.0 L) was added concentrated sulfuric acid (47.7 g, 486 mmol). The reaction mixture was stirred at 60°C for 17 hours. The reaction mixture was evaporated to 800 ml. The resulting mixture was cooled to 20°C and slowly poured into water (400 ml) over 30 min. Water (1200 ml) was added at 20°C for 3 hours and the resulting mixture was stirred at 20°C for 1 hour. The precipitated solid was collected by vacuum filtration (four batches combined) and washed three times with water/methanol (1000 ml, 9: 1) or until the filtrate reached pH > 3. The solid was dried in vacuo at 45°C to obtain methyl 3-hydroxy-2-methyl benzoate (700 g, 80.4% yield) as a gray solid . 1H NMR: 400 MHz DMSO-de δ: 9.70 (s, 1H), 7.18 (t, J=6.8 Hz, 1H), 7.09 (t, J=7.6 Hz, 1H) , 7.00 (t, J=6.8 Hz, 1H), 3.81 (s, 3H), 2.29 (s, 3H).

Methyl 3-[tert-butyl(dimethyl)silyl]oxy-2-methylbenzoate. Two batches (240 g each) were processed in parallel. To a solution of methyl 3-hydroxy-2-methylbenzoate (240 g, 1.44 mol) in N,Ndimethylformamide (1.40 L) was added imidazole (246 g, 3.61 mol) and tert-butyldimethylsilyl chloride (238 g, 1 .58 mol) at 5°C. After addition, the mixture was heated to 20°C and stirred for 6 hours. Isopropyl acetate (1700 ml) was added and then water (2000 ml) was added slowly, maintaining the temperature below 30°C. The resulting mixture was stirred, followed by separation of the organic phase. The combined organic layers (two batches combined) were washed with water (1700 ml x 3) and evaporated to ~1500 ml (KF <0.05%). The product was stored as an isopropyl acetate solution, which was used in the next step without further purification.

Methyl 2-(bromomethyl)-3-[tert-butyl(dimethyl)silyl]oxybenzoate. Two batches (~375 g each) were processed in parallel. To an isopropyl acetate solution of methyl 3-[tert-butyl(dimethyl)silyl]hydroxy2-methylbenzoate (~375 g, 1.34 mol) was added N-bromosuccinimide (274 g, 1.54 mol) and azobisisobutyronitrile (4.40 g, 26 .8 mmol). The reaction mixture was heated to 70°C for at least 1 hour and stirred at 70°C for 4 hours. The reaction mixture was cooled to 20°C and kept at 20°C for at least 1 hour. Two batches of the solid (succinimide) were removed filtration and washed with isopropyl acetate (700 ml). The filtrate was washed with a solution of sodium sulfite (700 g) in water (6000 ml) and then with water (1500 ml). The organic layer was distilled in vacuo at 45°C to dryness to obtain methyl 2(bromomethyl)-3-[tert-butyl(dimethyl)silyl]oxybenzoate (920 g, 95.5% yield) as a dark orange oil. 1H NMR: 400 MHz DMSO-de δ: 7.45 (d, J=6.8 Hz, 1H), 7.36 (t, J=8.0 Hz, 1H), 7.13 (t, J= 7.2 Hz, 1H), 4.95 (s, 2H), 1.02 (s, 9H), 0.29 (s, 6H).

tert-Butyl (4S)-5-αmino-4-[4-[tert-butyl(dimethyl)silyl]oxy-1-oxoisoindolin-2-yl]-5-oxopentanoate. To a solution of tert-butyl (4S)-4,5-diamino-5-oxopentanoate (130 g, 643 mmol) in acetonitrile (4.0

- 41 046729

l) methyl 2-(bromomethyl) 3-[tert-butyl(dimethyl)silyl]oxybenzoate (210 g, 584 mmol) and diisopropylethylamine (113 g, 877 mmol) were added. The reaction mixture was stirred at 50°C for 16 hours. The reaction mixture was evaporated to remove most of the acetonitrile, the residue was dissolved in methyl tert-butyl ether (2.0 L) and water (1.5 L), the organic layer was washed with saturated potassium monophosphate ( 1.0 l x 2), saline solution. (1.0 L), dried over anhydrous sodium sulfate, filtered and evaporated to give crude tert-butyl (4S)-5-amino-4-[4-[tert-butyl(dimethyl)silyl]oxy-1oxoisoindoline-2- yl]-5-oxopentanoate (524 g), which was used in the next step without further purification.

tert-Butyl (4S)-5-amino-4-(4-hydroxy-1-oxoisoindolin-2-yl)-5-oxopentanoate. To a solution of tert-butyl (4S)-5-amino-4-[4-[tert-butyl(dimethyl)silyl]oxy-1-oxoisoindolin-2-yl]-5-oxopentanoate (275 g, 613 mmol) in methanol (2 .0 L) tetrabutylammonium fluoride trihydrate (38.7 g, 123 mmol) was added. The mixture was stirred at 18°C for 16 hours. The reaction mixture was evaporated to remove most of the methanol, and the residue was dissolved in dichloromethane/water (3 L/2 L). The organic layer was separated and washed with brine (1.0 L), dried over anhydrous sodium sulfate, filtered and evaporated to give a crude product, which was purified by a silica gel column to give the product (260 g). The product was added to acetonitrile (750 ml) and the mixture was stirred at 60°C for 2 hours, cooled to 18°C and stirred for a further 2 hours. The solid was filtered and the precipitate was dried to give tert-butyl (4S)-5-amino -4-(4-hydroxy-1-oxoisoindolin-2-yl)-5-oxopentanoate (248 g, 60.5% yield) as a gray solid. 1H NMR 400 MHz DMSO<δ: 10.00 (s, 1H), 7.54 (s, 1H), 7.29 (t, J=7.6 Hz, 1H), 7.14 (d, J= 4.8 Hz, 2H), 4.72-4.68 (m, 1H), 4.49-4.28 (m, 2H), 2.17-1.97 (m, 4H), 1.31 (s, 9H).

4-(4-(4-(chloromethyl)benzyl)piperazin-1-yl)3-fluorobenzonitrile. 1,4-bis(chloromethyl)benzene (51.2 g, 292 mmol) was placed in a flask with acetonitrile (195 ml) and NN-dimethylformamide (195 ml). The reaction mixture was stirred at ambient temperature until all solids dissolved. Diisopropylamine (51.1 mL, 292 mmol) was then added along with 3-fluoro-4-(piperazin-1yl)benzonitrile (20 g, 97 mmol). The reaction mixture was heated to 60°C for 1 hour. Acetonitrile was removed under reduced pressure. The remaining mixture was partitioned between ethyl acetate (1.0 L), water (700 ml) and saline (300 ml). The organic layer was separated, and the aqueous layer was extracted twice with ethyl acetate. Volatile organic compounds were combined and removed under reduced pressure. The solid was dissolved in a minimal amount of dichloromethane and purified on a silica gel column (0-100% ethyl acetate in hexanes, >3 L). Fractions containing the desired product were combined and volatile organic compounds were removed under reduced pressure. The residue was dissolved in a minimal amount of dichloromethane and purified a second time on a silica gel column (10% isocratic ethyl acetate in hexanes greater than 800 ml, then 20-80% ethyl acetate in hexanes greater than 4 L). Fractions containing the desired product were combined and VOCs were removed under reduced pressure to give 4-(4-(4-(chloromethyl)benzyl)piperazin-1-yl)-3-fluorobenzonitrile (22.7 g, 66.0 mmol , yield 67.7%) as an off-white solid. 1H NMR (400 MHz, CDCl ₃ ) δ ppm 7.33-7.39 (m, 5H) 7.29 (d, J=1.96 Hz, 1H) 7.25 (d, J=1.96 Hz, 1H) 6.91 (t, J= 8.56 Hz, 1H) 4.60 (s, 2H) 3.58 (s, 2H) 3.19-3.27 (m, 4H) 2.58-2.66 (m, 4H). MS(ESI) m/z 344.2 [M+1]+.

(R)-tert-butyl 5-amino-4-(4-((4-((4-(4-cyano-2-fluorophenyl)piperazin-1-yl)methyl)benzyl)oxy)-1oxoisoindoline-2- yl)-5-oxopentanoate. (R)-tert-butyl 5-amino-4-(4-hydroxy-1-oxoisoindolin-2-yl)5-oxopentanoate (22.05 g, 65.9 mmol) was placed in a flask with 4-(4-( 4-(chloromethyl)benzyl)piperazin-1-yl)-3fluorobenzonitrile (22.67 g, 65.9 mmol), potassium carbonate (18.23 g, 132 mmol) and N,Ndimethylformamide (330 ml). The reaction mixture was heated to 45°C for 16 hours. The reaction mixture was diluted with ethyl acetate (50 ml) and filtered. The filtrate was partitioned between ethyl acetate (900 ml), water (600 ml) and saline (200 ml). The organic layer was separated and washed with water (600 ml). The organic layer was dried over sodium sulfate and volatiles were removed under reduced pressure. The residue was treated with 20% ethyl acetate in hexanes and the volatiles were removed under reduced pressure to give (R)-tert-butyl 5-amino-4-(4-((4-((4-(4-cyano-2-fluorophenyl)piperazine -1-yl)methyl)benzyl)oxy)-1-oxoisoindolin-2-yl)-5-oxopentanoate (44.02 g, 68.6 mmol, 104% yield) as an off-white solid. The yield was slightly higher than quantitative, since some DMF remained. 1H NMR (400 MHz, CDCl ₃ ) δ ppm 7.43-7.49 (m, 2H) 7.40 (s, 4H) 7.36 (dd, J=8.38, 1.28 Hz, 1H) 7.29 (d, J=1.96 Hz, 1H) 7.26 (d, J=1.83 Hz, 1H) 7.11 (dd, J=7.64, 1.16 Hz, 1H) 6.92 (t, J=8.50 Hz , 1H) 6.23 (lat. s, 1H) 5.24-5.32 (m, 1H) 5.15 (s, 2H) 4.86- 4.94 (m, 1H) 4.38-4 .55 (m, 2H) 3.61 (s, 2H) 3.18-3.32 (m, 4H) 2.58-2.70 (m, 4H) 2.09-2.47 (m, 4H ) 1.43 (s, 8H). MS(ESI) m/z 642.4 [M+1]+.

^)-4-(4-(4-(((2-(2,6-Dioxopiperidin-3-yl)-1-oxoisoindolin-4-yl)oxy)methyl)benzyl)piperazin1-yl)-3-fluorobenzonitrile (connection 1). (I)-tert-butyl 5-amino-4-(4-((4-((4-(4-cyano-2-fluorophenyl)piperazin-1-yl)methyl)benzyl)oxy)-1-oxoisoindolin- 2-yl)-5-oxopentanoate (12.1 g, 18.86 mmol) was placed in a vial with acetonitrile (189 ml) and benzenesulfonic acid (3.96 g, 24.51 mmol). The reaction mixture was placed under vacuum and purged with nitrogen. This was repeated once more and then the mixture was heated to 85°C overnight under a nitrogen atmosphere. The warm reaction mixture was poured

- 42 046729 directly into 2 separatory funnels containing dichloromethane (1000 ml) and ethyl acetate (300 ml). To this mixture were added saturated sodium bicarbonate solution (900 ml), water (100 ml) and saline solution (450 ml). The organic layer was separated and the aqueous layer was extracted with dichloromethane (800 ml) and ethyl acetate (200 ml). The combined organic layers were dried over anhydrous magnesium sulfate and evaporated. Purification by standard methods yielded the title compound. 1H NMR (400 MHz, DMSO-de) δ ppm 10.96 (s, 1H) 7.68 (dd, J=13.45, 1.83 Hz, 1H) 7.56 (dd, J=8.44, 1.83 Hz, 1H) 7.43- 7.52 (m, 3H) 7.29-7.39 (m, 4H) 7.11 (t, J=8.80 Hz, 1H) 5.24 (s, 2H) 5.11 (dd, J=13.20, 5.14 Hz, 1H) 4.22-4.46 (m, 2H) 3.54 (s, 2H) 3.12-3.22 (m, 4H) 2.85-2 .97 (m, 1H) 2.53-2.62 (m, 2H) 2.38-2.48 (m, 2H) 1.93-2.03 (m, 1H). MC (IER) m/z 568.2 [M+1]+.

Recrystallization to give form K of compound 1. ^)-4-(4-(4-(((2-(2,6-dioxopiperidin3-yl)-1-oxoisoindolin-4-yl)oxy)methyl)benzyl)piperazin- 1-yl)-3-fluorobenzonitrile was dissolved in acetone (35 L/kg) and filtered to increase chiral purity. The filtrate was passed through a 0.45 μM filter into a clean reactor. After distillation (target volume 15 L/kg), water (2 L/kg) was added to create supersaturation. The batch was inoculated with form K and kept at constant temperature with wet grinding through a recirculation loop at 40°C. Water (6 L/kg) was charged at a constant rate of 1 (L/kg)/h while wet grinding at reduced tip speed. The batch was cooled to 25°C and then held for 3 hours. The batch was filtered, washed and dried. Wet aging followed by grinding was performed to obtain (S)-4-(4-(4-(((2-(2,6dioxopiperidin-3-yl)-1-oxoisoindolin-4-yl)oxy)methyl)benzyl)piperazine -1-yl)-3-fluorobenzonitrile as form K. ¹ H NMR (400 MHz, DMSOO δ ppm 10.96 (s, 1H) 7.68 (dd, J=13.45, 1, 83 Hz, 1H) 7.56 (dd, J=8.44, 1.83 Hz, 1H) 7.43-7.52 (m, 3 H) 7.29-7.39 (m, 4H) 7 .11 (t, J=8.80 Hz, 1H) 5.24 (s, 2H) 5.11 (dd, J=13.20, 5.14 Hz, 1H) 4.22-4.46 (m , 2H) 3.54 (s, 2H) 3.12-3.22 (m, 4H) 2.85-2.97 (m, 1H) 2.53-2.62 (m, 2H) 2.38 -2.48 (m, 2H) 1.93-2.03 (m, 1H). MS (ESI) m/z 568.2 [M+1]+.

6.2 Antiproliferative effects on multiple myeloma cells

Cell culture materials: Human multiple myeloma cell lines were purchased from suppliers and cultured at 37°C with 5% CO2 in medium as indicated in Table 1. 1. Cell lines resistant to lenalidomide and pomalidomide were generated by methods generally described previously (Lopez-Girona et al Leukemia 2012; 26 (11): 2335). All cell lines were maintained in log phase, and cell density and viability were monitored by trypan blue exclusion using a Vicell XR Cell Viability Analyzer (Beckman Coulter, Brea, CA).

Table 1 Multiple myeloma cell lines tested

MM cell line Supplier/Source Catalog number Cultivation conditions NCI-H929 ATCC (Manassas, Virginia) CRL-9068 RPMI-1640, 10% FBS NCI-H929-1051 own development, resistant to lenalidomide No data RPMI-1640, 10% FBS OPM2 DSMZ (Brunschweig, Germany) ACC-50 RPMI-1640, 10% FBS OPM2-P10 own development, resistant to 10 µM pomalidomide No data RPMI-1640, 10% FBS

Preparation of Study Solutions: Compound 1 was plated in black 384-well plates (Corning Inc.) to a final volume of 0.1% DMSO, assuming a maximum volume of 50 μL. A 10-point dose-response starting at 10 μM with a 1:3 dilution was printed in duplicate by acoustic dispensing using the EDC ATS-100 platform. Alternatively, a 10-point dose response was used, starting at 10 μM at a 1:10 dilution or starting at 100 nM at a 1:3 dilution.

Cell Proliferation Assays: The effect of Compound 1 on the proliferation/viability of hematological cell lines (Table 1) was assessed after 120 hours of incubation using CTG (Promega) according to the manufacturer's instructions. Hematology cell lines were dispensed into compound plates using a Multidrop Combi Reagent Dispenser (Thermo Scientific, Waltham, MA) at a concentration of 0.1 x 10 ⁶ cells per ml in a total volume of 50 μl. After 120 h, 25 μl of CTG per well was dispensed using a Multidrop Combi Reagent Dispenser, and adenosine triphosphate (ATP) release from viable cells was measured as relative luminescence units after 30 min using the Envision platform.

Results. Compound 1 demonstrates antiproliferative activity against MM cell lines. The MM cell lines selected for this study were those sensitive and resistant to lenalidomide and/or pomalidomide (Table 1), two agents approved for the treatment of patients with myeloma. Proliferation was assessed using CellTitre-Glo®

- 43 046729 test. Results for cultures incubated with compound 1 were normalized to results for control cultures for each cell line. The IC50 for cell growth inhibition by Compound 1 was determined for each cell line using ActivityBase software. Compound 1 effectively inhibited cell proliferation in four cell lines, as determined by quantifying the levels of ATP present in the medium after 120 hours. The antiproliferative IC50 values of Compound 1 ranged from 0.07 nM to 4.3 nM (Table 2) . Compound 1 showed very effective proliferative activity against multiple myeloma, even against cell lines resistant to lenalidomide and/or pomalidomide.

table 2

Inhibition of Cell Growth by Compound 1 in MM Cell Lines in Liquid Culture Medium

Conn. No. NCI-H929 120h IC50 NCI-H929.1051 120h IC50 OPM-2 120h IC50 OPM-2.P10 120h IC50 1 0.07 nM 1.0 nM 0.07 nM 4.3 nM

6.3 Off-target effects of compound 1.

α1-Adrenergic receptors and dopamine D2 receptors. Methods: Binding and functional assays for α1-adrenergic and dopamine D2 receptors were performed by Eurofins Cerep according to their methods.

a1-Adrenergic receptor. Binding at 10 pM. The binding assay assessed the affinity of the test drug for the non-selective α1-adrenergic receptor in the rat cerebral cortex. Membrane homogenates of the cerebral cortex were incubated in duplicate for 60 min at room temperature with 0.25 nM [ ³ H]prazosin in the absence or presence of the test samples at 10 μM. After the incubation period, the samples were filtered through glass fiber filters, the filters were dried and then the radioactivity was counted using a scintillation counter. Results are expressed as the average percentage inhibition of control radioligand binding.

IC ₅₀ binding. To determine the IC ₅₀ of non-selective α1-adrenergic receptor binding, various concentrations of the test substance were incubated in duplicate with 0.25 nM [ ³ H]prazosin. The previously described compound 3-(4-((4-((4-(2,4-difluorophenyl)piperazin-1-yl)methyl)benzyl)oxy)-1-oxoisoindolin-2-yl)piperidin-2,6- dione (Example 5,285 in US Pat. No. 8,518,972) (compound A) was tested at 0.01-30 pM. Compound B, the S-enantiomer of Compound A, was tested at 0.0003-10 μM. Compound 1 was analyzed at 0.03-100 μM. Radioactivity was measured as described above. The IC50 was defined as the concentration causing half the inhibition of specific binding of the control.

Antagonistic activity. The antagonistic effects of test compounds on the α _1A and α _1B adrenergic receptors were measured using human receptor-transfected Chinese hamster ovary (CHO) cells. Antagonist activity was determined by measuring the effect of the compound on agonist (epinephrine)-induced calcium mobilization in the α _1A receptor assay or cAMP levels in the α _1B receptor assay. In these experiments, CHO cells were incubated in duplicate at room temperature with the test substance and epinephrine at 3 nM in the α _1A receptor assays or at 3000 nM in the α _1B receptor assay. Compound A was tested in the a _1A receptor assay at 0.01-30 μM. Compound B was tested in the a _1A and a _1B receptor assays at 0.0003-30 pM. Compound 1 was assayed at concentrations from 0.03 to 30 pM in the a _1A receptor assay and from 0.03 to 100 μM in the a _1B receptor assay. In the a _1A receptor assay, cytosolic calcium levels were measured fluorometrically using the Fluo4 Direct fluorescent probe. Intracellular cAMP levels in the adrenergic receptor a _1B assay were measured using homogeneous time-resolved fluorescence (HTRF). The IC ₅₀ of antagonism was defined as the concentration that caused half the response of the control agonist.

Dopamine D2 receptor. Binding at 10 pM. The binding assay assessed the affinity of test samples for the dopamine D2 receptor in transfected human embryonic kidney (HEK)-293 cells. To determine binding in the D _2S receptor assay, the test article was incubated with 0.3 nM [ ³ H]methylspiperone or 1 nM [ ³ H]7-hydroxy-2-KHdipropylaminotetraline (7-OH-DPAT). [ ³ H]Methylspiperone at a concentration of 0.3 nM was also used as a control ligand in the D2L binding assay. Cell membrane homogenates were incubated in duplicate at room temperature for 60 min with ligand in the absence or presence of test samples at 10 μM. After the incubation period, the samples were filtered through glass fiber filters, the filters were dried and then the radioactivity was counted using a scintillation counter. Results are expressed as the average percentage inhibition of control radioligand binding.

IC50 binding. To determine IC50 binding in D2 receptor assays, HEK-293 was tested.

- 44 046729 as described above, but with different concentrations of the test substance. Compound A was tested at 0.01-30 pM in a D2S radioligand binding assay. Compound B was tested at 0.0003-10 pM in D2S and D2L binding assays. Compound 1 was tested at 0.03-100 pM in the D2S assay and 0.01-100 pM in the D2L assay. The IC50 was defined as the concentration causing half the inhibition of specific binding of the control.

Agonistic activity. The agonism of test compounds at the dopamine D2S receptor was assessed using HEK-293 cells transfected with the human receptor. Agonistic activity was determined by measuring the effect of the compound on impedance modulation. In these experiments, HEK-293 cells were incubated in duplicate at 28°C with the test article. Compound A was tested at 0.01-30 μΙΗ. Compound B was tested at 0.0003-10 pM, while Compound 1 was tested at 0.01-10 pM. Dopamine (3 pM) was used as an agonist control. Impedance measurements were monitored for 10 min after ligand addition using cellular dielectric spectroscopy. _EC50 was defined as the concentration that produced half the maximum response compared to the response to the control agonist (dopamine).

Results. Binding at 10 μM at α1-adrenergic and dopamine D2 receptors was assessed for Compound 1, Compound A, Compound B, and a number of compounds exemplified in US Pat. No. 8,518,972 (as indicated by their example number Ex) (Table 3) While the previously disclosed compounds completely inhibited ligand binding at both receptors, surprisingly, compound 1 showed a significantly reduced ability to inhibit ligand binding, showing only 67/62% ( α1-adrenergic receptor) and 55/52% (dopamine D2S) inhibition of ligand binding, respectively.

Table 3

Effect of Compound A, Compound B, Compound 1 and Previously Described Compounds on the α1-Adrenergic and Dopamine D2 Receptor

ABOUT

RiAr ^:? ' _-R 2

Conn. No. R ¹ R ² X Stereo Adrenergic α1 Dopamine D _2S % Inhibition (@10 µM) % Inhibition (@10 µM) 1 CN F CH2 S 62 52 A F F CH2 rac 102 99 B F F CH2 S 98 99 Etc. 5 229 H H CH2 rac 98.3 98.7 Etc. 5 273 F H CH2 rac 100.3 94.7 Etc. 5 289 F H CO rac 97.9 92.4

6.4 Compatibility study of the drug and excipient

A binary drug-excipient compatibility study was conducted to identify suitable excipients for capsule formulation. The list of excipients and ratios of API to excipients are given in the following

- 45 046729 table. These binary mixtures were studied in an open dish and subjected to 5°C (control) and 50°C/75% relative humidity conditions for 3 and 6 weeks, respectively. Selective samples were tested for chemicals (impurities/degradants) and racemization (loss of chiral purity). Possible degradation pathways that may limit the shelf life of compound 1 include hydrolysis, oxidation, and racemization (conversion of the S isomer to the R isomer). The results of impurities from hydrolysis and oxidation are shown in FIG. 3, and the results of chiral impurities are shown in FIG. 4.

Table 4

List of samples assessed for compatibility with the drug substance

Ingredients Trade name Function API to Excipient Ratio Connection 1 API 1 Microcrystalline cellulose Avicel 102 Diluent 1:200 Lactose monohydrate, spray dried Fast Flo 316 Diluent Spray dried mannitol Pearlitol SD200 Diluent Partially pregelatinized starch Starch 1500 Diluent Tween 80 Tween 80 Wetting agent/solubilizer/crystallization stabilizer Poloxamer 188 Lutrol F68 Wetting agent/solubilizer/crystallization stabilizer Sodium Lauryl Sulfate (SLS) No data Wetting agent/solubilizer/crystallization stabilizer HPBCD Kleptosis Solubilizer/crystallization stabilizer Vitamin E-TPGS No data Solubilizer/crystallization stabilizer Croscarmellose sodium Ac-di-Sol SD-711 Baking powder Crospovidone Kollidon CL Baking powder Sodium starch glycolate (SSG) Explotab Baking powder HPMCAS-LF AQOAT AS Crystallization stabilizer HPMC E3 Hypromellose Binder/crystallization stabilizer PVP VA64 Kollidon VA64 Binder/crystallization stabilizer PVP K30 Plasdone K30 Binder/crystallization stabilizer HPC EXF Kluckel (hydroxypropylcellulose) Binder/crystallization stabilizer succinic acid No data Acidifier Fumaric acid Pharmaceutical grade Acidifier Silica dimethyl silylate Aerosil R-972 Substance that promotes anti-slip/anti-agglomerate Silica Aerosil 200 Substance that promotes slipping

- 46 046729

Silica Aerosil 300 Substance that promotes slipping Talc No data Release agent Stearic acid Kolliwax Lubricant Magnesium stearate Hyqual Lubricant HPMC capsule Vcaps capsules (size #4) Capsule shell 1 Gelatin capsule (Size #4) Capsule shell

Compound 1 undergoes hydrolysis at elevated levels of temperature and humidity (50°C/75% RH). Diluents such as mannitol, starch and lactose were evaluated as primary and secondary carriers. Lactose was found to be compatible with a slight decrease in chiral purity. Mannitol and starch have also been found to be compatible with some hydrolytic chemical degradation. A surfactant such as sodium lauryl sulfate (SLS) resulted in loss of chiral purity and increased chemical degradation, mainly due to hydrolysis. Of the disintegrants tested, sodium starch glycolate (SSG) was found to be compatible, while crospovidone enhanced chemical degradation through oxidation and hydrolysis. Among the polymers evaluated as precipitation inhibitors, HPMC-E3 caused a significant increase in unknown impurity (RRT 0.39), while PVP-K30 and PVP-VA64 resulted in the formation of a potential oxidative degradation product (RRT 0.66). In addition, PVP-K30 and PVP-VA64 not only cause chemical degradation through hydrolysis, but also cause a loss of chiral purity. Empty capsules based on HPMC and gelatin showed good compatibility. Of the shortlisted acidulants, succinic acid was found to be chemically incompatible, causing hydrolysis and also leading to loss of chiral purity. Fumaric acid was indicated as a more compatible acidifier, although at an increased ratio of 1:40 w/w. it may cause chemical decomposition through hydrolysis and loss of chiral purity. Glidants/deagglomerants were also evaluated; Aerosil R972 was found to be the most compatible, while Aerosil 300 and Aerosil 200 caused some chemical incompatibility due to hydrolysis and loss of chiral purity. Of the hydrophilic Aerosils (200 and 300), 200 was found to have better compatibility than Aerosil 300 due to the latter's large surface area - 300 m ² /g versus 200 m ² /g, catalyzing greater hydrolysis. Stearic acid was found to be chemically compatible but caused some loss of chiral purity.

Thus, lactose, mannitol, pregelatinized starch, fumaric acid, stearic acid, Aerosil R972, Aerosil 300, HPBCD, sodium starch glycolate and crospovidone were selected for further evaluation in the formulas. SLS, PVP-K30, PVP-VA64 and Aerosil 300 were found to result in both loss of chiral purity and chemical degradation, whereas Aerosil 200 affected only chemical degradation.

6.5 Stability screening of mannitol and starch-lactose BIC formulations

The stability of a prototype mixture in capsules (BIC) based on mannitol and starch lactose as primary carriers was evaluated. The mixtures were prepared by dry mixing using a Turbula mixer and filled with size 4 white opaque HPMC capsules using Profill 100 and the components of the mixtures are presented in the following table. To quickly assess the stability of the BIC formulation, an accelerated 3-day stability screening study was conducted.

Table 5

Prototype BIC formulations based on mannitol and starch-lactose (Cap-1 to Cap-12)

Serial number Cap-1 Cap-2 Cap-3 Cap-4 Cap-5 Cap-6 Cap-7 Cap-8 Cap-9 Cap10 Cap11 Cap12 Ingredients % w/w Connection 1 0.13 0.13 0.13 0.13 0.13 0.13 0.13 0.13 0.13 0.13 0.13 0.5 Silica dimethyl silylate (Aerosil R972) 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 Colloidal dioxide silicon (Aerosil 300 Pharma) 0.2 Partially Pregelatinized Corn Starch (Starch 1500) 30.0 30.0 64.67 64.67 44.0

- 47 046729

Lactose anhydrous (Supertab 24 AN) 66.67 64.67 30.0 28.0 22.0 Mannitol (Pearlitol SD 200) 66.0 66.0 68.0 91.67 94.67 94.67 94.3 Crospovidone (Kollidon CL) 2.0 2.0 2.0 2.0 2.0 Sodium starch glycolate (Explotab) 2.0 Polyvinylpyrrolidone (PVPK30) 26.67 HPBCD (Kleptose) 26.67 26.67 Hypromellose (HPMC-E3) 26.67 Sodium lauryl sulfate 1.0 Fumaric acid Pharmaceutical Grade (Powder) 2.0 2.0 2.0 2.0 2.0 4.0 2.0 2.0 2.0 Stearic acid (Hystrene 5016 Veg) 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0 Total (% w/w) 100.0 Total capsule filling mass (mg) 75.0 100.0

The results of the accelerated stability screening are shown in FIG. 5A-5D and FIG. 6.

RRT 4.2 min: Capsule-7 (mannitol BIC) containing HPMC-E3 was the least stable, followed by Capsule-5 (starch-lactose BIC) containing HPBCD, and Capsule-8 (mannitol BIC) containing HPBCD . Polymers and complexing agents were found to increase the degradation product at an RRT of 4.2 min. Increasing humidity levels catalyze the decomposition product at an RRT of 4.2 min.

The stability ranking order for formulations showing an increase in degradation product at 4.2 min is: Capsule-9 & 10 > Cap-12 > Cap-11 > Cap-1 > Cap-2 > Cap-4 > Cap3-> Cap6. > Cap-8 > Cap-5 > Cap-7.

Overall, mannitol-based batches containing fumaric acid were the most stable, followed by starch-lactose-based batches with or without fumaric acid.

RRT 7.6 min: Capsule-3 (starch-lactose BIC) was found to be the least stable, followed by Cap-7 (mannitol BIC) containing HPMC-E3, and Cap-2 (starch-lactose BIC). Interestingly, Cap-2 and Cap-3 contain the same excipients, except for the starch-lactose proportions. For Cap-2 formulation, starch is 30% w/w and lactose is 64.67% w/w, whereas for Cap-3 formulation, starch is 64.67% w/w and lactose is 30% w/w, indicating that a higher proportion of starch may lead to an increase in the degradation product RRT 7.6 min. Cap-5 (starch-lactose BIC) containing HPBCD was also found to be unstable. Cap-6 (mannitol BIC) containing PVP-K30 was also found to be less stable compared to the mannitol-based BIC formulation without PVP-K30, indicating instability induced by PVP-K30.

The stability ranking order for formulations demonstrating an increase in degradation product at 7.6 min is as follows: Capsule-9 & 11 > Cap-12 > Cap-10 > Cap-8 > Cap-4 > Cap-6 > Cap-1 > Cap5 > Cap-2 > Cap-7 > Cap-3.

In general, mannitol-based BIC formulations containing fumaric acid were found to be more stable.

RRT 7.1 min: Capsule-3 (starch-lactose BIC) containing crospovidone was the least stable. Capsule-1 (starch-lactose BIC) without crospovidone was found to have a significantly better stability profile. Capsule-7 (mannitol BIC) containing HPMC-E3 was also found to be unstable, followed by Cap-6 (mannitol BIC) containing PVP-K30, and Cap-5 (starch lactose BIC) containing HPBCD. RRT 7.1 min decomposition impurities increased with increasing temperature (70°C) at 0% relative humidity.

The stability ranking order for formulations showing an increase in degradation product RRT at 7.1 min is as follows: Capsule-11 > Cap-10 > Cap-12 > Cap-1 > Cap-4 > Cap-9 > Cap-8 > Cap5 > Cap -6 > Cap-7 > Cap-2 & Cap- 3.

RRT 11.3 min: Capsule-9 (mannitol BIC) containing SLS was the least stable. In general, all mannitol-based formulations with Aerosil R972 or Aerosil 300 Pharma were found to be the least stable. Of the starch-lactose formulations, Cap-3 with crospovidone was the least stable. In contrast, starch-lactose BIC formulations without crospovidone were found to have a significantly better stability profile (Cap-1), but mannitol-based BIC formulations showed higher degradation.

Stability ranking order for formulations showing an increase in product RRT

- 48 046729 decomposition in 11.3 min, the following: Capsule-6 > Cap-1 > Cap-8 > Cap-5 > Cap-4 > Cap-2 > Cap-7 > Cap-3 > Cap-12 > Cap- 11 > Cap-10 > Cap-9.

Loss of Chiral Purity: From FIG. 6, Cap-9 (mannitol BIC) containing SLS was found to enhance conversion to the R-isomer. In general, starch- and lactose-based BIC formulations with or without crospovidone and fumaric acid also enhance conversion to the R-isomer (capsules 1, 2, 3, and 4). Overall, mannitol-based BIC formulations with PVP-K30 (Cap-6) and HPBCD (Cap-8) were found to be the most stable. Mannitol-based BIC formulations with HPMC-E3 and starch-lactose HPBCD were also found to be more stable than formulations without polymers or complexing agents. Increasing temperature and decreasing humidity have also been found to contribute to the loss of chiral purity.

The stability ranking order for formulations exhibiting increasing loss of chirality purity is as follows: Capsule 6 > Cap-8 > Cap-12 > Cap-5 > Cap-7 > Cap-10 > Cap-11 > Cap-4 > Cap-2 > Cap -3 > Cap-1 & Cap-9.

In summary, mannitol-based BIC formulations containing fumaric acid were found to exhibit the best overall stability profiles. Although the inclusion of polymers or complexing agents has been found to increase degradation products, they have nevertheless been found to prevent loss of chiral purity. Starch lactose-based BIC formulations without fumaric acid (Cap-1, Cap-2 and Cap-3) were found to be unstable (chiral and related impurities) compared to formulations with fumaric acid (Cap-4 and Cap-5). Therefore, fumaric acid was chosen as an acidulant to improve the stability of the drug. Additionally, the stability profile of the drug product was found to improve with higher drug loading, as seen in Cap-10 (0.13% w/w Compound 1) compared to Cap-12 mannitol-based BIC formulations ( 0.5% w/w Compound 1). Starch-lactose ratios were optimized by evaluating their stability profiles at 1:2 w/w. compared to 2:1 w/w. Formulations with a higher proportion of lactose (2 parts) were found to be more stable than formulations containing a higher proportion of starch (2 parts), as seen in the stability results for starch-lactose BIC formulations Cap-1 to Cap-4 .

6.6 Precipitation risk assessment of prototype BIC formulations

The risk of precipitation of prototype BIC formulations was studied using a two-stage method (Stage 1 under immersion conditions and Stage 2 under non-immersion conditions) at a high concentration (2 mg). The formulation compositions are presented in the following table.

Table 6

Prototype batches of formulations to assess the risk of precipitation during two-step dissolution of 2 mg BIC

An object Screening for sedimentation inhibition 2 m BIC Serial number Cap19 Cap20 Cap21 Cap22 Ingredients % w/w % w/w % w/w % wt./wt. Connection 1 0.5 0.5 0.5 0.5 Silica dimethyl silylate (Aerosil R972) 2.0 2.0 2.0 2.0 Mannitol (Pearlitol 200 SD) 91.5 77.55 Partially Pregelatinized Corn Starch (Starch 1500) 29.9 23.9 Lactose anhydrous (Supertab 24 AN) 61.6 53.65 Polyvinylpyrrolidone (PVP-K30) HPBCD, 97% 13.95 13.95 Pharmaceutical Grade Fumaric Acid (Powder) 3.0 3.0 3.0 3.0 Stearic Acid (Hystrene 5016 Veg) 3.0 3.0 3.0 3.0 Total (% w/w) 100.0 100.0 100.0 100.0 Total capsule filling mass (mg) 400.0 400.0 400.0 400.0

A two-stage dissolution study was carried out on prototype compositions, in which the dissolution medium for stage 1 was 0.01 N. HCl. From two-step dissolution studies, no precipitation was observed at pH 6.8 (90-150 min) for any mannitol-starch based BIC (Figure 7). Based on the two-step dissolution results, the risk of precipitation is considered low for Compound 1.

The dissolution release profiles for starch and lactose based batches (Cap-19 and Cap-20) are observed to be slower than those for mannitol based batches (Cap-21 and Cap-22) with and without HPBCD. It is believed that the retardation of dissolution is due to the following: plug formation (gelation) inside the HPMC capsule and the level of Aerosil R972 hydrophobic silica on

- 49 046729 level 2% w/w. Thus, optimization of R972 was investigated and it was found that a level of 0.5% Aerosil R972 in a starch-lactose BIC formulation achieved a dissolution profile comparable to that of a mannitol-based BIC formulation.

When combinations of mannitol with HPBCD (Cap-22) and starch-lactose with HPBCD (Cap-20) were evaluated compared with formulations without HPBCD (Cap-19 and Cap-21), the dissolution release profiles of the formulations without HPBCD (Cap-19 and Cap-21) Cap-21) were faster than their combinations (Cap-20 and Cap-22). Thus, based on the two-step dissolution results, no precipitation was observed for the BIC formulation without any precipitation inhibitor or solubilizer. Therefore, it was determined that there was no need to include a precipitation inhibitor or solubilizer in the capsule formulation and that the risk of precipitation was considered low for Compound 1.

6.7 Screening of Prototype Formulations to Achieve High Dissolution Release Rates from Starch-Lactose BIC Formulations

Due to plugging, the dissolution release profile for the 2 mg batch of starch-lactose BIC (Cap-19) was slower than the batch of mannitol-based BIC (Cap-21) as shown in FIG. 7. Formulation optimization was carried out to achieve high release rate using different disintegrants as shown in the following table (Cap24, Cap-25 and Cap-26). The dissolution release profiles of these formulations were compared to a formulation of Cap-13, 0.1 mg BIC (same formulation as 2.0 mg Cap-19 BIC), and Cap-27 (2 mg binary BIC).

Table 7

Prototype batches of starch-lactose formulations to optimize dissolution rates by evaluating different grades of disintegrants

An object Dissolution screening of 0.1 mg BIC based on starch-lactose 2.0 mg binary BIC Serial number Cap13 Cap24 Cap25 Cap26 Cap27 Ingredients % w/w % wt./wt. % wt./wt. % wt./wt. % w/w Connection 1 0.13 0.13 0.13 0.13 90.09 Silica dimethyl silylate (Aerosil R972) 2.0 2.0 2.0 2.0 9.91 Partially Pregelatinized Corn Starch (Starch 1500) 30.0 28.0 27.0 27.0 Lactose anhydrous (Supertab 24 AN) 61.87 62.87 62.87 62.87 Crospovidone (Kollidon CL) 2.0 Sodium starch glycolate (Explotab) 2.0 Croscarmellose sodium (Ac-di-Sol SD-711) 2.0 Pharmaceutical Grade Fumaric Acid (Powder) 3.0 3.0 4.0 4.0 Stearic Acid (Hystrene 5016 Veg) 3.0 2.0 2.0 2.0 Total (% w/w) 100.0 100.0 100.0 100.0 100.0

The dissolution release profiles were found not to differ significantly between different disintegrants (Cap-24, Cap-25 and Cap-26). The BIC formulation Cap-25 (2% w/w sodium starch glycolate) showed a slightly better dissolution release profile than Cap-24 (2% w/w crospovidone) and Cap-26 (2% w/w . croscarmellose sodium), respectively. The decay time profiles for Cap-24, Cap-25 and Cap-26 were not found to be significantly different, but were nevertheless better and more consistent for Cap-25 containing 2% w/w. SSG, compared to Cap24 and Cap-26. The obvious role of hydrophobic Aerosil R972 compared to formulations with disintegrants is evident from the dissolution profile of 2 mg binary BIC with 10% w/w. Aerosil R972 (Cap-27), which showed the slowest dissolution release profile among the five drugs. The control mannitol formulation (Cap-13 without disintegrant) showed a slightly better overall dissolution release profile, which also contained 2% Aerosil R972, indicating that the effect of hydrophobic Aerosil R972 on slowing the dissolution release profile is more significant for BIC-based formulations. starch and lactose. Overall, no significant difference in disintegration or dissolution was observed with the inclusion of SSG, croscarmellose sodium, or crospovidone disintegrant. The inclusion of SSG resulted in a constant dissolution profile.

Further exploratory dissolution studies were conducted to determine the slower dissolution release profile for starch-lactose BIC with sodium starch glycolate selected as the disintegrant of choice due to better stability of the binary mixture and lack of negative effects on dissolution. By varying levels of hydrophobic silica (Aerosil R972), dissolution rates (50 rpm vs. 75 rpm) were evaluated to resolve the slower dissolution release profile associated with starch-based formulations and

- 50 046729 lactose, as shown in the following table (Cap-29, 33 and 34).

Table 8

Pilot batches of starch-lactose formulations to optimize dissolution rates by varying levels of Aerosil R972 and varying levels of sodium starch glycolate

An object Dissolution screening of 0.1 mg BIC based on starch lactose Serial number Cap25 Cap29 Cap33 Cap34 Ingredients % w/w % wt./wt. % wt./wt. % w/w Connection 1 0.13 0.13 0.13 0.13 Silica dimethyl silylate (Aerosil R972) 2.0 1.0 1.0 0.5 Partially Pregelatinized Corn Starch (Starch 1500) 27.0 24.0 22.0 22.0 Lactose anhydrous (Supertab 24 AN) 62.87 65.87 60.87 61.37 Sodium starch glycolate (Explotab) 2.0 2.0 10.0 10.0 Pharmaceutical Grade Fumaric Acid (Powder) 4.0 5.0 4.0 4.0 Stearic Acid (Hystrene 5016 Veg) 2.0 2.0 2.0 2.0 Total (% w/w) 100.0 100.0 100.0 100.0

As shown in FIG. 8, the dissolution release profile of Cap-34 is significantly superior (~80% drug release in 30 min) compared to Cap-25 (control - 30% drug release in 30 min). Although drug release is still not 100%, it can be concluded that decreasing Aerosil R972 levels from 2% to 0.5%, increasing sodium starch glycolate levels from 2% to 10%, and increasing dissolution rate from 50 rpm. up to 75 rpm had a synergistic effect on the overall dissolution characteristics of Cap-34.

Based on the dissolution results, the level of Aerosil R972 was reduced to 0.5 wt.%. For 0.1 mg BIC formulations based on starch and lactose.

6.8 Evaluation of different acidulants and their levels for mannitol and starch lactose based BIC formulations

Since compound 1 degrades at pH above 5, a low pH environment for the drug product is desirable. To achieve a low pH microclimate for the drug product, an acidifier was included in the BIC.

Three acidulants, namely fumaric acid (pKa- 3.0), maleic acid (pKa- 1.91) and succinic acid (pKa- 4.18), were evaluated to achieve low pH of starch-lactose media and BIC formulations based on mannitol. Of these acids, fumaric acid and maleic acid were further evaluated based on their lower pKa, while succinic acid was found to be less compatible with the drug substance.

The pH of mixtures of formulations with and without acidifiers was measured using the suspension method. 200 to 400 mg of starch-lactose BIC and mannitol-based BIC mixtures were weighed and diluted with 0.5 ml distilled water. The dispersion was shaken for 2 min to obtain a homogeneous suspension for measuring its pH.

Based on the evaluated data, as shown in the following tables, the starch-lactose BIC formulation without acidifier has a pH of 6.3. This high pH environment may be detrimental to drug stability. Addition of fumaric acid and maleic acid at levels of 2%, 3% and 5% was evaluated to lower the pH for starch-lactose based BIC batches. Because minor adjustments to the levels of the stronger acidifier (maleic acid) significantly lowered the pH, which could also have a detrimental effect on formulation stability, fumaric acid was chosen as the acidifier. However, due to the slower dissolution of the starch-lactose based formulation, the inclusion of disintegrants was important. Additional pH evaluations were performed to optimize the pH of starch-lactose BIC formulations containing fumaric acid in the presence of various disintegrants. The pH of the mannitol-based BIC composition with 3% fumaric acid was found to be 2.2, which is within the desired pH range of 2-3.

Table 9

Evaluation of the type and level of acidifiers to optimize starch-lactose and mannitol-based BIC formulations

Composition type Acidifier/Level pH Starch-lactose No data 6.3 Starch-lactose Fumaric acid-2% 2.3

- 51 046729

Starch-lactose Fumaric acid-3% 2.2 Starch-lactose Fumaric acid-5% 2.1 Starch-lactose Maleic acid-2% 1.7 Starch-lactose Maleic acid-3% 1.4 Starch-lactose Maleic acid-5% 1.2 Mannitol Fumaric acid-3% 2.2

Table 10

Evaluation of fumaric acid levels for optimization of starch-lactose formulations with or without ___________________________disintegrants___________________________

Composition type Baking Powder/Level Acidifier/Level pH Starch-lactose Fumaric acid-3% 3.2 Starch-lactose Crospovidone-2% Fumaric acid-3% 3.4 Starch-lactose SSG-2% Fumaric acid-3% 3.9 Starch-lactose Croscarmellose sodium-2% Fumaric acid-3% 4.4 Starch-lactose SSG-2% Fumaric acid-4% 2.8 Starch-lactose Croscarmellose sodium-2% Fumaric acid-4% 2.9 Starch-lactose SSG-10% Fumaric acid-5% 2.9

Based on these pH studies, a 3% fumaric acid level was selected for mannitol-based BIC formulations, while a 4% fumaric acid level was selected for starch-lactose BIC formulations containing 10% sodium starch glycolate to maintain the pH of the mixture at range 2-3.

6.9 Comparative evaluations of direct mix (low shear) and high shear mixing processes for BIC formulations to achieve acceptable content uniformity

Mannitol and starch-lactose formulations were evaluated for content uniformity using low and high shear mixing processes. Capsules were collected throughout the encapsulation cycle and content homogeneity was assessed using stratified CU sampling. All prototype formulations with their compositions and manufacturing processes are listed in the following table.

Table 11

Prototype formulations with their processes for the production of 0.1 mg BIC based on mannitol and starch-lactose to achieve acceptable content uniformity

Manufacturing process High shear mixing and co-grinding Dry blending with co-grinding HSM with joint grinding Serial number Cap 13 Cap 15 Cap 16 Cap 18 Cap28 Cap31 Ingredients % wt./ wt. % wt./wt. % wt./wt. % wt./wt. % wt./wt. % w/w Connection 1 0.13 0.13 0.13 0.133 0.13 0.13 Silica dimethyl silylate (Aerosil R972) 2.0 2.0 2.0 2.0 1.0 1.0 Mannitol (Pearlitol 200 SD) 91.87 77.87 91.87 91.87 Partially pre-gelatinized corn starch (Starch 1500) 30.0 24.0 Anhydrous lactose (Supertab 24 AN) 61.87 53.87 HPBCD, 97% 14.0 14.0 Fumaric acid Pharmaceutical Grade (Powder) 3.0 3.0 3.0 3.0 5.0 5.0 Stearic acid (Hystrene 5016 Veg) 3.0 3.0 3.0 3.0 2.0 2.0 Total (% w/w) 100.0 100.0 100.0 100.0 100.0 100.0

- 52 046729

Total capsule filling weight (mg) 75.0 75.0 75.0 75.0 75.0 75.0

Overall, starch-lactose BIC batches were found to be superior in achieving an activity distribution with high shear mixing and co-milling, while mannitol-based batches had a relatively poor activity distribution with either a high shear mixing process or a high shear mixing process. by force and co-grinding, or with direct mixing and co-grinding, respectively. This may be due to material adhering to processing equipment and tamping pins during production. Preliminary content uniformity studies have shown that Cap-15 (starch-lactose) containing HPBCD produced by high shear mixing with co-milling has the best activity distribution (mean - 99.1%, RSD - 1.9%) with an acceptability value (AV) of 4.4. However, the chemical degradation profile in the presence of HPBCD showed increased degradation levels.

The ranking order of content uniformity for the evaluated BIC formulations is Capsule-15 > Capsule-13 > Capsule-18 > Capsule-28 > Capsule-16 > Capsule-31. The ranking order was based on activity distribution (%RSD), assay mean (%) and increase in acceptance score (AV) as shown in the following table.

Table 12

Content homogeneity ranking order based on activity distribution (%RSD), assay mean (%) and acceptance score (AV)

The consignment Compound Manufacturing process Average analysis value (%) % RSD Acceptability Index (AV) Capsule-15 Starch lactose Mixing with joint grinding with high shear forces 99.1 1.9 4.4 Capsule-13 Starch lactose Mixing with joint grinding with high shear forces 97.4 1.7 5.1 Capsule-18 Mannitol Mixing with joint grinding with high shear forces 97.6 3.9 8.2 Capsule-28 Mannitol Dry blending with co-grinding 94.4 2.4 9.8 Capsule-16 Mannitol Mixing with joint grinding with high shear forces 86.0 1.8 17 Capsule-31 Mannitol Mixing with joint grinding with high shear forces 88.3 9.6 33.3

The relatively poor recovery in the analysis of mannitol-based formulations can be attributed to high shear mixing causing the mixture to adhere to the treated surfaces. Therefore, the direct mixing (low shear) method was chosen as the method for further development.

6.10 Starch-lactose BIC vs. mannitol BIC - accelerated stability assessment

Accelerated stability assessments were performed for starch-lactose BIC and mannitol-based BIC formulations under the following storage conditions: 50°C/50% RH for T=3 days, 60°C/30% RH for T=3 days, 70°C/0% RH for T=3 days and 70°C/50% RH for T=3 days. Data were compared with samples stored at 5°C (control).

According to accelerated stability studies (Fig. 9), the inclusion of HPBCD (Cap-15) in the formulations

- 53 046729 based on starch and lactose helped maintain chiral purity compared to formulations without a stabilizer (Cap-13). For mannitol-based formulations, HPBCD (Cap-18) also helped maintain chiral purity compared to formulations without HPBCD (Cap-28 and Cap-31). High levels of fumaric acid (5% w/w) for Cap-28 and Cap-31 also had a negative effect on chiral purity compared to (3% w/w) fumaric acid for mannitol-based formulations Cap-16 . Overall, the loss of chiral purity for starch-lactose formulations without HPBCD (Cap-13) was lower than for mannitol-based formulations without HPBCD (Cap-16).

The inclusion of a stronger acid (maleic acid) was also analyzed to evaluate its effect on the stability profile of the BIC formulations, as shown in the following table.

Table 13

Comparative accelerated stability assessment of mannitol-based BIC with fumaric acid and maleic acid

Manufacturing process Direct mixing with joint grinding HSM with co-grinding HSM with co-grinding Serial number Cap28 Cap31 Cap32 Ingredients % w/w % w/w % w/w Connection 1 0.13 0.13 0.13 Silica dimethyl silylate (Aerosil R972) 1.0 1.0 1.0 Mannitol (Pearlitol 200 SD) 91.87 91.87 91.87 Pharmaceutical Grade Maleic Acid 5.0 Pharmaceutical Grade Fumaric Acid (Powder) 5.0 5.0 Stearic Acid (Hystrene 5016 Veg) 2.0 2.0 2.0 Total (% w/w) 100.0 100.0 100.0 Total capsule filling weight (mg) 75.0 75.0 75.0

In fig. 10 shows that the inclusion of maleic acid in the formulation (Cap-32) resulted in better chiral stability, but resulted in an increase in the amount of chemical degradation products as shown in FIG. 11 compared to formulations containing fumaric acid (Cap-28 and Cap-31). Thus, fumaric acid was chosen as the acidulant at levels of 3% w/w. for BIC compositions based on mannitol and 4% w/w. for BIC compositions based on starch - lactose with 10% sodium starch glycolate as a disintegrant.

To evaluate the effect on drug product stability of the inclusion of a disintegrant (crospovidone, SSG or croscarmellose sodium) for starch-lactose BIC formulations, comparative accelerated stability screening studies were conducted for 0.1 mg starch-lactose BIC formulations with (Cap-24, Cap -25, Cap-26) and no disintegrant (Cap-13) compared to a BIC formulation based on 0.1 mg mannitol (Cap-16).

From fig. 12, Cap-25 containing a starch-lactose based BIC with SSG is less prone to oxidation (oxidative degradant 1, RRT 0.66) except at very high temperature and humidity (70°C/50% RH). A formulation containing crospovidone (Cap-24) undergoes significant oxidation at higher temperatures. The stability of the starch-lactose based BIC profile without any disintegrant (Cap-13) was found to be the most stable. The mannitol based formulation (Cap-16) undergoes oxidation primarily at elevated temperature and low humidity conditions (60°C/30% RH and 70°C).

Similarly, Cap-26 containing croscarmellose sodium was found to be prone to oxidation (oxidative degradant 2), but its level remained unchanged under all storage conditions (Fig. 13). Cap-24 containing crospovidone was found to have such same stability profile as Cap-25 containing SSG, however Cap-25 is superior due to its better tolerability of oxidative degradant 1. In general, formulations containing starch-lactose without disintegrants (Cap-13) demonstrate better tolerability relative to to oxidative decomposition agents compared to formulations with disintegrants (Cap-24, Cap-25 and Cap-26) or mannitol-based formulations (Cap-16).

The inclusion of disintegrants helped reduce hydrolytic impurities (FIG. 14 and FIG. 15), and based on the disintegrants evaluated, Cap-25 containing SSG was the most stable.

From fig. 16 it can be seen that the inclusion of disintegrants in the composition helped maintain chiral purity. Of the disintegrants evaluated, Cap-24 containing crospovidone and Cap-25 containing SSG were found to have better stability.

Shelf life prediction of mannitol-based BIC formulation was assessed by the Accelerated Stability Assessment Program (ASAP) for chiral purity and degradation products. Based on

- 54 046729 on ASAP study for BIC formulation based on 0.1 mg mannitol (Cap-16), the predicted shelf life of BIC formulation based on 0.1 mg mannitol at loss of chiral purity (Spec-3%) is 3+ years when stored at room temperature without desiccant.

Based on ASAP predictive modeling for a 0.1 mg mannitol based BIC formulation (Cap16), the predicted shelf life of 0.1 mg mannitol BIC is 3+ years when stored at room temperature without a drying agent for oxidative degradation.

Based on ASAP predictive modeling for BIC formulations based on 0.1 mg mannitol, the shelf life of 0.1 mg mannitol BIC when stored at room temperature without a desiccant is predicted to be 1.5+ years for the primary hydrolysis degradant.

Based on ASAP predictive modeling for mannitol-based BIC formulations, the shelf life of 0.1 mg BIC mannitol when stored at room temperature without a desiccant is predicted to be 3+ years for a secondary hydrolysis degrader.

Since hydrolysis has been found to be a major limitation on shelf life, adding 0.5 g silica to HDPE bottles is predicted to slow hydrolysis and increase shelf life from the previously predicted shelf life of 1.5+ years to 2.9+ years respectively.

Thus, from the preliminary ASAP study, the predicted shelf life at room temperature for a mannitol-based BIC formulation without silica gel is about 1.5+ years, while with 0.5 g silica gel at room temperature is about 2.9+ years.

Similarly, shelf life prediction for starch-lactose based BIC formulation was also assessed by the Accelerated Stability Assessment Program (ASAP) for chiral purity and associated impurities. Based on the ASAP study for the 0.1 mg starch-lactose BIC formulation (Cap34), the predicted shelf life of the 0.1 mg starch-lactose BIC formulation upon loss of chiral purity is 3+ years when stored at room temperature without silica gel desiccant.

Based on ASAP predictive modeling for BIC formulations with 0.1 mg starch-lactose, the shelf life of 0.1 mg BIC starch-lactose when stored at room temperature without a desiccant is predicted to be 2.5+ years for the primary hydrolysis degradant.

Similarly, based on ASAP predictive modeling for BIC formulations with 0.1 mg starch-lactose, the shelf life of 0.1 mg BIC starch-lactose when stored at room temperature without a desiccant is predicted to be 3+ years for the secondary hydrolysis degrader.

Based on ASAP predictive modeling for the starch-lactose BIC formulation (Cap-34), the predicted shelf life of 0.1 mg starch-lactose BIC is 3+ years when stored at room temperature without an oxidative degradant desiccant.

Thus, the shelf life limiting degradation pathway for the 0.1 mg starch-lactose (Cap-34) BIC formulation may be hydrolysis without silica gel desiccant. Predicted shelf life is 2.5+ years when stored at room temperature.

In general, the predicted shelf life for both formulations is 1.5-2+ years when stored at room temperature, and the shelf life limitation pathway is hydrolysis.

6.11 Evaluation of hydrophobic and hydrophilic silica in 0.1 mg formulations

Agglomeration and surface adhesion of cohesive APIs is a challenge in the development of solid dosage forms, especially for formulations with low strength. The crystal morphology of compound 1 is hydrophobic agglomerated clusters, which were wet-milled at the seeding stage. Compound 1 tends to stick, and poor flow and surface adhesion were observed during processing.

Scanning electron microscope (SEM) images of a crystal of Compound 1, uncoated and coated with Aerosil R-972, are shown in FIG. 17A and FIG. 17V, respectively. The images show that predispersing the crystals of Compound 1 with hydrophobic silica Aerosil R-972 (coating Compound 1 with Aerosil R-972) overcame the agglomeration and adhesion problem.

The effectiveness of controlling hydrophobic silica agglomeration and cohesion is evident in a comparison of hydrophilic silicas (Aerosil 200 and Aerosil 300) as shown in the SEM images of FIG. 18A-18D.

The adhesion of Compound 1 to process containers was further reduced by pre-treating the containers with a lubricant mixture.

6.12 Evaluation of alternative silica grades by dissolution release profile for 2 mg BIC formulations.

Due to limitations associated with global regulatory acceptability of silica dimethyl silylate (Aerosil R972), as well as the low maximum permissible limit (0.5 mg w/w per unit for IR capsules) according to the USFDA Inactive Ingredients Database (IIG) , alternative silica grades for higher doses (0.5 mg and 2 mg BIC) have been evaluated. The drug-excipient compatibility is shown in FIG. 19 show that Aerosil R972, due to its hydrophobicity, prevents drug degradation compared to Aerosil 200 Pharma or Aerosil 300 Pharma. The degree of stability was as follows: Aerosil R972> Aerosil 200> Aerosil 300.

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It was found that Aerosil 200 is not only relatively more compatible than Aerosil 300, but also offers a lower surface area of 200 m ² /g compared to the 300 m ² /g offered by Aerosil 300, the overall interaction of silica with drug substance can be minimized , thereby reducing degradation products and preventing loss of chiral purity. The stability of the low strength capsule (0.1 mg) was very difficult and acceptable stability could be achieved by coating the API with hydrophobic silica (Aerosil R972), so for the 0.1 mg concentration Aerosil R 972 was chosen as the glidant/deagglomerating/ stabilizing agent. For higher concentrations (0.5 and 2 mg), since the stability problem was less severe, the use of hydrophilic silica (Aerosil 200) was evaluated.

To evaluate the effect of replacing Aerosil R972 with Aerosil 200 Pharma on the properties of the drug product, dissolution studies were carried out. Because starch and lactose based batches were found to have slower dissolution profiles than mannitol based batches, starch-lactose 2 mg BIC pilot batches containing 0.5%, 1% and 2% Aerosil 200 were evaluated as shown in the following table.

Table 14

Comparative evaluation of different levels of Aerosil 200 Pharma based on release profile at

dissolution 2 mg starch-lactose BIC Manufacturing process Dry mix with manual filling Dry mix with manual filling Dry mix with manual filling Serial number Cap38 Cap39 Cap40 Ingredients % w/w % w/w % w/w Connection 1 1.0 1.0 1.0 Colloidal silicon dioxide (Aerosil 200 Pharma) 2.0 1.0 0.5 Partially Pregelatinized Corn Starch (Starch 1500) 20.0 20.0 20.0 Lactose anhydrous (Supertab 24 AN) 59.0 60.0 60.5 Pharmaceutical Grade Fumaric Acid (Powder) 4.0 4.0 4.0 Stearic Acid (Hystrene 5016 Veg) 4.0 4.0 4.0 Total (% w/w) 100.0 100.0 100.0 Total capsule filling weight (mg) 200 200 200

As shown in FIG. 20, in vitro dissolution studies show no effect on dissolution release profiles when varying Aerosil 200 levels from 0.5% w/w, 1% w/w. and 2% w/w, however, the release rates at 75 rpm were faster compared to the release rates at 50 rpm blade speed.

6.13 Production of mannitol and starch-lactose batches 0.1, 0.5 and 2 mg BIC

Theoretical unit formulas for mannitol and starch-lactose 0.1, 0.5 and 2 mg BIC batches are presented in the following table.

Table 15

Theoretical unit formulas for BIC formulations based on 0.1 mg, 0.5 mg and 2 mg mannitol and starch lactose

Manufacturing process Direct mixing with co-grinding 0.1 mg BIC 0.5 mg BIC 2 mg BIC Serial number Cap-37 Cap-41 Cap-42 Cap-43 Cap-45 Cap-46 Ingredients % wt./wt. % wt./wt. % wt./wt. % wt./wt. % wt./wt. % wt./wt. Connection 1 0.133 0.13 0.50 0.50 1.00 1.00 Silica dimethyl silylate (Aerosil R972) 0.50 0.50 Colloidal silicon dioxide (Aerosil 200 Pharma) 1.00 1.00 2.00 1.00 Mannitol (Pearlitol 200 SD) 92.37 91.5 90.00 Partially preliminary gelatinized corn starch (Starch 1500) 20.00 19.50 19.00 Lactose anhydrous (Supertab 24 AN) 61.37 61.00 61.00

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Sodium starch glycolate (Explotab) 10.0 10.0 10.0 Pharmaceutical Grade Fumaric Acid (Powder) 3.00 4.00 3.00 4.00 3.00 4.00 Stearic acid (Hystrene 5016 Veg) 4.00 4.00 4.00 4.00 4.00 4.00 Total (% w/w) 100.0 100.0 100.0 100.0 100.0 100.0 Total capsule filling mass (mg) 75.0 75.0 100.0 100.0 200.0 200.0 HPMC Vcaps Plus capsule shell Size 4 White Opaque V Cap Plus Size 3, yellow, ivory bones opaque V Cap Plus Size 2 Orange Red V Cap Plus

Mannitol-based 0.1 mg BIC manufacturing process

In one example, a 0.1 mg mannitol-based BIC was prepared according to the development process described below.

Preparation of the pre-treatment mixture: Compound 1 tends to stick, therefore, to minimize sticking to equipment surfaces, the pre-treatment mixture was prepared by mixing a portion of mannitol and stearic acid in a Turbula mixer. All equipment used in production has been pre-treated with a pre-treatment mixture.

Mixing 1: A weighed amount of Compound 1 and Aerosil R972 was added to the pre-treated turbule mixer and mixed for 170 revolutions.

Mixing 2: Suspended fumaric acid was added to the Turbula mixer and mixed at 170 rpm.

Blend 3: Mannitol and Blend 2 were added to a 4 L surface treated Turbula mixer and mixed for 340 rpm.

Rub 1: Mixture 3 was passed through a pre-treated Co-mil surface equipped with a 457 µm round screen at 1600 rpm. The Co-mil was washed with a portion of the pretreatment mixture.

Mixing 4: A portion of the pre-treatment mixture, the co-ground mixture and a portion of the mannitol were added to a V-mixer, and the powder was mixed at 300 rpm.

Rub 2: Mix 4 and the remaining pre-treatment mixture were passed through a sifter equipped with a 457 µm round sieve at 1600 rpm. The remaining mannitol was passed through co-mil for washing.

Sifting: The remaining stearic acid was manually sifted through a No. 25 sieve.

Mixing 5 (Lubricant): A mixture of anhydrous 2 and sieved stearic acid was loaded into a V-mixer and mixed at 300 rpm. The final lubricated mixture was discharged into one of two surface-treated collection bags (1 or 2).

Encapsulation: The final mixture was loaded into the hopper of a Bosch encapsulator. The 75 mg mixture was filled into a size #4 HPMC Vcap plus capsule using a size #4 disc at a rate of 75-85 capsules per minute. Average weight and weight change were checked during encapsulation every 15 min, and capsules were collected for stratified CU testing.

Production processes for 0.5 mg and 2 mg mannitol BIC

In one example, 0.5 and 2 mg mannitol-based BICs were prepared according to the development process described below.

Preparation of the pre-treatment mixture: Compound 1 tends to stick, therefore, to minimize sticking to equipment surfaces, the pre-treatment mixture was prepared by mixing a portion of mannitol and stearic acid in a Turbula mixer. All equipment used in production has been pre-treated with a pre-treatment mixture.

Mixing 1: A weighed amount of Compound 1, a portion of mannitol and fumaric acid were added to the pre-treated turbule mixer and mixed for 340 revolutions.

Rub 1: Mixture 1 and a portion of the mannitol were passed through a pre-treated Co-mil surface equipped with a 457 µm round screen at 1600 rpm. The Co-mil was washed with a portion of the pretreatment mixture.

Mixing 2: A portion of the pre-treatment mixture, a mixture from dehydrator 1 and a portion of mannitol were added to the V-mixer, and the powder was mixed for 300 revolutions.

Rub 2: Mixture 2 and a portion of the mannitol were passed through a co-mil equipped with a 457 µm round sieve at 1600 rpm. The remaining mannitol was passed through co-mil for washing.

Sieving: Colloidal silica (Aerosil 200) and stearic acid were manually sifted through a No. 25 sieve.

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Mixing 3 (Lubricant): A mixture of pureed 2 and sieved stearic acid colloidal silica was loaded into a V-mixer and mixed for 300 revolutions.

The final lubricated mixture was discharged into one of two surface-treated collection bags (1 or 2).

Encapsulation: The final mixture was loaded into the hopper of a Bosch encapsulator. For 0.5 mg BIC, the 100 mg mixture was filled into a size #3 HPMC Vcap plus capsule using a size #3 disc at a speed of 75-85 capsules per minute. For 2 mg BIC, the 200 mg mixture was filled into a size #2 HPMC Vcap plus capsule using a size #2 disc at a rate of 75-85 capsules per minute. Average weight and weight change were checked during encapsulation every 15 min, and capsules were collected for stratified CU testing.

Production process of 0.1 mg BIC based on starch and lactose

In one example, 0.1 mg starch-lactose BIC was prepared according to the development process described below.

Preparation of Pretreatment Mixture: Compound 1 tends to stick, therefore, to minimize sticking to equipment surfaces, the pretreatment mixture was prepared by mixing a portion of lactose anhydrous and stearic acid in a Turbula mixer. All equipment used in production has been pre-treated with a pre-treatment mixture.

Mixing 1: A weighed amount of Compound 1 and Aerosil R972 was added to the pre-treated turbule mixer and mixed for 170 revolutions.

Mixing 2: Suspended fumaric acid was added to the Turbula mixer and mixed at 170 rpm.

Mixing 3: A portion of lactose anhydrous, mixture 2, and a portion of the partially pregelatinized corn starch were added to a 4 L surface treated turbula mixer and mixed for 340 revolutions.

Rub 1: Mixture 3 was passed through a pre-treated Co-mil surface equipped with a 457 µm round screen at 1600 rpm. The Co-mil was washed with a portion of the pretreatment mixture.

Mixing 4: A portion of the anhydrous lactose, the co-milled mixture, a portion of the pregelatinized corn starch, and the sodium starch glycolate were added to a V-mixer and the powder was mixed for 300 revolutions.

Rub 2: Mix 4 and the remaining pre-treatment mixture were passed through a sifter equipped with a 457 µm round sieve at 1600 rpm.

Sifting: The remaining stearic acid was manually sifted through a No. 25 sieve.

Mixing 5 (Lubricant): A mixture of anhydrous 2 and sieved stearic acid was loaded into a V-mixer and mixed at 300 rpm. The final lubricated mixture was discharged into one of two surface-treated collection bags (1 or 2).

Encapsulation: The final mixture was loaded into the hopper of a Bosch encapsulator. The 75 mg mixture was filled into a size #4 HPMC Vcap plus capsule using a size #4 disc at a rate of 75-85 capsules per minute. Average weight and weight change were checked during encapsulation every 15 min, and capsules were collected for stratified CU testing.

Starch-lactose production process 0.5 and 2 mg BIC

In one example, 0.5 and 2 mg starch-lactose BICs were prepared according to the development process described below.

Preparation of Pretreatment Mixture: Compound 1 tends to stick, therefore, to minimize sticking to equipment surfaces, the pretreatment mixture was prepared by mixing a portion of lactose anhydrous and stearic acid in a Turbula mixer. All equipment used in production has been pre-treated with a pre-treatment mixture.

Mixing 1: A weighed amount of Compound 1, a portion of anhydrous lactose, a portion of pregelatinized corn starch, and fumaric acid were added to a pretreated Turbula mixer and mixed for 340 revolutions.

Rub 1: Mix 1 and a portion of the pregelatinized corn starch were passed through a pretreated Co-mil surface equipped with a 457 µm round screen at 1600 rpm. The Co-mil was washed with part of the pre-treatment mixture.

Mixing 2: A portion of pre-treatment mixture, dehydration mixture 1 and a portion of anhydrous lactose were added to the V-mixer, and the powder was mixed for 300 revolutions.

Grinding 2: Mixture 2 was passed through a co-mil grinder equipped with a 457 µm round screen at 1600 rpm. The sodium starch glycolate was passed through co-mil for washing.

Sieving: Colloidal silica (Aerosil 200) and stearic acid were manually sifted through a No. 25 sieve.

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Mixing 3 (Lubricant): A mixture of pureed 2 and sieved stearic acid colloidal silica was loaded into a V-mixer and mixed for 300 revolutions.

The final lubricated mixture was discharged into one of two surface-treated collection bags (1 or 2).

Encapsulation: The final mixture was loaded into the hopper of a Bosch encapsulator. For 0.5 mg BIC, the 100 mg mixture was filled into a size #3 HPMC Vcap plus capsule using a size #3 disc at a speed of 75-85 capsules per minute. For 2 mg BIC, the 200 mg mixture was filled into a size #2 HPMC Vcap plus capsule using a size #2 disc at a rate of 75-85 capsules per minute. Average weight and weight change were checked during encapsulation every 15 min, and capsules were collected for stratified CU testing.

6.14 Characteristics of mannitol and starch-lactose batches 0.1, 0.5 and 2 mg BIC

Flowability assessment

A Dietmar-Schulze ring shear tester (RST) was used to evaluate the flowability of the powder mixture. Specimens were prepared using shear chamber No. 1, and standard loads ranging from 400 Pa to 3600 Pa were applied. The flow function coefficient (ffc) values were determined from the resulting principal consolidation stress (σ1) Pa and its corresponding unconfined yield stress ( _fc ) Pa using the following equation:

//с = σΐ/fc

Based on the obtained ffc value, the powder flow behavior can be categorized numerically: ffc <1 (does not flow); 1 <ffc<2 (very cohesive); 2 <ffc<4 (cohesive); 4 <ffc<10 (easy to flow); ffc>10 (free flow).

The flow ranking order for the BIC mixtures analyzed is Cap-37 > Cap41 > Cap-42 > Cap-43 > Cap-45 > Cap-46 > Cap-44. From the flow charts shown in FIG. 21, batches of 0.1 mg mannitol (Cap 37) and starch-lactose BIC (Cap 41) showed better overall fluidity and can be attributed to the low drug loading in the presence of 0.5% Aerosil R972. Batches of 0.5 mg mannitol BIC (Cap 42) and starch-lactose BIC (Cap 43) also showed good flow profiles. 2 mg mannitol BIC (Cap 45) containing 2% Aerosil 200 was found to have acceptable flowability (free flow limit). 2 mg of BIC starch-lactose mixture with 2% Aerosil 200 (Cap44) was found to flow easily.

It can be noted that the flow tendency decreases with increasing levels of Aerosil 200-1% for the 0.5 mg BIC batches, 2% for the mannitol BIC batch and the starch-lactose BIC batch. The flowability of 2 mg starch-lactose mixture BIC (Cap44) was impacted during the encapsulation process where high weight variability was observed. This high mass variability resulted in a wider distribution of activity, as shown in FIG. 22. To overcome this, Aerosil 200 levels were reduced to 1% from previous levels of 2% (Cap44), the flow of a mixture of 2 mg starch lactose (Cap46) BIC with 1% Aerosil 200 was found to flow freely with a tighter activity distribution.

Water Activity Testing

Water activity is a measure of how tightly water is bound to a substance. To measure water activity, the sample is placed in a sealed vessel and allowed to reach equilibrium. The final relative humidity (RH) in the vessel is indicated as a fraction called Aw. Pure water has A _w 1.00. Because Form K of Compound 1 is a channel hydrate that converts to the anhydrous form at <20% relative humidity, water activity was tested to assess the likelihood of form conversion. A HygroLab 3 benchtop indicator was used to determine water activity. Before testing samples, the equipment is calibrated at various relative humidity levels using relative humidity standards (0%, 35% and 80%) and at various temperatures to ensure the accuracy of the sensors. A known amount of sample is sealed inside the sample cell by turning the cap downward. The initial temperature is recorded before the test begins. If the temperature before and after the test changes by no more than 0.5°C, Aw is recorded.

Both the filled capsules and the final mixture show water activity above Aw=0.300 for all formulations. Therefore, at standard room relative humidity of 35-40%, it is unlikely that Compound 1 will become anhydrous.

Dissolution profiles

For the 0.1 mg BIC batches, the mannitol-based BIC exhibited ~95% drug release in 30 min compared to ~80% drug release for the starch-lactose based batch as shown in FIG. 23. The relatively slower drug release for starch-lactose BIC batches is attributed to the formation of a plug that slows drug release. The drug release profile is slower but acceptable for starch-lactose based formulations. Overall, release close to 100% was achieved for both formulations by 60 min.

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At a dosage of 0.5 mg, both formulations have similar dissolution release profiles as shown in FIG. 24. In general, release close to 100% was achieved for both formulations by 60 min.

Dissolution release profile in FIG. 25 for a 2 mg concentration shows that ~90% drug release was observed for the mannitol-based BIC in 30 min, compared to ~75% drug release for the starch lactose-based BIC formulation. However, by 60 min, both BIC formulations demonstrated 100% drug release and exhibited acceptable release profiles upon dissolution.

Resilience Profiles

The stability of BIC mannitol capsules in a HDPE bottle without desiccant was evaluated at 25°C/60% RH and 40°C/75% RH after 1 month, as shown in FIG. 26. 0.1 mg BIC was found to be very stable under all conditions studied. Higher doses (0.5 mg and 2 mg) of BIC were found to be sensitive to hydrolytic degradation at 40°C/75% RH for 1M. The difference between these strengths lies in the grade and levels of silica. For 0.1 mg BIC (Cap-37), Aerosil R972 (hydrophobic silica) coated the API surface well and prevented API degradation, while hydrophilic silica (Aerosil 200) was used for 0.5 mg and 2 mg BIC, respectively. The increase in hydrolytic degradation correlates with an increase in the level of hydrophilic Aerosil 200.

The increase in R-isomer levels for 0.5 mg BIC and 2 mg BIC follows a similar trend to that of increasing hydrolytic degradation levels, indicating the effect of Aerosil 200 on the loss of chiral purity, as shown in FIG. 27.

Dissolution stability

The stable release and dissolution profiles for 0.1 mg mannitol based BIC batches were found to be reproducible under all conditions, whereas the dissolution release profiles for 0.1 mg starch-lactose based BIC batches at 40°C/75% RH at 1M and 25°C/60% RH were slower than at T=0 (initial), as shown in Fig. 28.

From fig. 29 the dissolution release profiles for 0.5 mg BIC for both mannitol and starch-lactose were found to be reproducible. A slight slowdown was observed in the dissolution release profile for 0.5 mg starch-lactose BIC at 40°C/75% RH at T=1M at initial time points, but complete drug release was eventually observed. Overall, both formulations were found to exhibit reproducible dissolution profiles indicating dissolution stability.

Likewise, the dissolution release profiles for 2 mg BIC formulations for both mannitol and starch-lactose formulations were found to be reproducible under all conditions tested (Figure 30).

Particle size distribution for mannitol-based batches

Particle size distribution using the sieving method was done using a Rotap sifter by stacking 6 sieves in decreasing order of sieve opening from top to bottom (larger on top followed by smaller on bottom). A tray was placed at the bottom to collect finely ground material. The experiment was performed in duplicate using a 425 μm screen, a 250 μm screen, a 180 μm screen, a 125 μm screen, a 90 μm screen, and a 75 μm screen. The D10, D50 and D90 of all lubricant mixtures were determined for the 0.1 mg, 0.5 mg and 2 mg mannitol based batches and their values are presented in the following table.

Table 16

Particle size distribution of lubricant mixtures for 0.1, 0.5 and 2 mg mannitol-based batches

Particle size distribution Batch 0.1 mg (µm) Batch 0.5 mg (µm) Batch 2 mg (µM) D10 ~87.5 ~75 ~75 D50 ~137.5 ~131.25 ~143.75 D90 ~193.75 ~181.25 ~206.25

Comparative assessment of content uniformity (CU) between mannitol-based BIC and starch-lactose BIC formulations

The activity distribution, average assay recovery, and average CU for mannitol-based BICs (Cap-37, Cap-42, and Cap-45) and starch-based BICs (Cap-41, Cap-43, and Cap-46) are shown in the following table.

Table 17

Content Uniformity (CU) Summary Data for 0.1, 0.5 and 2 mg BIC Mannitol and Starch-Lactose Formulations

0.1 mg mannitol BIC 0.1 mg starch lactose BIC 0.5 mg mannitol BIC 0.5 mg starch lactose BIC 2 mg mannitol BIC 2 mg starch lactose BIC Serial number Cap-37 Cap-41 Cap-42 Cap-43 Cap-45 Cap-46

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Average CU 96.13 94.16 97.0 99.9 98.89 100.61 % RSD 2.7 1.32 1.3 1.28 0.69 2.0 Acceptability Index (AV) 8.6 7.5 4.6 3.0 1.67 4.86 Analysis mean 99.7 96.1 97.7 99.4 99.0 100.5 Average weight Cap (mg) 113.1 113.3 147.6 147.8 262.4 262.8 Target mass Cap (mg) 111.8 146.9 260.6

Due to the adhesive nature of the mixture, differences in activity distribution were observed for mannitol-based BIC formulations between different formulations made using low or high shear processes. However, pre-treatment of surfaces prior to mass production produced acceptable data with a narrow activity distribution, as shown in the table.

The table shows that not only was the %RSD more stringent for starch-lactose BICs, but also the acceptance rates were lower. Similarly, for 0.5 mg BIC batches, the mean assay recovery, mean CU and AV for the starch-lactose BIC (Cap-43) were higher than the mannitol-based BIC (Cap-42), while the %RSD turned out to be the same for both parties. Finally, the mean assay recovery and mean CU were found to be higher for the starch-lactose-based BIC (Cap-46), but the %RSD and AV were better for the mannitol-based BIC (Cap-45). It was also demonstrated that mean recovery scores improved with increasing dose strength, whereas acceptability scores decreased with increasing dose strength, indicating vulnerability to CU for low doses.

The embodiments presented herein are not intended to be limited in scope to the specific embodiments presented in the examples, which are intended to illustrate several aspects of the embodiments provided, and any implementations that are functionally equivalent are covered by this disclosure. Indeed, various modifications to the embodiments presented herein in addition to those shown and described herein will become apparent to those skilled in the art and are intended to fall within the scope of the appended claims.

A number of references have been cited, the descriptions of which are incorporated herein by reference in their entirety.

Claims (6)

1. Pharmaceutical composition containing: 1) connection 1: or an enantiomer thereof, or a pharmaceutically acceptable salt in an amount of from 0.05 to 3% w/w; And 2) mannitol in an amount from 80 to 98% wt./wt. 2. The pharmaceutical composition according to claim 1, characterized in that compound 1 or its enantiomer, or a pharmaceutically acceptable salt is the free base of compound 1. 3. The pharmaceutical composition according to claim 2, characterized in that the free base of compound 1 is a crystalline free base of compound 1. 4. The pharmaceutical composition according to claim 2, characterized in that the free base of compound 1 is characterized by a pXRD diagram containing peaks at 14.6, 18.2 and 18.3° 2θ ± 0.2° 2θ. 5. Pharmaceutical composition according to any one of claims 1 to 4, characterized in that the amount of compound 1 or its enantiomer, or a pharmaceutically acceptable salt is from 0.1 to 1.5% wt./wt. 6. The pharmaceutical composition according to claim 5, characterized in that the amount of compound 1 or its enantiomer, or a pharmaceutically acceptable salt is from 0.13 to 1% wt./wt. 7. Pharmaceutical composition according to any one of claims 1 to 6, characterized in that the amount of mannitol is from 85 to 95% wt./wt. 8. Pharmaceutical composition according to claim 7, characterized in that the amount of mannitol is from 90 to 93% wt./wt. 9. Pharmaceutical composition according to any one of claims 1 to 8, additionally containing a substance that improves glidability in an amount of from 0 to 10% wt./wt. 10. Pharmaceutical composition according to claim 9, characterized in that the glidant is silica dimethyl silylate or colloidal silicon dioxide. 11. Pharmaceutical composition according to claim 9 or 10, characterized in that the amount of glidant improves from 0.25 to 3% wt./wt. 12. Pharmaceutical composition according to claim 11, characterized in that the amount of glidant improves glidability is from 0.5 to 2% wt./wt. 13. Pharmaceutical composition according to any one of claims 1-12, additionally containing an acidifier in an amount from 0 to 6% wt./wt. 14. Pharmaceutical composition according to claim 13, characterized in that the acidulant is fumaric acid. 15. Pharmaceutical composition according to claim 13 or 14, characterized in that the amount of acidifier is from 2 to 5% wt./wt. 16. Pharmaceutical composition according to claim 15, characterized in that the amount of acidifier is 3% wt./wt. 17. Pharmaceutical composition according to any one of claims 1-16, additionally containing a lubricant in an amount of from 0 to 8% wt./wt. 18. Pharmaceutical composition according to claim 17, characterized in that the lubricant is stearic acid. 19. Pharmaceutical composition according to claim 17 or 18, characterized in that the amount of lubricant is from 2 to 6% wt./wt. 20. Pharmaceutical composition according to claim 19, characterized in that the amount of lubricant is 4% wt./wt. 21. Pharmaceutical composition according to claim 1, containing: 1) compound 1 in an amount of 0.13% w/w; 2) mannitol in an amount of 92.37% w/w; 3) silica dimethyl silylate in an amount of 0.5% w/w; 4) fumaric acid in an amount of 3% w/w; And 5) stearic acid in an amount of 4% wt./wt. 22. Pharmaceutical composition according to claim 21, having a total weight of 75 mg. 23. The pharmaceutical composition according to claim 22, which is contained in a size 4 capsule. 24. Pharmaceutical composition according to claim 1, containing: - 62 046729 1) compound 1 in an amount of 0.5% w/w; 2) mannitol in an amount of 91.5% w/w; 3) colloidal silicon dioxide in an amount of 1% w/w; 4) fumaric acid in an amount of 3% w/w; And 5) stearic acid in an amount of 4% wt./wt. 25. Pharmaceutical composition according to claim 24, having a total weight of 100 mg. 26. Pharmaceutical composition according to claim 25, which is contained in a capsule size 3. 27. Pharmaceutical composition according to claim 1, containing: 1) compound 1 in an amount of 1% w/w; 2) mannitol in an amount of 90% wt./wt. 3) colloidal silicon dioxide in an amount of 2% w/w; 4) fumaric acid in an amount of 3% w/w; And 5) stearic acid in an amount of 4% wt./wt. 28. Pharmaceutical composition according to claim 27, having a total weight of 200 mg. 29. The pharmaceutical composition according to claim 28, which is contained in a size 2 capsule. 30. Pharmaceutical composition containing: 1) connection 1: or an enantiomer thereof, or a pharmaceutically acceptable salt in an amount of from 0.05 to 3% w/w; And 2) a mixture of starch and lactose in an amount from 70 to 90% wt./wt. 31. The pharmaceutical composition according to claim 30, characterized in that compound 1 or its enantiomer, or a pharmaceutically acceptable salt is the free base of compound 1. 32. The pharmaceutical composition according to claim 31, characterized in that the free base of compound 1 is a crystalline free base of compound 1. 33. The pharmaceutical composition according to claim 31, characterized in that the free base of compound 1 is characterized by a pXRD diagram containing peaks at 14.6, 18.2 and 18.3° 2θ ±0.2° 2θ. 34. Pharmaceutical composition according to any one of claims 30-33, characterized in that the amount of compound 1 or its enantiomer, or a pharmaceutically acceptable salt is from 0.1 to 1.5% wt./wt. 35. The pharmaceutical composition according to claim 34, characterized in that the amount of compound 1 or its enantiomer, or a pharmaceutically acceptable salt is from 0.13 to 1% wt./wt. 36. Pharmaceutical composition according to any one of claims 30-35, characterized in that the starch is partially pregelatinized starch. 37. Pharmaceutical composition according to any one of claims 30-36, characterized in that the lactose is anhydrous lactose. 38. Pharmaceutical composition according to any one of claims 30-37, characterized in that the amount of starch is from 17 to 22% w/w, and the amount of lactose is from 53 to 68% w/w. 39. Pharmaceutical composition according to any one of claims 30-37, characterized in that the amount of the mixture of starch and lactose is from 75 to 85% wt./wt. 40. The pharmaceutical composition according to claim 39, characterized in that the amount of starch is from 18 to 21% wt./wt., and the amount of lactose is from 57 to 64% wt./wt. 41. Pharmaceutical composition according to any one of claims 30-37, characterized in that the amount of the mixture of starch and lactose is from 80 to 82% wt./wt. 42. The pharmaceutical composition according to claim 41, characterized in that the amount of starch is from 19 to 20% w/w, and the amount of lactose is from 61 to 62% w/w. 43. Pharmaceutical composition according to any one of claims 30-37, characterized in that the mass ratio of starch to lactose is from 1:2 to 1:4. 44. Pharmaceutical composition according to claim 43, characterized in that the mass ratio of starch to lactose is from 1:3 to 1:3.3. 45. Pharmaceutical composition according to any one of claims 30-44, additionally containing a substance that improves glidability in an amount of from 0 to 10% wt./wt. 46. The pharmaceutical composition according to claim 45, characterized in that the glidant is silica dimethyl silylate or colloidal silicon dioxide. 47. Pharmaceutical composition according to claim 45 or 46, characterized in that the amount of substance, - 63 046729 improving slip, ranges from 0.25 to 3% w/w. 48. The pharmaceutical composition according to claim 47, characterized in that the amount of the glidant agent is from 0.5 to 1% wt./wt. 49. Pharmaceutical composition according to any one of claims 30-48, additionally containing an acidifier in an amount from 0 to 8% wt./wt. 50. Pharmaceutical composition according to claim 49, characterized in that the acidulant is fumaric acid. 51. Pharmaceutical composition according to claim 49 or 50, characterized in that the amount of acidifier is from 2 to 6% wt./wt. 52. The pharmaceutical composition according to claim 51, characterized in that the amount of acidifier is 4% wt./wt. 53. Pharmaceutical composition according to any one of claims 30-52, additionally containing a lubricant in an amount of from 0 to 8% wt./wt. 54. Pharmaceutical composition according to claim 53, characterized in that the lubricant is stearic acid. 55. Pharmaceutical composition according to claim 53 or 54, characterized in that the amount of lubricant is from 2 to 6% wt./wt. 56. Pharmaceutical composition according to claim 55, characterized in that the amount of lubricant is 4% wt./wt. 57. Pharmaceutical composition according to any one of claims 30-56, additionally containing a disintegrant in an amount from 0 to 20% wt./wt. 58. Pharmaceutical composition according to claim 57, characterized in that the disintegrant is sodium starch glycolate. 59. Pharmaceutical composition according to claim 57 or 58, characterized in that the amount of disintegrant is from 5 to 15% wt./wt. 60. Pharmaceutical composition according to claim 59, characterized in that the amount of disintegrant is 10% wt./wt. 61. Pharmaceutical composition according to claim 30, containing: 1) compound 1 in an amount of 0.13% w/w; 2) partially pregelatinized starch in an amount of 20% wt./wt. and anhydrous lactose in an amount of 61.4% w/w; 3) silica dimethyl silylate in an amount of 0.5% w/w; 4) fumaric acid in an amount of 4% w/w; 5) stearic acid in an amount of 4% w/w; And 6) sodium starch glycolate in an amount of 10% wt./wt. 62. Pharmaceutical composition according to claim 61, having a total weight of 75 mg. 63. The pharmaceutical composition according to claim 62, which is contained in a size 4 capsule. 64. Pharmaceutical composition according to claim 30, containing: 1) compound 1 in an amount of 0.5% w/w; 2) partially pregelatinized starch in an amount of 19.5% wt./wt. and anhydrous lactose in an amount of 61% w/w; 3) colloidal silicon dioxide in an amount of 1% w/w; 4) fumaric acid in an amount of 4% w/w; 5) stearic acid in an amount of 4% w/w; And 6) sodium starch glycolate in an amount of 10% wt./wt. 65. Pharmaceutical composition according to claim 64, having a total weight of 100 mg. 66. The pharmaceutical composition according to claim 65, which is contained in a size 3 capsule. 67. Pharmaceutical composition according to claim 30, containing: 1) compound 1 in an amount of 1% w/w; 2) partially pregelatinized starch in an amount of 19% wt./wt. and anhydrous lactose in an amount of 61% w/w; 3) colloidal silicon dioxide in an amount of 1% w/w; 4) fumaric acid in an amount of 4% w/w; 5) stearic acid in an amount of 4% w/w; And 6) sodium starch glycolate in an amount of 10% wt./wt. 68. Pharmaceutical composition according to claim 67, having a total weight of 200 mg. 69. The pharmaceutical composition according to claim 68, which is contained in a size 2 capsule. 70. A method of treating multiple myeloma, comprising administering a therapeutically effective amount of a pharmaceutical composition according to any one of claims 1 to 69 to a patient in need thereof. 71. A method for preparing a pharmaceutical composition according to any one of claims 1 to 69, comprising coating compound 1 or its enantiomer, or a pharmaceutically acceptable salt with hydrophobic silica. - 64 046729 72. The method of claim 71, wherein the coating is carried out by mixing compound 1 or its enantiomer, or a pharmaceutically acceptable salt, and hydrophobic silica to form a binary mixture before mixing with additional excipients. 73. The method according to claim 71 or 72, characterized in that the hydrophobic silica is silica dimethyl silylate.