Crystalline compositions for improving embryo implantation and methods of using the same

Abstract

The disclosure provides crystalline forms of (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1,1'-biphenyl-4-5 yl)carbonyl]pyrrolidin-3-one O-methyloxime, as well as methods of using this crystalline material to improve embryo implantation outcomes in patients undergoing an assisted reproduction technology (ART) procedure, such as in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI).

Description

[0001] CRYSTALLINE COMPOSITIONS FOR IMPROVING EMBRYO IMPLANTATION AND METHODS OF USING THE SAME

[0002] Field of the Invention

[0003] The disclosure relates to the field of assisted reproduction technology and provides compositions and methods for treating subjects undergoing embryo transfer procedures so as to enhance, for example, endometrial receptivity and reduce the likelihood of embryo implantation failure.

[0004] Background of the Invention

[0005] Despite recent progress in assisted reproductive technology, the overall effectiveness of even advanced treatments, such as in vitro fertilization (IVF) followed by embryo transfer (IVF-ET), remains relatively low, resulting in an average of about 30% live births per treatment cycle (Andersen et al., Human Reproduction 24:1267-1287 (2009)). Moreover, embryo implantation success rate tends to decrease with age. Many current treatment strategies to promote successful embryo implantation in a subject undergoing embryo transfer therapy have focused on the inhibition of uterine contractions prior to embryo transfer. Such treatment modalities include the administration of p-adrenergic receptor agonists and non-steroidal anti-inflammatory drugs (NSAIDS), which have not been shown to provide sufficient clinical benefit (Bernabeu et al., Human Reproduction 21 :364-368 (2006); Moon et al., Fertility and Sterility 82:816-820 (2004); and Tsirigotis et al., Human Reproduction 15:10 (2000)). There remains a need for improved compositions and methods for promoting successful embryo implantation, for instance, by enhancing endometrial receptivity upon embryo transfer in patients undergoing assisted reproductive technology procedures.

[0006] Summary of the Invention

[0007] The present disclosure provides compositions and methods for treating subjects (e.g., female human subjects) undergoing embryo transfer procedures, such as in vitro fertilization (IVF), among others described herein. Particularly, the disclosure features crystalline forms of an oxytocin receptor antagonist, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O- methyloxime, as well as methods of administering the same to a subject undergoing an embryo transfer procedure. Using the compositions and methods described herein, administration of the oxytocin receptor antagonist to such a subject may improve the subject’s likelihood of successful embryo implantation, as well as reduce the probability that the subject will experience a miscarriage following embryo transfer. These beneficial results can help a patient undergoing embryo transfer therapy to successfully establish pregnancy and to remain pregnant until delivery following a full gestational period.

[0008] Examples of beneficial responses to oxytocin receptor antagonist treatment that a subject may exhibit include reduced uterine contractility and augmented blood flow to the endometrium. Together, uterine contractility and endometrial blood flow constitute important components of endometrial receptivity to a transferred embryo. Without being limited by mechanism, an oxytocin receptor antagonist administered to a subject using the compositions and methods described herein may enhance uterine perfusion and suppress uterine contractions that could otherwise lead to embryo expulsion. These effects can collectively serve to create an environment within the endometrium that is conducive to successful embryo implantation, which can ultimately enhance the likelihood of achieving and sustaining a successful pregnancy.

[0009] In an aspect, the disclosure features a crystalline form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) wherein the crystalline form of the compound exhibits characteristic X-ray powder diffraction (XRPD) peaks at 12.86 ± 0.20° 26, 16.32 ± 0.20° 26, and 22.69 ± 6.26° 26.

[0010] In some embodiments, the crystalline form of said compound exhibits characteristic XRPD peaks at 17.81 ± 6.26° 26, 21.69 ± 6.26° 26, and 24.86 ± 6.26° 26. In some embodiments, the crystalline form of said compound exhibits characteristic XRPD peaks at 23.94 ± 6.26° 26 and 27.93 ± 6.26° 26. In some embodiments, the crystalline form of said compound exhibits characteristic XRPD peaks at 25.87 ± 6.26° 26, 26.99 ± 6.26° 26, and 28.74 ± 6.26° 26. In some embodiments, the crystalline form of said compound exhibits characteristic XRPD peaks at 7.67 ± 6.26° 26, 12.27 ± 6.26° 26, 14.17 ± 6.26° 26, and 18.79 ± 6.26° 26. In some embodiments, the crystalline form of said compound exhibits characteristic XRPD peaks at 21.34 ± 6.26° 26 and 25.72 ± 6.26° 26. In some embodiments, the crystalline form of said compound exhibits characteristic XRPD peaks at 36.56 ± 6.26° 26, 31 .22 ± 6.26° 26, 32.56 ± 6.26° 26, 33.63 ± 6.26° 26, 33.35 ± 6.26° 26, 33.79 ± 6.26° 26, 34.67 ± 6.26° 26, 34.96 ± 6.26° 26, 35.67 ± 6.26° 26, 36.68 ± 6.26° 26, 37.42 ± 6.26° 26, 37.84 ± 6.26° 26, 38.25 ± 6.26° 26, and 39.72 ± 6.26° 26.

[0011] In some embodiments, the crystalline form of said compound has the XRPD spectrum as shown in FIG. 5A. In some embodiments, the XRPD peaks at diffraction angle 26 (°) are collected by irradiating with Cu Ka.

[0012] In some embodiments, the crystalline form of said compound has a differential scanning calorimetry (DSC) endotherm at about 99 °C. In some embodiments, the crystalline form of said compound has the DSC curve substantially as depicted in FIG. 5F.

[0013] In some embodiments, the crystalline form of said compound exhibits1H nuclear magnetic resonance (NMR) peaks centered at a chemical shift (6) of about 1 .6 ppm, about 2.3 ppm, about 2.5 ppm, about 3.3 ppm, about 3.5 ppm, about 3.7 ppm, about 7.2 ppm, about 7.3 ppm, about 7.4 ppm, and about 7.6 ppm. In some embodiments, the crystalline form is characterized by a1H NMR spectrum substantially as depicted in FIG. 5B.

[0014] In some embodiments, the crystalline form exhibits a weight loss of from 0.01% to 5% when heated from 36 °C to 160 °C as measured by thermogravimetric analysis (TGA). In some embodiments, the crystalline form exhibits a weight loss of about 0.01% to 0.05% when heated from 36 °C to 160 °C as measured by TGA. In some embodiments, the crystalline form exhibits a weight loss of about 0.03% when heated from 36 °C to 160 °C as measured by TGA. In some embodiments, the crystalline form exhibits a TGA curve substantially as depicted in FIG. 5G.

[0015] In some embodiments, the crystalline form exhibits a weight gain of from 0.01% to 5% when the relative humidity is increased from 5% to 95% as measured by dynamic vapor sorption (DVS). In some embodiments, the crystalline form exhibits a weight gain of from 1% to 5% when the relative humidity is increased from 5% to 95% as measured by DVS. In some embodiments, the crystalline form exhibits a weight gain of about 1 .65% when the relative humidity is increased from 5% to 95% as measured by DVS. In some embodiments, the crystalline form exhibits a weight loss of from 0.01% to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS. In some embodiments, the crystalline form exhibits a weight loss of from 1% to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS. In some embodiments, the crystalline form exhibits a weight loss of about 1 .67% when the relative humidity is decreased from 95% to 5% as measured by DVS. In some embodiments, the crystalline form exhibits a DVS curve substantially as depicted in FIG. 5H.

[0016] In some embodiments, the crystalline form exhibits a Raman curve substantially as depicted in FIG. 5I or FIG. 5J.

[0017] In some embodiments, the crystalline form has a solubility of from 0.100 mg / mL to 0.200 mg / mL in water after 24 hours of stirring. In some embodiments, the crystalline form has a solubility of about 0.189 mg / mL in water after 24 hours of stirring.

[0018] In some embodiments, the crystalline form has an intrinsic dissolution rate (IDR) of from 0.010 mg / cm2 / min to 0.020 mg / cm2 / min in water. In some embodiments, the crystalline form has an IDR of about 0.013 mg / cm2 / min in water.

[0019] In some embodiments, the crystalline form is obtainable by recrystallization from a solvent. In some embodiments, the crystalline form is obtainable by recrystallization from ethanol, methanol, or water.

[0020] In another aspect, the disclosure provides a crystalline form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by form wherein the crystalline form exhibits a weight loss of from 0.01% to 5% when heated from 36 °C to 160 °C as measured by TGA, optionally wherein the weight loss is from 0.01% to 0.05%, further optionally wherein the weight loss is about 0.03%.

[0021] In another aspect, the disclosure provides a crystalline form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) having (a) the XRPD spectrum as shown in FIG. 5A and optionally at least one of the following properties:

[0022] (b) a DSC endotherm at about 99 °C;

[0023] (c) a DSC substantially as depicted in FIG. 5F;

[0024] (d)1H NMR peaks centered at a chemical shift (6) of about 1 .0 ppm, about 2.3 ppm, about 2.5 ppm, about 3.3 ppm, about 3.5 ppm, about 3.7 ppm, about 7.2 ppm, about 7.3 ppm, about 7.4 ppm, and about 7.6 ppm;

[0025] (e) a1H NMR spectrum substantially as depicted in FIG. 5B;

[0026] (f) a weight loss of from 0.01% to 5% when heated from 36 °C to 160 °C as measured by TGA, optionally wherein the weight loss is from 0.01 % to 0.05%, further optionally wherein the weight loss is about 0.03%;

[0027] (g) a TGA curve substantially as depicted in FIG. 5G;

[0028] (h) a weight gain of from 0.01% to 5% when the relative humidity is increased from 5% to 95% as measured by DVS, optionally wherein the weight gain is from 1% to 5%, further optionally wherein the weight gain is about 1 .65%;

[0029] (i) a weight loss of from 0.01% to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS, optionally wherein the weight loss is from 1% to 5%, further optionally wherein the weight loss is about 1 .67%; (j) a DVS curve substantially as depicted in FIG. 5H;

[0030] (k) a Raman curve substantially as depicted in FIG. 51 or FIG. 5J;

[0031] (l) a solubility of from 0.100 mg / mL to 0.200 mg / mL in water after 24 hours of stirring;

[0032] (m) an IDR of from 0.010 mg / cm2 / min to 0.020 mg / cm2 / min in water; or

[0033] (n) combinations thereof.

[0034] In another aspect, the disclosure provides a crystalline form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by form characterized by unit cell parameters substantially equal to the following cell dimensions: a = 14.410 A; b = 14.410 A; c = 8.247 A; a = 90 degrees; p = 90 degrees; y = 120 degrees; and

[0035] Space group = P3i.

[0036] In another aspect, the disclosure provides a solid form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by form comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 80% (e.g., about 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0037] In another aspect, the disclosure provides a solid form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by form comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 84% (e.g., about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0038] In another aspect, the disclosure provides a solid form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by form comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 87% (e.g., about 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0039] In another aspect, the disclosure provides a solid form of a compound, (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by form comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 89% (e.g., about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0040] In a further aspect, the disclosure features a pharmaceutical composition containing the crystalline form of any one of the preceding aspects or embodiments of the disclosure, in combination with one or more carriers, diluents, or excipients.

[0041] In another aspect, the disclosure features a method of treating a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form or pharmaceutical composition of any one of the preceding aspects or embodiments of the disclosure.

[0042] In a further aspect, the disclosure features a method of reducing the likelihood of embryo implantation failure in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form or pharmaceutical composition of any one of the preceding aspects or embodiments of the disclosure.

[0043] In a further aspect, the disclosure features a method of improving endometrial receptivity in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form or pharmaceutical composition of any one of the preceding aspects or embodiments of the disclosure.

[0044] In a further aspect, the disclosure features a method of reducing uterine contractility in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form or pharmaceutical composition of any one of the preceding aspects or embodiments of the disclosure.

[0045] In some embodiments, the method comprises transferring the one or more embryos to the uterus of the subject. In some embodiments, the method further comprises inducing follicular maturation in the subject, optionally wherein the inducing of follicular maturation comprises administering human chorionic gonadotropin (hCG) to the subject.

[0046] In some embodiments, from 1 to 2 embryos are transferred to the subject. For example, in some embodiments, one embryo is transferred to the subject. In some embodiments, two embryos are transferred to the subject.

[0047] In some embodiments, the subject is a mammal and the one or more embryos are mammalian embryos. In some embodiments, the mammal is a human and the one or more mammalian embryos are human embryos.

[0048] In some embodiments, the one or more embryos are produced ex vivo by in vitro fertilization (IVF). In some embodiments, the one or more embryos are produced ex vivo by IVF of one or more ova derived from the subject.

[0049] In some embodiments, the one or more embryos are produced ex vivo by intracytoplasmic sperm injection (ICSI). In some embodiments, the one or more embryos are produced ex vivo by ICSI into one or more ova derived from the subject.

[0050] In some embodiments, the one or more ova are derived from one or more oocytes isolated from the subject. In some embodiments, the one or more oocytes comprise from 1 to 4 mature oocytes. In some embodiments, a gonadotropin-releasing hormone (GnRH) antagonist is administered to the subject prior to isolation of the one or more oocytes from the subject. In some embodiments, hCG is administered to the subject prior to isolation of the one or more oocytes from the subject. In some embodiments, progesterone is administered to the subject following isolation of the one or more oocytes from the subject.

[0051] In some embodiments, the one or more ova are isolated directly from the subject. In some embodiments, a GnRH antagonist is administered to the subject prior to isolation of the one or more ova from the subject. In some embodiments, hCG is administered to the subject prior to isolation of the one or more ova from the subject. In some embodiments, progesterone is administered to the subject following isolation of the one or more ova from the subject.

[0052] In some embodiments, the one or more embryos each comprise from 6 to 8 blastomeres immediately prior to the transfer of the one or more embryos to the subject. In some embodiments, the blastomeres are of approximately equal sizes as assessed by visual microscopy.

[0053] In a further aspect, the disclosure features a kit comprising the crystalline form or pharmaceutical composition of any of the foregoing aspects or embodiments of the disclosure, as well as a package insert instructing a user of the kit to administer the crystalline form to a subject in accordance with any one or more of the preceding methods.

[0054] In some embodiments of any of the preceding compositions or methods, the compound represented by formula (I) (i.e., (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime) is substantially pure. For instance, in some embodiments, the compound represented by formula (I) has a purity of at least 85%, such as a purity of from 85% to 99.9% or more (e.g., a purity of 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or more). The purity of the compound represented by formula (I) may be assessed, for instance, using NMR techniques and / or chromatographic methods, such as HPLC procedures, that are known in the art and described herein.

[0055] In some embodiments, the compound represented by formula (I) is substantially pure with respect to diastereomers of this compound and other by-products that may be formed during the synthesis of this compound. For instance, in some embodiments, the compound represented by formula (I) has a purity of at least 85%, such as a purity of from 85% to 99.9% or more (e.g., a purity of 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or more) with respect to diastereomers of this compound and other by-products that may be formed during the synthesis of this compound, such as a by-product that is formed during the synthesis of this compound. The purity of the compound represented by formula (I) may be assessed, for instance, using NMR techniques and / or chromatographic methods, such as HPLC procedures, that are known in the art and described herein.

[0056] In some embodiments, the compound represented by formula (I) is substantially pure with respect to its (3E) diastereomer, (3E,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime. For instance, in some embodiments, the compound represented by formula (I) has a purity of at least 85%, such as a purity of from 85% to 99.9% or more (e.g., a purity of 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or more) with respect to (3E,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime. For instance, compound (I) may be administered in the form of a composition (e.g., a tablet, such as a dispersible tablet, capsule, gel cap, powder, liquid solution, or liquid suspension) that contains less than 15% of the (3E) diastereomer. For example, compound (I) may be administered in the form of a composition (e.g., a tablet, such as a dispersible tablet, capsule, gel cap, powder, liquid solution, liquid suspension, granulate, microemulsion, or self-emulsifying dispersed system) that contains less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.1%, less than 0.01%, less than 0.001%, or less of the (3E) diastereomer. The purity of the compound represented by formula (I) may be assessed, for instance, using NMR techniques and / or chromatographic methods, such as HPLC procedures, that are known in the art and described herein.

[0057] In some embodiments of any of the above aspects of the disclosure, the subject is a human female subject, such as a human female subject of up to 44 years of age, such as a human female subject of from 18 to 44 years of age, such as a human female subject of 18 years, 19 years, 20 years, 21 years, 22 years, 23 years, 24 years, 25 years, 26 years, 27 years, 28 years, 29 years, 30 years, 31 years, 32 years, 33 years, 34 years, 35 years, 36 years, 37 years, 38 years, 39 years, 40 years, 41 years, 42 years, 43 years, or 44 years of age. In some embodiments of any of the above aspects of the disclosure, the subject is a human female subject of up to 42 years of age, such as a human female subject of from 18 to 42 years of age, such as a human female subject of 18 years, 19 years, 20 years, 21 years, 22 years, 23 years, 24 years, 25 years, 26 years, 27 years, 28 years, 29 years, 30 years, 31 years, 32 years, 33 years, 34 years, 35 years, 36 years, 37 years, 38 years, 39 years, 40 years, 41 years, or 42 years of age. In some embodiments of any of the above aspects of the disclosure, the subject is a human female subject of up to 36 years of age, such as a human female subject of from 18 to 36 years of age, such as a female subject of 18 years, 19 years, 20 years, 21 years, 22 years, 23 years, 24 years, 25 years, 26 years, 27 years, 28 years, 29 years, 30 years, 31 years, 32 years, 33 years, 34 years, 35 years, or 36 years of age. Definitions

[0058] As used herein, the term “about” refers to a value that is within 10% above or below the value being described. For instance, the phrase “about 50 mg” refers to a value between and including 45 mg and 55 mg.

[0059] As used herein, the term “affinity” refers to the strength of a binding interaction between two molecules, such as a ligand and a receptor. The term "Ki", as used herein, is intended to refer to the inhibition constant of an antagonist for a particular molecule of interest, and can be expressed as a molar concentration (M). Ki values for antagonist-target interactions can be determined, e.g., using methods established in the art. Methods that can be used to determine the Ki of an antagonist for a molecular target include competitive binding experiments, such as competitive radioligand binding assays, for instance, as described in US Patent No. 9,670,155, the disclosure of which is incorporated herein by reference in its entirety. The term "Kd", as used herein, is intended to refer to the dissociation constant, which can be obtained, for example, from the ratio of the rate constant for the dissociation of the two molecules (kd) to the rate constant for the association of the two molecules (ka) and is expressed as a molar concentration (M). Kd values for receptor-ligand interactions can be determined, e.g., using methods established in the art. Methods that can be used to determine the Kd of a receptor-ligand interaction include surface plasmon resonance, e.g., through the use of a biosensor system such as a BIACORE® system.

[0060] As used herein, the term “assisted reproductive technology” or “ART” refers to a fertility treatment in which one or more female gametes (ova) and male gametes (sperm cells) are manipulated ex vivo so as to promote ovum fertilization and formation of a zygote or embryo. The zygote or embryo is then transferred to the uterus of a female subject, for instance, using the compositions and methods described herein. Exemplary assisted reproductive technology procedures include in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) techniques described herein and known in the art.

[0061] As used herein, the terms “benefit” and “response” in the context of a subject undergoing embryo transfer therapy are used interchangeably and refer to any clinical improvement in the subject’s condition or ability to undergo successful embryo implantation and development. Exemplary “benefits” (“responses”) in this context, such as in the context of a subject treated with an oxytocin receptor antagonist prior to, concurrently with, and / or after the transfer of one or more embryos to the subject, include, without limitation, an increase in the subject’s endometrial perfusion, a reduction in the subject’s uterine contractility, increase in the subject’s endometrial receptivity toward a transferred embryo, a reduction in the likelihood of embryo implantation failure, the prevention of a miscarriage, and the achievement and maintenance of a successful pregnancy, for example, until delivery at a full gestational age, in a subject following transfer of one or more embryos to the subject. A subject can be determined to benefit, for instance, from oxytocin receptor antagonist treatment as described herein by observing an elevated endometrial receptivity in the subject (for instance, as assessed by detecting a reduction in prostaglandin F2a (PGF2a) signal transduction as described herein and / or by assessing the subject’s ability to sustain a pregnancy for at least 14 days, 6 weeks, 10 weeks, or more, following the transfer of one or more embryos to the subject and / or following the retrieval of one or more oocytes or ova from the subject, and / or by detecting the ability of the subject to give birth to a live offspring at least 24 weeks following the transfer of one or more embryos to the subject. Additionally or alternatively, a subject can be determined to benefit from oxytocin receptor antagonist treatment as described herein by monitoring the subject for a miscarriage following the transfer of one or more embryos to the subject and observing that the subject has not undergone a miscarriage.

[0062] As used herein, the term “controlled ovarian hyperstimulation” refers to a procedure in which ovulation is induced in a subject, such as a human subject, prior to oocyte or ovum retrieval for use in embryo formation, for instance, by in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). Controlled ovarian hyperstimulation procedures may involve administration of human chorionic gonadotropin (hCG) and / or a gonadotropin-releasing hormone (GnRH) antagonist to the subject so as to promote “follicular maturation,” a term that refers to the development of an oocyte into a “mature oocyte,” as defined herein. “Final follicular maturation” refers to the last administration of an agent that promotes follicular maturation to a subject prior to oocyte retrieval. Controlled ovarian hyperstimulation methods are known in the art and are described, for instance, in US Patent Nos. 7,405,197 and 7,815,912, the disclosures of each of which are incorporated herein by reference as they pertain to methods for inducing follicular maturation and ovulation in conjunction with assisted reproductive technology.

[0063] As used herein, the term “crystalline” or “crystalline form” means having a physical state that is a regular three-dimensional array of atoms, ions, molecules or molecular assemblies. Crystalline forms have lattice arrays of building blocks called asymmetric units that are arranged according to well-defined symmetries into unit cells that are repeated in three-dimensions. In contrast, the term “amorphous” or “amorphous form” refers to an unorganized (no orderly) structure. The physical state of a therapeutic compound may be determined by exemplary techniques such as x-ray diffraction, polarized light microscopy, thermal gravimetric analysis, and / or differential scanning calorimetry.

[0064] As used herein, the term “derived from” in the context of a cell derived from a subject refers to a cell, such as a mammalian ovum, that is either isolated from the subject or obtained from expansion, division, maturation, or manipulation (e.g., ex vivo expansion, division, maturation, or manipulation) of one or more cells isolated from the subject. For instance, an ovum is “derived from” a subject or an oocyte as described herein if the ovum is directly isolated from the subject or obtained from the maturation of an oocyte isolated from the subject, such as an oocyte isolated from the subject from about 1 day to about 7 days prior to the subject undergoing an embryo transfer procedure (e.g., an oocyte isolated from the subject from about 3 days to about 5 days prior to the subject undergoing an embryo transfer procedure).

[0065] As used herein, the term "dispersible tablet" refers to a tablet capable of rapidly disintegrating in water and that is swallowed by a subject, or that is intended to be disintegrated rapidly in water and subsequently swallowed by a subject, such as a subject undergoing embryo transfer therapy as described herein. As used herein, the term “dose” refers to a quantity of a therapeutic agent, such as an oxytocin receptor antagonist described herein, that is administered to a subject at a particular point in time for the treatment of a disorder or condition, such as to enhance endometrial receptivity and promote successful embryo implantation in the context of assisted reproductive technology. A therapeutic agent as described herein may be administered in a single dose or in multiple doses. In each case, the therapeutic agent may be administered using one or more unit dosage forms of the therapeutic agent. For instance, a single dose of 100 mg of a therapeutic agent may be administered using, e.g., two 50 mg unit dosage forms of the therapeutic agent. Similarly, a single dose of 300 mg of a therapeutic agent may be administered using, e.g., six 50 mg unit dosage forms of the therapeutic agent or two 50 mg unit dosage forms of the therapeutic agent and one 200 mg unit dosage form of the therapeutic agent, among other combinations. Similarly, a single dose of 900 mg of a therapeutic agent may be administered using, e.g., six 50 mg unit dosage forms of the therapeutic agent and three 200 mg unit dosage forms of the therapeutic agent or ten 50 mg unit dosage form of the therapeutic agent and two 200 mg unit dosage forms of the therapeutic agent, among other combinations. Similarly, a single dose of 400 mg of the therapeutic agent may be administer using, e.g., one 400 mg unit dosage form of the therapeutic agent.

[0066] As used herein, the term “embryo” refers to a multicellular, post-zygotic derivative of a fertilized ovum. An embryo may contain two or more blastomeres. For instance, embryos for use with the compositions and methods of the disclosure include those that contain from 6 to 8 blastomeres. Embryos may be produced ex vivo, for instance, by in vitro fertilization (IVF) of an ovum, such as an ovum isolated from a subject undergoing embryo transfer therapy or from a donor, or an ovum produced by maturation of an oocyte isolated from a subject undergoing embryo transfer therapy or from a donor. Embryos may be produced ex vivo, for instance, by intracytoplasmic sperm injection (ICS I) of an ovum, such as an ovum isolated from a subject undergoing embryo transfer therapy or from a donor, or an ovum produced by maturation of an oocyte isolated from a subject undergoing embryo transfer therapy or from a donor. An embryo may have a variety of multicellular forms resulting from ovum fertilization and mitosis of the ensuing zygote. For instance, an embryo may have the form of a morula, which is typically formed from about 3 days to about 4 days following ovum fertilization, and contains two or more cells (such as from 2 to 16 cells, for instance, from 6 to 8 cells) packed contiguously in a spherical arrangement. An embryo may have the form of a blastula (e.g., a mammalian blastocyst), which is typically formed from about 5 days to about 7 days following ovum fertilization, characterized by a spherical morphology containing an outer lining of cells (e.g., a mammalian trophoblast or trophectoderm) surrounding an inner cell mass and a fluid-filled cavity (e.g., a mammalian blastocoele). A blastocyst may contain, for instance, from about 20 to about 300 cells (e.g., about 20 cells, 25 cells, 30 cells, 35 cells, 40 cells, 45 cells, 50 cells, 55 cells, 60 cells, 65 cells, 70 cells, 75 cells, 80 cells, 85 cells, 90 cells, 95 cells, 100 cells, 105 cells, 110 cells, 115 cells, 120 cells, 125 cells, 130 cells, 135 cells, 140 cells, 145 cells, 150 cells, 155 cells, 160 cells, 165 cells, 170 cells, 175 cells, 180 cells, 185 cells, 190 cells, 195 cells, 200 cells, 205 cells, 210 cells, 215 cells, 220 cells, 225 cells, 230 cells, 235 cells, 240 cells, 245 cells, 250 cells, 255 cells, 260 cells, 265 cells, 270 cells, 275 cells, 280 cells, 285 cells, 290 cells, 295 cells, or 300 cells) or more.

[0067] As used herein, the terms “embryo transfer therapy” and “embryo transfer procedure” are used interchangeably and refer to a procedure in which one or more embryos are transferred to the uterus of a subject, such as a mammalian subject (e.g., a human subject) so as to promote implantation of the one or more embryos into the endometrium of the subject, thereby establishing pregnancy. The embryo may be produced ex vivo, for instance, by in vitro fertilization (IVF) or by intracytoplasmic sperm injection (ICSI), optionally using one or more ova derived from the subject (e.g., one or more ova obtained from maturation of one or more oocytes isolated from the subject) or using one or more ova derived from a donor (e.g., one or more ova obtained from maturation of one or more oocytes isolated from a donor). The embryo may be freshly transferred to the subject, for example, by performing intrauterine embryo transfer using one or more embryos produced by fertilization within about 1 day to about 7 days, such as within about 3 days to about 5 days, of oocyte retrieval from the subject or donor. Embryo transfer is considered “fresh” when ovarian hyperstimulation and ovum / oocyte retrieval from the subject are performed during the same menstrual cycle as embryo transfer to the subject. Alternatively, the embryo may be cryopreserved for long-term storage and subsequently thawed prior to embryo transfer. This process is referred to herein as frozen embryo transfer (FET).

[0068] As used herein, a subject is considered to be “undergoing” an embryo transfer procedure if the subject is currently preparing for, is actually in the process of receiving, or has recently received, a transfer of one or more embryos to the uterus of the subject with the intention of establish pregnancy. For example, a subject is considered to be “undergoing” an embryo transfer procedure if the subject is (i) currently preparing for embryo transfer, e.g., by taking medication that stimulates follicular maturation and / or that promotes the ability of the endometrium to receive a transferred embryo, (ii) in the process of physically being transferred one or more embryos, or (iii) has undergone an embryo transfer procedure and is still clinically pregnant. Examples of subjects that are “preparing for embryo transfer” include subjects that are receiving, or that have recently received, one or more therapeutic agents designed to promote follicular maturation, such as human chorionic gonadotropin (hCG). Additional examples of subjects that are “preparing for embryo transfer” include subjects that are in the process of oocyte retrieval, as well as subjects that are receiving luteal phase support following oocyte retrieval, for example, with the aim of improving the endometrial condition so as to improve the likelihood of a successful embryo implantation.

[0069] As used herein, the term “luteal phase support” refers to administration of a progestin or progesterone to a subject undergoing embryo transfer therapy with the aim of enhancing the quality of the endometrium, for example, prior to the transfer of one or more embryos to the uterus of the subject. Exemplary luteal phase supports useful in conjunction with the compositions and methods of the disclosure include periodic administration of progesterone (e.g., intravaginally) starting within 24-48 hours of oocyte retrieval. As used herein, the term “endogenous” describes a molecule (e.g., a polypeptide, nucleic acid, or cofactor) that is found naturally in a particular organism (e.g., a human) or in a particular location within an organism (e.g., an organ, a tissue, or a cell, such as a human cell).

[0070] As used herein, the term “endometrial receptivity” refers to the ability of the uterus to provide optimal conditions to promote proper implantation and development of an embryo, such as an embryo produced ex vivo by in vitro fertilization of, or intracytoplasmic sperm injection into, an ovum (e.g., an ovum obtained directly from a subject undergoing an embryo transfer procedure therapy or by maturation of one or more oocytes obtained from a subject undergoing an embryo transfer procedure, or an ovum obtained directly from a donor not undergoing an embryo transfer procedure or by maturation of one or more oocytes obtained from a donor not undergoing an embryo transfer procedure). Exemplary factors that contribute to a subject’s level of endometrial receptivity include uterine perfusion and uterine contractility. Without being limited by mechanism, heightened uterine blood flow can augment the ability of the endometrium to successfully receive a transferred embryo in a manner conducive to implantation. Another factor that contributes to a subject’s level of endometrial receptivity is uterine contractility. Uterine contractions occurring at the time of embryo transfer can serve to expel a transferred embryo. Thus, high endometrial receptivity may be achieved when a subject’s uterine contractility is low. Endometrial receptivity may be enhanced (i.e., increased) using the compositions and methods described herein, for instance, by administration of an oxytocin receptor antagonist to a subject undergoing embryo transfer therapy prior to, concurrently with, and / or following the transfer of one or more embryos to the subject.

[0071] Enhanced endometrial receptivity may manifest clinically in one or more ways. For instance, a subject exhibiting enhanced endometrial receptivity (e.g., in response to treatment with an oxytocin receptor antagonist prior to, concurrently with, and / or following the transfer of one or more embryos to the subject) may exhibit decreased prostaglandin F2a (PGF2a) signaling in the subject’s endometrial and / or myometrial tissue. For instance, a subject can be determined to exhibit enhanced endometrial receptivity in response to oxytocin receptor antagonist administration if the subject demonstrates a reduced concentration of one or more secondary messengers involved in PGF2a signal transduction, such as diacylglycerol (DAG), inositol-1 ,4,5-trisphosphate (IP3), and / or intracellular calcium (Ca2+) released from Ca2+stores, such as sarcoplasmic reticula. For instance, a subject can be determined to exhibit enhanced endometrial receptivity in response to oxytocin receptor antagonist treatment as described herein by detecting a decrease in the concentration of one or more of the foregoing secondary messengers in a tissue sample, cell sample, or blood sample isolated from the subject’s endometrium and / or myometrium of 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, or more, relative to a measure of the secondary messenger prior to administration of the oxytocin receptor antagonist. Enhanced endometrial receptivity in a subject undergoing embryo transfer therapy can also be observed by assessing the ability of the subject to sustain pregnancy for a period of time following embryo transfer to the uterus of the subject. For instance, a subject exhibiting enhanced endometrial receptivity in response to oxytocin receptor antagonist therapy may sustain pregnancy for at least 14 days following transfer of one or more embryos to the subject, as assessed, for instance, by a blood pregnancy test, such as by detecting the presence and / or quantity of human chorionic gonadotropin (hCG) in a blood sample isolated from the subject using hCG tests known in the art and / or described herein. A subject exhibiting enhanced endometrial receptivity in response to oxytocin receptor antagonist therapy may sustain pregnancy for at least 6 weeks, such as for 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 13 weeks, 14 weeks, 15 weeks, 16 weeks, 17 weeks, 18 weeks, 19 weeks, 20 weeks, 21 weeks, 22 weeks, 23 weeks, 24 weeks, 25 weeks, 26 weeks, 27 weeks, 28 weeks, 29 weeks, 30 weeks, 31 weeks, 32 weeks, 33 weeks, 34 weeks, 35 weeks, 36 weeks, 37 weeks, 38 weeks, 39 weeks, or 40 weeks, following transfer of one or more embryos to the subject and / or following the retrieval of one or more oocytes or ova from the subject, as assessed, for instance, by detecting intrauterine embryo heartbeat. A subject exhibiting enhanced endometrial receptivity in response to oxytocin receptor antagonist therapy may give birth to a live offspring at a gestational age of at least 24 weeks, for instance, at a gestational age of 24 weeks, 25 weeks, 26 weeks, 27 weeks, 28 weeks, 29 weeks, 30 weeks, 31 weeks, 32 weeks, 33 weeks, 34 weeks, 35 weeks, 36 weeks, 37 weeks, 38 weeks, 39 weeks, or 40 weeks.

[0072] As used herein, the term “exogenous” describes a molecule (e.g., a polypeptide, nucleic acid, or cofactor) that is not found naturally in a particular organism (e.g., a human) or in a particular location within an organism (e.g., an organ, a tissue, or a cell, such as a human cell). Exogenous materials include those that are provided from an external source to an organism or to cultured matter extracted there from.

[0073] As used herein, the term “gestational age” describes how far along a particular pregnancy is, and is measured from the first day of a pregnant female subject's last menstrual cycle to the current date. As used herein, the term “labor” (which may also be termed birth) relates to the expulsion of the fetus and placenta from the uterus of a pregnant female subject. For a normal pregnancy, labor may occur at a gestational age of about 40 weeks. “Preterm labor” as used herein refers to a condition in which labor commences more than three weeks before the full gestation period, which is typically about 40 weeks. That is, preterm labor occurs at any stage prior to, e.g., 38 weeks of gestation. Preterm labor typically leads to the occurrence of labor, or physiological changes associated with labor in a pregnant female subject, if not treated. Preterm labor may or may not be associated with vaginal bleeding or rupture of uterine membranes. Preterm labor may also be referred to as premature labor. The avoidance of preterm labor in a subject will prolong the term of pregnancy and may therefore avoid preterm delivery, thus reducing the risk of neonatal mortality and morbidity.

[0074] As used herein, the term “gonadotropin-releasing hormone antagonist” or “GnRH antagonist” refers to a compound capable of inhibiting the gonadotropin-releasing hormone receptor, e.g., such that release of one or more gonadotropins (such as follicle stimulating hormone and luteinizing hormone) is inhibited. GnRH antagonists include 2-phenylethylpyrimidine-2,4(1 H,3H)-dione derivatives, such as those described in US Patent Nos. 7,056,927; 7,176,21 1 ; and 7,419,983; the disclosures of each of which are incorporated herein by reference in their entirety. Exemplary GnRH antagonists include elagolix, relugolix, ASP-1707, and SKI2670, among others.

[0075] As used herein, the term “ICso” refers to the concentration of a substance (antagonist) that reduces the efficacy of a reference agonist or the constitutive activity of a biological target by 50%, for instance, as measured in a competitive ligand binding assay or in a cell-based functional assay, such as a Ca2+mobilization assay. Exemplary Ca2+mobilization assays that can be used to determine the IC50 of oxytocin receptor antagonist include fluorimetric imaging assays, such as those described in US Patent No. 9,670,155, the disclosure of which is incorporated herein by reference in its entirety.

[0076] As used herein, the term “in vitro fertilization” (IVF) refers to a process in which an ovum, such as a human ovum, is contacted ex vivo with one or more sperm cells so as to promote fertilization of the ovum and zygote formation. The ovum can be derived from a subject, such as a human subject, undergoing embryo transfer therapy. For instance, the ovum may be obtained from maturation of one or more oocytes isolated from the subject, e.g., from about 1 day to about 7 days prior to embryo transfer to the subject (such as from about 3 days to about 5 days prior to embryo transfer to the subject). The ovum may also be retrieved directly from the subject, for instance, by transvaginal ovum retrieval procedures known in the art. Alternatively, the ovum may be derived or isolated from a donor.

[0077] As used herein, the term “intracytoplasmic sperm injection” (ICSI) refers to a process in which a sperm cell is injected directly into an ovum, such as a human ovum, so as to promote fertilization of the ovum and zygote formation. The sperm cell may be injected into the ovum, for instance, by piercing the oolemma with a microinjector so as to deliver the sperm cell directly to the cytoplasm of the ovum. ICSI procedures useful in conjunction with the compositions and methods described herein are known in the art and are described, for instance, in WO 2013 / 158658, WO 2008 / 051620, and WO 2000 / 009674, among others, the disclosures of which are incorporated herein by reference as they pertain to compositions and methods for performing intracytoplasmic sperm injection.

[0078] As used herein, the term “miscarriage” refers to a naturally-occurring, spontaneous termination of a pregnancy at a stage in which the embryo or fetus is incapable of surviving independently of the mother. For instance, in human subjects, an embryo or fetus may be incapable of surviving independently of the mother at a gestational age of less than about 20 weeks (e.g., a gestational age of less than about 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 13 weeks, 14 weeks, 15 weeks, 16 weeks, 17 weeks, 18 weeks, 19 weeks, or 20 weeks).

[0079] As used herein, the term "oral bioavailability" refers to the fraction of a compound administered to a subject, such as a mammal (e.g., a human) that reaches systemic circulation in the subject, and that is not sequestered in a non-target organ or excreted without absorption via the gastrointestinal tract. The term refers to a blood plasma concentration that is integrated over time and is typically expressed as a percentage of the orally administered dose. As used herein, the terms “ovum” and “mature oocyte” refer to a mature haploid female reproductive cell or gamete. In the context of assisted reproductive technology as described herein, ova may be produced ex vivo by maturation of one or more oocytes isolated from a subject undergoing embryo transfer therapy. Ova may also be isolated directly from the subject, for example, by transvaginal ovum retrieval methods described herein or known in the art.

[0080] As used herein, the terms “oxytocin receptor antagonist,” “OTR antagonist,” “oxytocin antagonist,” and the like are used interchangeably and refer to a compound capable of inhibiting the oxytocin receptor, for example, such that activity of one or more downstream signaling molecules in the oxytocin signal transduction cascade is inhibited. Oxytocin receptor antagonists for use with the compositions and methods described herein include (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime.

[0081] As used herein, the term “pharmaceutical composition” refers to a mixture containing a therapeutic compound, such as an oxytocin receptor antagonist described herein, to be administered to a subject, such as a mammal, e.g., a human, in order to prevent, treat or control a particular disease or condition affecting or that may affect the mammal, such as to reduce the likelihood of embryo implantation failure in a subject undergoing embryo transfer therapy.

[0082] As used herein, the term “pharmaceutically acceptable” refers to those compounds, materials, compositions and / or dosage forms, which are suitable for contact with the tissues of a subject, such as a mammal (e.g., a human) without excessive toxicity, irritation, allergic response and other problem complications commensurate with a reasonable benefit / risk ratio.

[0083] As used herein, the term “prostaglandin F2a signaling” or “PGF2a signaling” refers to the endogenous signal transduction cascade by which PGF2a potentiates the intracellular activity of the PGF2a receptor so as to effect one or more biological responses. PGF2a signaling encompasses the PGF2a-mediated stimulation of the PGF2a receptor (FP), a G protein-coupled receptor, which leads to the activation of the Gqprotein and, in turn phospholipase C (PLC), phosphatidylinositol-3-kinase (PI3K), and extracellular signal-regulated kinases (ERK) 1 and 2. PGF2a signaling can be detected by observing an increase in the concentration of phosphatidylinsolitol-4,5-bisphosphate (PIP2) and / or a decrease in the concentration of one or more secondary messengers involved in PGF2a signal transduction, such as diacylglycerol (DAG), inositol-1 ,4,5-trisphosphate (IP3), and / or intracellular calcium (Ca2+) released from Ca2+stores, such as sarcoplasmic reticula. The PGF2a signal transduction cascade is described in detail, for instance, in Xu et al., Reproduction 149:139-146 (2015), the disclosure of which is incorporated herein by reference as it pertains to the proteins and messengers involved in PGF2a signaling.

[0084] As used herein, the term “sample” refers to a specimen (e.g., blood, blood component (e.g., serum or plasma), urine, saliva, amniotic fluid, cerebrospinal fluid, tissue (e.g., placental or dermal), pancreatic fluid, chorionic villus sample, and / or cells) isolated from a subject.

[0085] As used herein, the phrases “specifically binds” and “binds” refer to a binding reaction which is determinative of the presence of a particular protein in a heterogeneous population of proteins and other biological molecules that is recognized, e.g., by a ligand with particularity. A ligand (e.g., a protein, peptide, or small molecule) that specifically binds to a protein will bind to the protein, e.g., with a KD of less than 100 nM. For example, a ligand that specifically binds to a protein may bind to the protein with a KD of up to 100 nM (e.g., between 1 pM and 100 nM). A ligand that does not exhibit specific binding to a protein or a domain thereof may exhibit a KD of greater than 100 nM (e.g., greater than 200 nM, 300 nM, 400 nM, 500 nM, 600 nm, 700 nM, 800 nM, 900 nM, 1 pM, 100 pM, 500 pM, or 1 mM) forthat particular protein or domain thereof. A variety of assay formats may be used to determine the affinity of a ligand for a specific protein. For example, solid-phase ELISA assays are routinely used to identify ligands that specifically bind a target protein. See, e.g., Harlow & Lane, Antibodies, A Laboratory Manual, Cold Spring Harbor Press, New York (1988) and Harlow & Lane, Using Antibodies, A Laboratory Manual, Cold Spring Harbor Press, New York (1999), for a description of assay formats and conditions that can be used to determine specific protein binding.

[0086] As used herein, the terms “subject” and “patient” are interchangeable and refer to an organism that receives treatment for a particular disease or condition as described herein. Examples of subjects and patients include mammals, such as humans, such as those undergoing embryo transfer therapy and are to receive medication so as to improve endometrial receptivity, reduce the likelihood of embryo implantation failure, or otherwise improve the likelihood of achieving and maintaining pregnancy.

[0087] As used herein, the term “substantially pure” refers to a compound that has a purity of at least 85%, as assessed, for instance, using nuclear magnetic resonance (NMR) and / or high-performance liquid chromatography (HPLC) techniques described herein or known in the art.

[0088] As used herein, the term “tmax” refers to the time following administration of a compound to a subject at which the compound exhibits a maximum concentration in the blood (e.g., serum or plasma) of the subject.

[0089] A compound, salt form, crystal polymorph, therapeutic agent, or other composition described herein may be referred to as being characterized by graphical data “substantially as depicted in” a figure. Such data may include, without limitation, powder X-ray diffractograms, NMR spectra, differential scanning calorimetry curves, and thermogravimetric analysis curves, among others. As is known in the art, such graphical data may provide additional technical information to further define the compound, salt form, crystal polymorph, therapeutic agent, or other composition. As is understood by one of skill in the art, such graphical representations of data may be subject to small variations, e.g., in peak relative intensities and peak positions due to factors such as variations in instrument response and variations in sample concentration and purity. Nonetheless, one of skill in the art will readily be capable of comparing the graphical data in the figures herein with graphical data generated for a compound, salt form, crystal polymorph, therapeutic agent, or other composition and confirm whether the two sets of graphical data are characterizing the same material or two different materials. For instance, a crystal form of (3Z,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime referred to herein as being characterized by graphical data "substantially as depicted in" a figure will thus be understood to include any crystal form of (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime characterized by the graphical data, optionally having one or more of small variations, e.g., one or more variations described above or known to one of skill in the art.

[0090] As used herein, the terms “treat” or “treatment” in the context of a subject undergoing embryo transfer therapy refer to treatment, for instance, by administration of an oxytocin receptor antagonist, with the intention of enhancing endometrial receptivity thereby reducing the likelihood of embryo implantation failure and promoting pregnancy in the subject. Those in need of treatment include, for example, female mammalian subjects, such as female human subjects, that are undergoing embryo transfer therapy, such as subjects undergoing oocyte or ovum retrieval followed by in vitro fertilization or intracytoplasmic sperm injection and subsequent embryo transfer. Those in need of treatment also include, for example, female mammalian subjects, such as female human subjects, that are undergoing embryo transfer therapy, for example, using embryos produced ex vivo by in vitro fertilization or intracytoplasmic sperm injections of one or more ova derived from a donor (e.g., isolated directly from a donor by transvaginal ovum retrieval or by maturation of one or more oocytes obtained directly from the donor). The subject may be undergoing fresh embryo transfer or frozen embryo transfer, and may be transferred, for instance, one, two, three, or more embryos according to the methods described herein. The subject may be one that has previously undergone embryo transfer therapy, either successfully or unsuccessfully, including subjects that have previously undergone one or more cycles (for instance, one, two, three, four, five, six, seven, eight, nine, ten, or more cycles) of failed embryo transfer therapy.

[0091] A subject can be considered to have been treated, for instance, by administration of an oxytocin receptor antagonist according to the methods described herein, if the subject exhibits one or more beneficial outcomes following administration of the oxytocin receptor antagonist. Exemplary beneficial outcomes that are indicative of treatment include an increase in the subject’s endometrial perfusion, a reduction in the subject’s uterine contractility, increase in the subject’s endometrial receptivity toward a transferred embryo, a reduction in the likelihood of embryo implantation failure, the prevention of a miscarriage, and the achievement and maintenance of a successful pregnancy, for example, until delivery at a full gestational age, in a subject following transfer of one or more embryos to the subject.

[0092] As an example, endometrial receptivity can be observed in a variety of clinical manifestations, including a reduction in prostaglandin F2a (PGF2a) signal transduction following oxytocin receptor antagonist administration, successful implantation of the embryo into the endometrium of the subject, as well as the subject’s capacity to achieve and sustain pregnancy following embryo transfer, such as for about 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23 days, 24 days, 25 days, 26 days, 27 days, 28 days, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 13 weeks, 14 weeks, 15 weeks, 16 weeks, 17 weeks, 18 weeks, 19 weeks, 20 weeks, 21 weeks, 22 weeks, 23 weeks, 24 weeks, 25 weeks, 26 weeks, 27 weeks, 28 weeks, 29 weeks, 30 weeks, 31 weeks, 32 weeks, 33 weeks, 34 weeks, 35 weeks, 36 weeks, or more, following the transfer of one or more embryos to the subject and / or following the retrieval of one or more oocytes or ova from the subject. Pregnancy can be assessed using methods described herein or known in the art, such as by detecting and / or quantifying human chorionic gonadotropin (hCG) in a blood sample isolated from the subject and / or by detecting intrauterine embryo heartbeat.

[0093] As used herein, the term “unit dosage form” refers to a single, discrete composition containing a therapeutic agent, such as an oxytocin receptor antagonist described herein, formulated in a manner appropriate for administration to a subject, such as a subject undergoing embryo transfer therapy as described herein. Unit dosage forms include solid and liquid formulations, such as tablets (e.g., dispersible tablets), capsules, gel caps, powders, liquid solutions, liquid suspensions, granulate, microemulsion, and self-emulsifying dispersed system. A subject may be administered a single dose of a therapeutic agent by administration of one or more unit dosage forms. As an example, a single dose of 100 mg of a therapeutic agent can be administered using two 50 mg unit dosage forms of the therapeutic agent.

[0094] As used herein, the term “uterine contractility” refers to a measurement of the frequency and / or amplitude of uterine contractions that a subject exhibits at a particular time. Uterine contractility can be measured, for example, by assessing the quantity of uterine contractions that the subject exhibits over a certain time period; this is the frequency of the subject’s uterine contractions. Another measure of uterine contractility is the work done by one or more uterine contractions that the subject exhibits. Methods of measuring uterine contractility are described, for example, in US Patent No. 9,670,155, the disclosure of which is incorporated herein by reference.

[0095] As used herein, the term "poloxamer" refers to a non-ionic triblock copolymer composed of a central hydrophobic chain of polyoxypropylene flanked by two hydrophilic chains of polyoxyethylene. Poloxamers are also known by the trade name of "Pluronics" or "Synperonics" (BASF). The block copolymer can be represented by the following formula: HO(C2H4O)x(C3H6O)y(C2H4O)zH. The lengths of the polymer blocks can be customized. As a result, many different poloxamers exist. Poloxamers suitable for use in conjunction with the compositions and methods of the present disclosure include those having an average molecular weight of at least about 7,000 g / mol. Since the synthesis of block copolymers is associated with a natural degree of variation from one batch to another, the numerical values recited above (and those used herein to characterize a given poloxamer) may not be precisely achievable upon synthesis, and the average value will differ to a certain extent. Thus, the term "poloxamer" as used herein can be used interchangeably with the term "poloxamers" (representing an entity of several poloxamers, also referred to as mixture of poloxamers) if not explicitly stated otherwise. The term "average" in relation to the number of monomer units or molecular weight of (a) poloxamer(s) as used herein is a consequence of the technical inability to produce poloxamers all having the identical composition and thus the identical molecular weight. Poloxamers produced according to state of the art methods will be present as a mixture of poloxamers each showing a variability as regards their molecular weight, but the mixture as a whole averaging the molecular weight specified herein. BASF and Sigma Aldrich are suitable sources of poloxamers for use in conjunction with the compositions and methods of the disclosure. As used herein, the term “co-crystal” refers to a crystalline form composed of two or more molecules in the same crystal lattice.

[0096] As used herein, the term “pseudopolymorph” refers to a crystalline form of a therapeutic agent which incorporates or introduces an additional molecule. The additional molecule may be, e.g., an excipient (e.g., a poloxamer), solvent, or water.

[0097] As used herein, the term “complex” refers to a crystalline form that is associated with an additional molecular entity (e.g., an excipient, such as a poloxamer). The bonding between the components is normally weaker than in a covalent bond.

[0098] As used herein, the term “crystalline Form B” and “Form B” are used interchangeably and refer to a solid form of compound (I) and Poloxamer 188 having one or more, or all, of the characteristic X-ray powder diffraction peaks as set forth in Table 1 . In some embodiments, crystalline Form B is a co-crystal of compound (I) and Poloxamer 188. In some embodiments, crystalline Form B is a pseudopolymorph, having a crystal lattice of compound (I) incorporated with Poloxamer 188. In some embodiments, the crystalline Form B is a complex of an anhydrous crystal lattice of compound (I), in association with Poloxamer 188. Crystalline Form B is characterized by one or more of the XRPD spectrum as shown in FIG. 5A, the DSC curve substantially as depicted in FIG. 5F, the1H NMR spectrum substantially as depicted in FIG. 5B, the TGA curve substantially as depicted in FIG. 5G, the DVS curve substantially as depicted in FIG. 5H, and the Raman curve substantially as depicted in FIG. 5I or FIG. 5J. Crystalline Form B is further characterized in Examples 2-4, below.

[0099] As used herein, the term “crystalline Form A” and “Form A” are used interchangeably and refer to a solid form of anhydrous compound (I) having one or more, or all, of the characteristic X-ray powder diffraction peaks as set forth in Table 2. Crystalline Form A is characterized by one or more of the XRPD spectrum as shown in FIG. 1A, the DSC curve substantially as depicted in FIG. 1 B or FIG. 1C, the TGA curve substantially as depicted in FIG. 1 D or 1 E, and the DVS curve substantially as depicted in FIG. 1 F. Crystalline Form A is further characterized in Examples 1 and 4, below.

[0100] Brief Description of the Figures

[0101] FIG. 1 A is an X-ray powder diffractogram (XRPD) for crystalline Form A of compound (I). Experiments were performed as described in Example 1 .

[0102] FIG. 1B is an exemplary differential scanning calorimetry (DSC) curve for crystalline Form A of compound (I) prepared by crystallization from diethyl ether. Experiments were performed as described in Example 1 .

[0103] FIG. 1C is an exemplary DSC curve for crystalline Form A of compound (I) prepared by crystallization from ethyl acetate. Experiments were performed as described in Example 1.

[0104] FIG. 1D is an exemplary thermogravimetric analysis (TGA) curve for crystalline Form A of compound (I) prepared by crystallization from diethyl ether. Experiments were performed as described in Example 1 . FIG. 1 E is an exemplary TGA curve for crystalline Form A of compound (I) prepared by crystallization from ethyl acetate. Experiments were performed as described in Example 1.

[0105] FIG. 1 F is an exemplary dynamic vapor sorption (DVS) curve for crystalline Form A of compound (I) prepared by crystallization from diethyl ether. Experiments were performed as described in Example 1.

[0106] FIGS. 2A-2P are XRPD spectra showing attempted crystallization of compound (I) in combination with various excipients. Experiments were performed as described in Example 2. Attempts to crystallize compound (I) from ethanol in the presence of calcium silicate, glyceryl dibehenate, povidone, lactose monohydrate, magnesium stearate, microcrystalline cellulose, saccharin sodium dihydrate, or sodium croscarmellose were unsuccessful. XRPD patterns of the resulting materials were consistent with the excipient or mixtures of amorphous compound (I) and the excipient.

[0107] FIG. 3A is an XRPD overlay of crystalline Form B and Form A. Experiments were performed as described in Example 2.

[0108] FIG. 3B is an XRPD overlay of crystalline Form B, Poloxamer 188, and amorphous compound (I). Experiments were performed as described in Example 2.

[0109] FIGS. 4A-4Q are XRPD spectra showing formation of crystalline Form B under various conditions. Experiments were performed as described in Example 2.

[0110] FIG. 5A is an XRPD of crystalline Form B. Experiments were performed as described in Example 3.

[0111] FIGS. 5B-5E show an exemplary proton NMR spectrum of crystalline Form B. Experiments were performed as described in Example 3.

[0112] FIG. 5F is an exemplary DSC curve of crystalline Form B. Experiments were performed as described in Example 3.

[0113] FIG. 5G is an exemplary TGA curve of crystalline Form B. Experiments were performed as described in Example 3.

[0114] FIG. 5H is an exemplary DVS curve of crystalline Form B. Experiments were performed as described in Example 3.

[0115] FIGS. 5I-5J are exemplary Raman spectra of crystalline Form B. Experiments were performed as described in Example 3.

[0116] FIG. 6 is a pH solubility profile of crystalline Form B after stirring at 25°C for 24 hours. Experiments were performed as described in Example 4.

[0117] FIG. 7A is an intrinsic dissolution rate (IDR) profile overlay of amorphous compound (I), crystalline Form A, crystalline Form B, and compound (I) in crystalline Form B in water. Experiments were performed as described in Example 4.

[0118] FIG. 7B is an IDR profile overlay of amorphous compound (I) and crystalline Form A in pH 7 phosphate buffer. Experiments were performed as described in Example 4.

[0119] FIGS. 8A-8J are XRPD of recovered solids following solubility experiments, compression experiments, and IDR experiments with amorphous compound (I) (FIGS. 8A-8C), crystalline Form A (FIGS. 8D-8G), and crystalline Form B (FIGS. 8H-8J). Experiments were performed as described in Example 4.

[0120] FIG. 9 is a DSC thermogram overlay of crystalline Form B and crystalline Form A. Experiments were performed as described in Example 4.

[0121] Detailed Description

[0122] The present disclosure provides compositions and methods that can be used to treat a subject (e.g., a female human subject) undergoing embryo transfer procedures. Exemplary responses to oxytocin receptor antagonist treatment include reduced uterine contractility and enhanced blood flow to the endometrium. Collectively, these phenotypes contribute to a subject’s endometrial receptivity toward a transferred embryo. Without being bound by theory, an oxytocin receptor antagonist administered to a subject using the compositions and methods described herein may enhance uterine perfusion and suppress uterine contractions that could otherwise lead to embryo expulsion, ultimately serving to create an environment within the endometrium that is conducive to successful embryo implantation and the establishment and maintenance of a healthy pregnancy.

[0123] Particularly, the disclosure features crystalline forms of the oxytocin receptor antagonist, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I), below.

[0124] Using the methods described herein, one can administer an oxytocin receptor antagonist, such as a crystalline form of compound (I), to a subject, such as a mammalian subject (e.g., a female human subject) in order to promote enhanced endometrial receptivity, reduce the likelihood of embryo implantation failure, and / or prevent miscarriage in a subject following the transfer of one or more embryos to the uterus of the subject.

[0125] The subject may be one that has previously undergone one or more successful or unsuccessful embryo implantation procedures. Alternatively, the subject may be one that has not undergone a previous embryo transfer cycle. According to the methods described herein, the one or more embryos that are ultimately transferred to the subject can be obtained, for instance, by in vitro fertilization (IFV) or intracytoplasmic sperm injection (ICSI) of an ovum isolated or derived from the subject or from a donor. For instances in which the ovum is isolated or derived from the subject, the ovum may be isolated from the subject directly or may be produced ex vivo by inducing maturation of one or more oocytes isolated from the subject. Following fertilization of the ovum by contact with one or more sperm cells, the subsequently formed zygote can be matured ex vivo so as to produce an embryo, such as a morula or blastula (e.g., a mammalian blastocyst), which can then be transferred to the uterus of the subject for implantation into the endometrium. Embryo transfers that can be performed using the methods described herein include fresh embryo transfers, in which the ovum or oocyte used for embryo generation is retrieved from the subject and the ensuing embryo is transferred to the subject during the same menstrual cycle. The embryo can alternatively be produced and cryopreserved for long-term storage prior to transfer to the subject.

[0126] The sections that follow provide a description of the foregoing oxytocin receptor antagonist in further detail, as well as a description of methods for administering this agent to subjects in order to enhance endometrial receptivity upon embryo transfer, reduce the likelihood of embryo implantation failure, and / or prevent the occurrence of a miscarriage in a subject undergoing an assisted reproduction procedure, among other beneficial phenotypes described herein.

[0127] (3Z,5S)-5-(hydroxymethyl)-1 -[(2'-methyl-1 ,T-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O- methyloxime (Compound I)

[0128] Compound (I) ((3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3- one O-methyloxime) is a non-peptide oxytocin receptor antagonist that can be used to enhance endometrial receptivity, promote successful embryo implantation, and reduce the likelihood of miscarriage in subjects undergoing or that have undergone embryo transfer therapy. Compound (I) is an orally-active oxytocin receptor antagonist capable of inhibiting human oxytocin receptor with a Ki of 52 nM and suppressing Ca2+mobilization in cultured HEK293EBNA cells with an IC50 of 81 nM. Additionally, compound (I) selectively inhibits the oxytocin receptor over the vasopressin Via receptor, as compound (I) inhibits the vasopressin Via receptor with a Ki of 120 nM.

[0129] Synthesis of compound (I)

[0130] An exemplary procedure for the synthesis of compound (I) is shown in Scheme 1 , below.

[0131] Scheme 1. Exemplary synthesis of compound (I)

[0132] Purity of Compound (I)

[0133] In some embodiments, the compound represented by formula (I) (i.e., (3Z,5S)-5-(hydroxymethyl)- 1-[(2'-methyl-1 ,1 '-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime) is substantially pure. For instance, in some embodiments, the compound represented by formula (I) has a purity of at least 85%, such as a purity of from 85% to 99.9% or more (e.g., a purity of 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1 %, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or more). The purity of the compound represented by formula (I) may be assessed, for instance, using NMR techniques and / or chromatographic methods, such as HPLC procedures, that are known in the art and described herein, such as those techniques that are described in US Patent No. 9,670,155, the disclosure of which is incorporated herein by reference in its entirety.

[0134] In some embodiments, the compound represented by formula (I) is substantially pure with respect to diastereomers of this compound and other by-products that may be formed during the synthesis of this compound. For instance, in some embodiments, the compound represented by formula (I) has a purity of at least 85%, such as a purity of from 85% to 99.9% or more (e.g., a purity of 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or more) with respect to diastereomers of this compound and other by-products that may be formed during the synthesis of this compound. The purity of the compound represented by formula (I) may be assessed, for instance, using NMR techniques and / or chromatographic methods, such as HPLC procedures, that are known in the art and described herein, such as those techniques that are described in US Patent No. 9,670,155.

[0135] In some embodiments, the compound represented by formula (I) is substantially pure with respect to its (3E) diastereomer, (3E,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime. For instance, in some embodiments, the compound represented by formula (I) has a purity of at least 85%, such as a purity of from 85% to 99.9% or more (e.g., a purity of 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or more) with respect to (3E,5S)-5- (hydroxymethyl)-1-[(2'-methyl-1 ,1'-biphenyl-4-yl)carbonyl]pyrrolidin-3-one O-methyloxime. For instance, compound (I) may be administered in the form of a composition (e.g., a tablet, such as a dispersible tablet, capsule, gel cap, powder, liquid solution, liquid suspension, granulate, microemulsion, or selfemulsifying dispersed system) that contains less than 15% of the (3E) diastereomer. For example, compound (I) may be administered in the form of a composition (e.g., a tablet, such as a dispersible tablet, capsule, gel cap, powder, liquid solution, liquid suspension, granulate, microemulsion, or selfemulsifying dispersed system) that contains less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.1%, less than 0.01%, less than 0.001%, or less of the (3E) diastereomer. The purity of the compound represented by formula (I) may be assessed, for instance, using NMR techniques and / or chromatographic methods, such as HPLC procedures, that are known in the art and described herein, such as those techniques that are described in US Patent No. 9,670,155.

[0136] Therapeutic activity

[0137] The present disclosure is based in part on the discovery of crystalline forms of compound (I), and the characterization of their ability to promote successful endometrial implantation of a transferred embryo in female human subjects. Compound (I) has been found to reduce the risk of embryo implantation failure in clinical studies conducted with human subjects that previously underwent ovarian hyperstimulation and oocyte retrieval. It has been discovered that compound (I) increases the rate of successful embryo implantation as assessed by a variety of metrics. These manifestations have been found to include an increase in the rate of positive pregnancy tests at 14 days, 6 weeks, and 10 weeks following embryo transfer and / or oocyte retrieval, as well as an increase in the rate of live births at a gestational age of at least 24 weeks.

[0138] In particular, compound (I) is particularly effective at enhancing endometrial receptivity by multiple modes of action. Particularly, compound (I) has been discovered to reduce uterine contractility and augment the flow of blood to the endometrium. These effects provide important therapeutic benefits to patients undergoing embryo transfer therapy. Together, the reduced uterine contractile activity and enhanced endometrial perfusion engendered by compound (I) create an environment in the uterus that is conducive to successful embryo implantation. Without being limited by mechanism, these biological activities represent ways in which compounds of the disclosure may reduce the likelihood of embryo implantation failure and of miscarriage.

[0139] Follicular Maturation and Oocyte / Ovum Retrieval

[0140] A variety of methods can be used in order to induce follicular maturation and to perform oocyte (e.g., mature oocyte) retrieval in conjunction with the compositions and methods described herein. In some embodiments, ova or oocytes are isolated from the subject from about 1 day to about 7 days prior to the transfer of the one or more embryos to the subject, such as from about 2 days to about 5 days prior to embryo transfer. The ova or oocytes isolated from the subject may include mature oocytes, such as from 1 to 4 mature oocytes that are ready for fertilization upon contact with one or more sperm cells. The ova or oocytes may be isolated from a subject undergoing embryo transfer therapy or from a donor, such as a familial donor.

[0141] A subject undergoing embryo transfer therapy or a donor may be prepared for ovum or oocyte retrieval by controlled ovarian hyperstimulation, for instance, according to methods described herein or known in the art. For example, a subject or donor may be administered a GnRH antagonist so as to prevent a premature increase in the serum concentration of luteinizing hormone (LH). Additionally or alternatively, final follicular maturation can be achieved by administration of hCG to the subject or donor prior to isolation of the one or more ova or oocytes. For instance, the hCG can be administered to the subject in a single dose or in multiple doses, for instance, by intravenous injection according to procedures known in the art.

[0142] In some embodiments, a luteal support is provided to the subject or donor following ovum or oocyte retrieval. This may be performed, for instance, by administering progesterone to the subject or donor following the retrieval procedure. For example, progesterone may be administered to the subject or donor intravaginally at a dose of from about 300 mg to about 600 mg. The progesterone may be administered to the subject in a single dose or in multiple doses. For instance, progesterone may be administered to the subject in regularly spaced intervals beginning within about 24 hours of isolation of the one or more ova or oocytes, such as within 12 hours of retrieval, and continuing for about 6 or more weeks following the transfer of the one or more embryos to the subject. Embryo Quality and Condition

[0143] Embryos for use in conjunction with the compositions and methods described herein include those that are at, for example, the morula or the blastula stage of embryonic development. For instance, embryos that may be transferred to a subject as described herein include those that contain from 6 to 8 blastomeres immediately prior to transfer of the one or more embryos to the subject. The blastomeres may be of approximately equal sizes as assessed by visual microscopy prior to the transfer of the one or more embryos to the subject.

[0144] Embryos for use in conjunction with the compositions and methods described herein include those that are formed, for instance, by IVF or ICSI methods known in the art. In some embodiments, the embryos are freshly transferred to the uterus of the subject, for instance, from about 1 day to about 7 days (e.g., from about 2 days to about 5 days) following the isolation of one or more oocytes or ova from the subject for IVF or ICSI. In some embodiments, the one or more embryos are frozen and cryopreserved for long-term storage prior to thaw and transfer to the subject. Methods for the cryopreservation of embryos are known in the art and have been described, for instance, in WO 1991 / 003935 and WO 2010 / 011766, the disclosures of each of which are incorporated herein by reference as they pertain to compositions and procedures for cryopreserving embryos for long-term storage.

[0145] Methods of Assessing Pregnancy

[0146] Techniques for assessing pregnancy for use in conjunction with the compositions and methods described herein include qualitative and quantitative assessments of a sample isolated from a subject, such as a sample of blood or urine. Methods for assessing pregnancy include detecting the presence and / or quantity of hCG in a sample isolated from a subject. This can be achieved, for instance, using conventional receptor-ligand binding assays known in the art, such as through the use of competitive radioligand binding assays, which are described for the detection of hCG in US Patent No. 4,094,963, the disclosure of which is incorporated herein by reference as it pertains to methods of detecting hCG in subject samples to assess pregnancy. Additionally or alternatively, test strips may be used to determine hCG concentrations, as described, for instance, in US Patent No. 7,989,217, the disclosure of which is incorporated herein by reference as it pertains to methods of detecting hCG in subject samples to assess pregnancy. Urine samples isolated from a subject can additionally be analyzed in order to determine pregnancy, as described, for instance, in US Patent No. 4,315,908, the disclosure of which is incorporated herein by reference as it pertains to methods of detecting hCG in subject samples to assess pregnancy.

[0147] Additionally or alternatively, pregnancy may be assessed by detecting intrauterine heartbeat, such as the heartbeat of the embryo or developing fetus following successful embryo implantation. Compositions and methods for detecting embryonic and fetal heartbeat are known in the art and are described, for instance, in US Patent Nos. 3,780,725 and 4,437,467, the disclosures of each of which are incorporated herein by reference as they pertain to methods of detecting heartbeat to assess pregnancy in a subject.

[0148] Following embryo transfer, for instance, as described herein, a subject may be subject to one or more pregnancy tests, for example, using one or more of the foregoing procedures. The subject may be tested for pregnancy at one or more points following embryo transfer therapy, such as at about 14 days, about 6 weeks, about 10 weeks, or longer, following embryo transfer and / or oocyte retrieval.

[0149] Pharmaceutical Compositions

[0150] The present disclosure features crystalline forms of compound (I). In some embodiments, the crystalline form may be a solid form of compound (I) and a poloxamer. Exemplary poloxamers are provided in the section that follows.

[0151] Poloxamers

[0152] Poloxamers that may be used in conjunction with the compositions and methods of the disclosure include those having an average molar mass of polyoxypropylene subunits of greater than 1 ,600 g / mol (e.g., an average molar mass of polyoxypropylene subunits of about 1 ,700 g / mol, 1 ,800 g / mol, 1 ,900 g / mol, 2,000 g / mol, 2,100 g / mol, 2,200 g / mol, 2,300 g / mol, 2,400 g / mol, 2,500 g / mol, 2,600 g / mol, 2,700 g / mol, 2,800 g / mol, 2,900 g / mol, 3,000 g / mol, 3,100 g / mol, 3,200 g / mol, 3,300 g / mol, 3,400 g / mol, 3,500 g / mol, 3,600 g / mol, 3,700 g / mol, 3,800 g / mol, 3,900 g / mol, 4,000 g / mol, 4,100 g / mol, 4,200 g / mol, 4,300 g / mol, 4,400 g / mol, 4,500 g / mol, 4,600 g / mol, 4,700 g / mol, 4,800 g / mol, 4,900 g / mol, or 5,000 g / mol).

[0153] In some embodiments, the poloxamer has an average molar mass of polyoxypropylene subunits of from about 1 ,600 g / mol to about 4,000 g / mol (e.g., about 1 ,600 g / mol, about 1 ,650 g / mol, about 1 ,700 g / mol, about 1 ,750 g / mol, about 1 ,800 g / mol, about 1 ,850 g / mol, about 1 ,900 g / mol, about 1 ,950 g / mol, about 2,000 g / mol, about 2,050 g / mol, 2,100 g / mol, 2,150 g / mol, 2,200 g / mol, 2,250 g / mol, 2,300 g / mol,

[0154] 2,350 g / mol, 2,400 g / mol, 2,450 g / mol, 2,500 g / mol, 2,550 g / mol, 2,600 g / mol, 2,650 g / mol, 2,700 g / mol,

[0155] 2,750 g / mol, 2,800 g / mol, 2,850 g / mol, 2,900 g / mol, 2,950 g / mol, 3,000 g / mol, 3,050 g / mol, 3,100 g / mol,

[0156] 3,150 g / mol, 3,200 g / mol, 3,250 g / mol, 3,300 g / mol, 3,350 g / mol, 3,400 g / mol, 3,450 g / mol, 3,500 g / mol,

[0157] 3,550 g / mol, 3,600 g / mol, 3,650 g / mol, 3,700 g / mol, 3,750 g / mol, 3,800 g / mol, 3,850 g / mol, 3,900 g / mol,

[0158] 3,950 g / mol, or 4,000 g / mol).

[0159] In some embodiments, the poloxamer has an average molar mass of polyoxypropylene subunits of from about 1 ,600 g / mol to about 2,000 g / mol (e.g., about 1 ,600 g / mol, 1 ,650 g / mol, 1 ,700 g / mol, 1 ,750 g / mol, 1 ,800 g / mol, 1 ,850 g / mol, 1 ,900 g / mol, 1 ,950 g / mol, or 2,000 g / mol).

[0160] In some embodiments, the poloxamer has an average molar mass of polyoxypropylene subunits of from about 1 ,700 g / mol to about 1 ,900 g / mol (e.g., about 1 ,700 g / mol, 1 ,710 g / mol, 1 ,720 g / mol, 1 ,730 g / mol, 1 ,740 g / mol, 1 ,750 g / mol, 1 ,760 g / mol, 1 ,770 g / mol, 1 ,780 g / mol, 1 ,790 g / mol, 1 ,800 g / mol, 1 ,810 g / mol, 1 ,820 g / mol, 1 ,830 g / mol, 1 ,840 g / mol, 1 ,850 g / mol, 1 ,860 g / mol, 1 ,870 g / mol, 1 ,880 g / mol, 1 ,890 g / mol, or 1 ,900 g / mol). In some embodiments, the poloxamer has an average ethylene oxide content of greater than 40% by mass (e.g., about 41 %, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51 %, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, or more).

[0161] In some embodiments, the poloxamer has an average ethylene oxide content of greater than 50% by mass (e.g., about 50%, 51 %, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, or more).

[0162] In some embodiments, the poloxamer has an average ethylene oxide content of greater than 60% by mass (e.g., about 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, or more).

[0163] In some embodiments, the poloxamer has an average ethylene oxide content of greater than 70% by mass (e.g., about 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, or more).

[0164] In some embodiments, the poloxamer has an average ethylene oxide content of from about 40% to about 90% (e.g., about 40%, 41 %, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51 %, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, or 90%).

[0165] In some embodiments, the poloxamer has an average ethylene oxide content of from about 70% to about 90% (e.g., about 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, or 90%).

[0166] In some embodiments, the poloxamer has an average molar mass of greater than 7,000 g / mol (e.g., about 7,100 g / mol, 7,200 g / mol, 7,300 g / mol, 7,400 g / mol, 7,500 g / mol, 7,600 g / mol, 7,700 g / mol, 7,800 g / mol, 7,900 g / mol, 8,000 g / mol, 8,100 g / mol, 8,200 g / mol, 8,300 g / mol, 8,400 g / mol, 8,500 g / mol, 8,600 g / mol, 8,700 g / mol, 8,800 g / mol, 8,900 g / mol, 9,000 g / mol, 9,100 g / mol, 9,200 g / mol, 9,300 g / mol, 9,400 g / mol, 9,500 g / mol, 9,600 g / mol, 9,700 g / mol, 9,800 g / mol, 9,900 g / mol, or 10,000 g / mol).

[0167] Poloxamer 188

[0168] Poloxamers that may be used in conjunction with the compositions and methods of the disclosure include “poloxamer 188” (also referred to in the art as “P 188”) having the approximate chemical formula HO(C2H4O)x(C3HeO)y(C2H4O)zH, wherein x and z are each independently about 80, and y is about 27. The average molecular weight of P188 is about 8,400 g / mol.

[0169] For clarity, the terms “average molar mass” and “average molecular weight” are used interchangeable herein to refer to the same quantity. The average molar mass, ethylene oxide content, and propylene oxide content of a poloxamer, as described herein, can be determined using methods disclosed in Alexandridis and Hatton, Colloids and Surfaces A: Physicochemical and Engineering Aspects 96:1-46 (1995), the disclosure of which is incorporated herein by reference in its entirety. Crystalline Form B

[0170] In some embodiments, the disclosure features a crystalline Form B, which is a solid form of compound (I) and Poloxamer 188, with a molar stoichiometry of approximately 100:1 compound (l):Poloxamer 188. In some embodiments, crystalline Form B is a co-crystal of compound (I) and Poloxamer 188. In some embodiments, crystalline Form B is a pseudopolymorph, having a crystal lattice of compound (I) incorporated with Poloxamer 188. In some embodiments, crystalline Form B is a complex of an anhydrous crystal lattice of compound (I), in association with Poloxamer 188. Crystalline Form B has one or more, or all, of the characteristic X-ray powder diffraction peaks as set forth in Table 1 , below. Crystalline Form B is characterized by one or more of the XRPD spectrum as shown in FIG. 5A, the DSC curve substantially as depicted in FIG. 5F, the1H NMR spectrum substantially as depicted in FIG. 5B, the TGA curve substantially as depicted in FIG. 5G, the DVS curve substantially as depicted in FIG. 5H, and the Raman curve substantially as depicted in FIG. 5I or FIG. 5J.

[0171] Table 1. Characteristic X-ray powder diffraction (XRPD) peaks of crystal Form B of compound (I)

[0172] Crystalline Form A

[0173] An alternative crystal form of compound (I) is crystalline Form A of compound (I), which is a solid form of anhydrous compound (I) having the characteristic X-ray powder diffraction peaks as set forth in Table 2, below. Crystalline Form A is characterized by one or more of the XRPD spectrum as shown in FIG. 1A, the DSC curve substantially as depicted in FIG. 1 B or FIG. 1C, the TGA curve substantially as depicted in FIG. 1 D or 1 E, and the DVS curve substantially as depicted in FIG. 1 F.

[0174] Table 2. Characteristic X-ray powder diffraction (XRPD) peaks of crystal Form A of compound (I)

[0175] As is described in Example 4, below, the inventors have shown that crystalline Form B is more stable and resistant to degradation in water and thermal stress than crystalline Form A, which enables storage of compound (I) for long-term usage. Compound (I) may be administered by a variety of routes, such as orally or intravenously. When formulated for oral administration, for instance, the compound may be administered in the form of a tablet, capsule, gel cap, powder, liquid solution, liquid suspension, granulate, microemulsion, or self-emulsifying dispersed system. Compound for Use

[0176] In another aspect, the disclosure provides compound (I) (e.g., a crystalline form of compound (I) described herein) for use in any of the methods described herein. For example, the disclosure features a crystalline form of compound (I) described herein for use in a method of treating a subject undergoing an embryo transfer procedure, reducing the likelihood of embryo implantation failure in a subject undergoing an embryo transfer procedure, improving endometrial receptivity in a subject undergoing an embryo transfer procedure, and / or reducing uterine contractility in a subject undergoing an embryo transfer procedure. The method may feature, for example, any one or more of the method steps recited herein.

[0177] Medicament

[0178] In still another aspect, the disclosure provides the use of compound (I) (e.g., a crystalline form of compound (I) described herein) in the manufacture of a medicament for performing any of the methods described herein. For example, the disclosure features the use of a crystalline form of compound (I) described herein in the manufacture of a medicament for use in a method of treating a subject undergoing an embryo transfer procedure, reducing the likelihood of embryo implantation failure in a subject undergoing an embryo transfer procedure, improving endometrial receptivity in a subject undergoing an embryo transfer procedure, and / or reducing uterine contractility in a subject undergoing an embryo transfer procedure. The method may feature, for example, any one or more of the method steps recited herein.

[0179] Examples

[0180] The following examples are put forth so as to provide those of ordinary skill in the art with a description of how the compositions and methods described herein may be used, made, and evaluated, and are intended to be purely exemplary of the disclosure and are not intended to limit the scope of what the inventors regard as their disclosure.

[0181] Example 1. Preparation and characterization of crystalline Form A of compound (I) Objective

[0182] The present Example is directed towards preparation and characterization of crystalline Form A of compound (I).

[0183] Methods

[0184] A series of experiments were conducted before successful crystallization was achieved. The failed crystallization attempts included use of acetone, acetonitrile, dichloromethane, 1 ,4-dioxane, ethanol, hexanes, isopropyl alcohol, methanol, methyl ethyl ketone, trifluoroethanol, tetrahydrofurane, Toluene and water, among other solvents. After each of these experiments, the resulting solid was tested and XRPD analysis showed the compound to be in the amorphous form.

[0185] Preparation of crystalline Form A was achieved by crystallization from diethyl ether or ethyl acetate, as described below. Results

[0186] Crystallization using Ethyl Acetate

[0187] Ethyl acetate was added to amorphous compound (I). The suspension was sonicated in ambient bath for 60 minutes to provide a clear solution that was further sonicated and cooled over 10 minutes. The clear solution was placed in the freezer for 4 days. The solvent was decanted and the solid was washed with hexane, then dried under nitrogen flow for 30 minutes. The resulting solid was analyzed by XRPD and confirmed to be crystalline compound (I). FIGS. 1C and 1E are the DSC and TGA curves, respectively, obtained from testing this sample. Characterization data is presented in Table 3.

[0188] Table 3. Characterization data of crystalline Form A from ethyl acetate

[0189] Crystallization using Diethyl Ether

[0190] Diethyl ether (40 mL) was added to amorphous compound (I) (4.0g). The solid was dissolved by swirling and the clear solution was left to stand at 18-23°C. Spontaneous nucleation occurred after ca. 10-15 minutes and the crystallization vessel and contents were allowed to stand undisturbed for 20 hours at 18-23°C. The crystals were isolated by filtration and dried under nitrogen for 4 hours at 18- 23°C. Yield 3.38g (84%), white crystals. Chemical purity 99.43%.

[0191] The resulting solid was analyzed by XRPD . FIGS. 1B and 1D are the DSC and TGA curves, respectively, obtained from testing this sample. Characterization data is presented in Table 4. Table 4. Characterization data of crystalline Form A from diethyl ether

[0192] Other Crystallization attempts using Diethyl Ether

[0193] Numerous attempts at preparing the crystalline compound using diethyl ether were performed, as set forth in Table 5. A representative DVS curve for crystalline Form A prepared from diethyl ether is shown in FIG. 1F.

[0194] Table 5. Crystalline preparation attempts from amorphous compound (I) using diethyl ether

[0195] a. RT=ambient temperature; vac=vacuum; P-N=Paratone-N (non-GMP). Reported times and temperatures are approximate; temperatures measured by NIST-traceable thermometer. b. B=birefringence; E=extinction; singles=sample contains crystal(s) which appeared suitable for submission for single crystal x-ray.

[0196] One such preparation involved adding 0.897 ml of diethyl ether to 240 mg of amorphous compound (I) and the mixture was left at ambient temperature for about 24 hours. The liquid was decanted and the solid transferred to a dry container to be air dried for 5 hours. The solid was then vacuum-dried for 20 hours at ambient temperature. The resulting solid was analyzed by XRPD and confirmed to be the crystalline compound. The XRPD pattern from this sample is shown in FIG. 1A.

[0197] 23 mg of the solid was then further vacuum-dried at 45°C for an additional 65 hours. The resulting solid was analyzed by XRPD and confirmed to be the crystalline compound. Characterization data is presented in Table 6.

[0198] Table 6. Characterization data of crystalline Form A from diethyl ether a. Temperatures rounded to nearest whole degree.

[0199] Example 2. Discovery of a novel crystalline form, Form B, of compound (I) Objective

[0200] The present Example is directed towards the screening and discovery of a novel crystalline form of compound (I).

[0201] Methods

[0202] Fast Evaporation (FE)

[0203] Clear liquid solutions of compound (I) or compound (I) and excipient were prepared in various solvents. Vials were left uncapped and solvent evaporated at ambient conditions.

[0204] Grinding

[0205] Compound (I) and excipients were ground with mortar and pestle for 2 to 5 minutes. Resulting solids were transferred to clean vials and in some cases underwent further purification by aqueous slurrying.

[0206] Heating Experiments

[0207] Compound (I) and excipient were placed on a glass slide together and melted at 90 °C or 120 °C. Slides were held at temperature for given period of time and then slowly cooled until solids were observed.

[0208] Milling

[0209] A drop of solvent was added to compound (I) in an agate milling jar with an agate ball. Jars were milled at 30 Hz for 3 10-minute cycles. The sides of the jars were scraped down in between cycles.

[0210] Slow Cooling (SC)

[0211] Concentrated solutions of compound (I) or compound (I) and excipient were prepared in various solvents at room temperature with stirring. Vials were capped cooled in the refrigerator (2 to 8 °C) and then further cooled in the freezer (~ -20 °C). If no solids were present, additional crystallization techniques were employed.

[0212] Slow Evaporation (SE)

[0213] Solutions of compound (I) and excipient were prepared in various solvents. The vial was capped with aluminum foil perforated with 1 -pin hole and solvent evaporated at ambient conditions.

[0214] Spontaneous Precipitation

[0215] A mixture of compound (I) in Et2O was sonicated. A clear solution was briefly formed followed by precipitation of white solids. Solids stirred in a slurry for 1 day at ambient conditions. Solids were collected by positive pressure filtration on a 0.2-pm nylon filter and flushed with 20 mL of air 5 times.

[0216] Slurry Experiments

[0217] Saturated solutions of crystalline Form B were prepared in water to remove excess Poloxamer 188. Mixtures were stirred at ambient temperature for the noted duration of time. Solids were collected by the stated technique.

[0218] Vapor Diffusion (VD)

[0219] Saturated solutions of compound (I) and excipient were prepared in a variety of solvents in 1- dram vials. The 1-dram vials were left uncapped and placed in a 20-mL vial containing a given antisolvent. The larger vial was capped and left at ambient temperature for a given amount of time. Any solids were collected, typically by decanting and drying under N2.

[0220] Vapor Stressing (VS)

[0221] A gel of compound (I) and Poloxamer 188 that resulted from another experiment was placed inside 20-mL vials containing water. The small vial was left uncapped and the larger vial was capped to allow vapor stressing to occur at the ambient temperature. No solids were isolated. X-ray Powder Diffraction (XRPD)

[0222] For transmission, XRPD pattern was collected with a PANalytical X'Pert PRO MPD or PANalytical Empyrean diffractometer using an incident beam of Cu radiation produced using a long, fine-focus source. An elliptically graded multilayer mirror was used to focus Cu Ka X-rays through the specimen and onto the detector. Prior to the analysis, a silicon specimen (NIST SRM 640e) was analyzed to verify the observed position of the Si 111 peak is consistent with the NIST-certified position. A specimen of the sample was sandwiched between 3-pm-thick films and analyzed in transmission geometry. A beam-stop, short antiscatter extension, and antiscatter knife edge were used to minimize the background generated by air. Soller slits for the incident and diffracted beams were used to minimize broadening and asymmetry from axial divergence. Diffraction patterns were collected using a scanning position-sensitive detector (X'Celerator) located 240 mm from the specimen and Data Collector software v. 2.2b or 5.5.

[0223] For reflection, XRPD patterns were collected with a PANalytical X'Pert PRO MPD diffractometer using an incident beam of Cu Ka radiation produced using a long, fine-focus source and a nickel filter. The diffractometer was configured using the symmetric Bragg-Brentano geometry. Prior to the analysis, a silicon specimen (NIST SRM 640e) was analyzed to verify the observed position of the Si 111 peak is consistent with the NIST-certified position. A specimen of the sample was prepared as a thin, circular layer centered on a silicon zero-background substrate. Antiscatter slits (SS) were used to minimize the background generated by air. Soller slits for the incident and diffracted beams were used to minimize broadening from axial divergence. Diffraction patterns were collected using a scanning position-sensitive detector (X'Celerator) located 240 mm from the sample and Data Collector software v. 2.2b.

[0224] Results

[0225] Crystallization studies were conducted to identify a novel crystalline form of compound (I). Attempts to crystallize neat compound (I) from ethanol or aqueous ethanol were unsuccessful (Table 7 and FIGS. 2A-2B). In addition to grinding and evaporation, cooling conditions and attempted spontaneous precipitation were also employed.

[0226] Next, amorphous compound (I) was combined with various excipients. The mixtures were evaporated from ethanol or ground with a small amount of aqueous ethanol with a mortar and pestle. Generated materials were analyzed by XRPD, comparing the resulting XRPD patterns to known patterns of compound (I) and the excipients. Attempts to crystallize compound (I) in the presence of calcium silicate, glyceryl dibehenate, povidone, lactose monohydrate, magnesium stearate, microcrystalline cellulose, saccharin sodium dihydrate, or sodium croscarmellose were also unsuccessful (Table 7 and FIGS. 2C-2P). XRPD patterns of the resulting materials were consistent with the excipient or mixtures of amorphous compound (I) and the excipient. Table 7. Attempts to crystallize compound (I) in the presence of various excipients

[0227] SC= slow cooling; FE= fast evaporation; RT= room temperature

[0228] All experiments utilizing compound (I) with Poloxamer 188 co-crystallized, producing crystalline

[0229] Form B. An exemplary XRPD overlay of the crystalline Form B and Form A is shown in FIG. 3A. Attempts with 9, 16, 19, 23, and 50 weight % Poloxamer 188 are summarized in Table 8 and FIGS. 4A-4Q.

[0230] Typically, unless an excess amount of Poloxamer 188 was used, a mixture of crystalline Form B and amorphous compound (I) resulted. A representative XRPD pattern of the mixture is compared to Poloxamer 188 and amorphous compound (I) in FIG. 3B. Co-crystallization with Poloxamer 188 still occurred in the absence of ethanol; grinding experiments using either dry methanol or no solvent were both successful in providing crystalline Form B (Table 8).

[0231] The ability to crystallize Form B from ethanol, methanol, or water represents an important advantage over crystalline Form A. Crystalline Form A can only be crystallized from ethyl acetate or diethyl ether (Example 1), which are less environmentally friendly solvents than ethanol, methanol, or water which may be used to produce crystalline Form B. The ability of Form B to crystallize from more environmentally friendly solvents than Form A is a surprising advantage from the perspective of drugsubstance manufacturing processes. Table 8. Crystallization of compound (I) in the presence of various amounts of Poloxamer 188

[0232]

[0233] Example 3. Characterization of crystalline Form B of compound (I)

[0234] Objective

[0235] The present Example is directed towards the characterization of crystalline Form B of compound (I).

[0236] Methods

[0237] Crystalline Form B was generated by combining amorphous compound (I) with 50 wt%

[0238] Poloxamer 188 and ground with a small amount of aqueous ethanol with a mortar and pestle. A large amount of Poloxamer 188 was utilized in an attempt to co-crystallize the entire amount of compound (I). Any excess polymer was removed though multiple water washes.

[0239] Differential Scanning Calorimetry (DSC)

[0240] DSC was performed using a Mettler-Toledo DSC3+ differential scanning calorimeter. Temperature calibration was performed using octane, phenyl salicylate, indium, tin, and zinc. The sample was placed into a hermetically sealed aluminum DSC pan, the weight was accurately recorded, the lid was pierced, and the sample was inserted into the DSC cell. A weighed aluminum pan configured as the sample pan was placed on the reference side of the cell. The sample was analyzed from -30 °C to 250 °C at 10 °C / min.

[0241] Dynamic Vapor Sorption (DVS)

[0242] Dynamic vapor sorption (DVS) data were collected on a Surface Measurement System DVS Intrinsic instrument. Samples were not dried prior to analysis. Sorption and desorption data were collected over a range from 5% to 95% RH at 10% RH increments under a nitrogen purge. The equilibrium criterion used for analysis was less than 0.0100% weight change in 5 minutes with a maximum equilibration time of 3 hours. Data were not corrected for the initial moisture content of the samples.

[0243] Liquid-State Nuclear Magnetic Resonance (NMR)

[0244] The solution1H NMR spectra were acquired by Spectral Data Services of Champaign, IL. The samples were dissolved in DMSO-d6.

[0245] Thermogravimetric Analysis (TGA)

[0246] TG analysis was performed using a Mettler-Toledo TGA / DSC3+ analyzer. Temperature calibration was performed using calcium oxalate, indium, tin, and zinc. The sample was placed in an aluminum pan. The pan was hermetically sealed, the lid pierced, then inserted into the TG furnace. A weighed aluminum pan configured as the sample pan was placed on the reference platform. The furnace was heated under nitrogen. The samples was analyzed from 25 °C to 350 °C at 10 °C / min.

[0247] X-ray Powder Diffraction (XRPD)

[0248] For transmission, XRPD pattern was collected with a PANalytical X'Pert PRO MPD or PANalytical Empyrean diffractometer using an incident beam of Cu radiation produced using a long, fine-focus source. An elliptically graded multilayer mirror was used to focus Cu Ka X-rays through the specimen and onto the detector. Prior to the analysis, a silicon specimen (NIST SRM 640e) was analyzed to verify the observed position of the Si 111 peak is consistent with the NIST-certified position. A specimen of the sample was sandwiched between 3-pm-thick films and analyzed in transmission geometry. A beam-stop, short antiscatter extension, and antiscatter knife edge were used to minimize the background generated by air. Soller slits for the incident and diffracted beams were used to minimize broadening and asymmetry from axial divergence. Diffraction patterns were collected using a scanning position-sensitive detector (X'Celerator) located 240 mm from the specimen and Data Collector software v. 2.2b or 5.5.

[0249] For reflection, XRPD patterns were collected with a PANalytical X'Pert PRO MPD diffractometer using an incident beam of Cu Ka radiation produced using a long, fine-focus source and a nickel filter. The diffractometer was configured using the symmetric Bragg-Brentano geometry. Prior to the analysis, a silicon specimen (NIST SRM 640e) was analyzed to verify the observed position of the Si 111 peak is consistent with the NIST-certified position. A specimen of the sample was prepared as a thin, circular layer centered on a silicon zero-background substrate. Antiscatter slits (SS) were used to minimize the background generated by air. Soller slits for the incident and diffracted beams were used to minimize broadening from axial divergence. Diffraction patterns were collected using a scanning position-sensitive detector (X'Celerator) located 240 mm from the sample and Data Collector software v. 2.2b.

[0250] Results

[0251] A representative XRPD spectrum of crystalline Form B is shown in FIG. 5A. Indexing results for crystalline Form B and Form A are summarized in Table 9. The space group was determined to be P3i. Form B has a longer c-axis and a larger unit cell volume than Form A. This indicates a different molecule packing and is manifest in the observation that the two polymorphs have distinct XRPD patterns.

[0252] Table 9. Indexing results for crystalline Form B

[0253] A representative proton NMR spectrum of crystalline Form B is shown in FIGS. 5B-5E and is consistent with a cocrystal of compound (I) and Poloxamer 188. Calculations based on the integration of the Poloxamer 188 peaks suggest a molar stoichiometry of approximately 100:1 compound (I) / Poloxamer 188. Residual ethanol is not evident in the spectrum.

[0254] Representative thermograms of crystalline Form B are provided in FIGS. 5F-5G. The DSC curve (FIG. 5F) exhibits a likely melt endotherm with an onset of 99 °C, followed by a decomposition exotherm at 237 °C (peak maximum). Negligible weight loss from 36 °C to 160 °C is observed by TGA (0.03% weight loss when heated from 36 °C -160 °C) (FIG. 5G), consistent with unsolvated material.

[0255] The DVS isotherm indicates crystalline Form B exhibits limited hygroscopicity (FIG. 5H). A weight change of ~1 .7% was observed though the sorption / desorption cycle with no significant hysteresis. A weight gain of 1 .65% was observed when the relative humidity was increased from 5% to 95%. A weight loss of 1 .67% was observed when the relative humidity was decreased from 95% to 5%. The material recovered from the DVS experiment remained crystalline Form B by XRPD.

[0256] Representative Raman spectra of crystalline Form B are shown in in FIGS. 5I and 5J.

[0257] Example 4. Comparative stability of compound (I) polymorphs Objective

[0258] The present Example is directed towards analyzing the comparative stability of the various crystal forms that compound (I) may adopt.

[0259] Comparative solubility studies may be used to evaluate the stability of compound (I) polymorphs. A lower solubility is indicative of a higher thermodynamic stability. This coincides with the hypothesis that a lower stability crystalline form, having weaker lattice energy, is more easily solubilized and has a readily disrupted lattice structure. Thus, a polymorph having a lower solubility is likely to be able to withstand long-term storage without degradation.

[0260] To further characterize the relative stability of the various crystal forms that compound (I) may adopt, the intrinsic dissolution rates (IDR) may be measured, which is defined as the mass dissolved in unit time from a unit surface area. As is described in Wen et al., Cryst. Growth Des. 2019, 19, 5471-5476, the IDR is proportional to the equilibrium solubility of the crystal. Therefore, a crystal form having a higher IDR, and therefore a higher solubility, is indicative of a lower thermodynamic stability. Accordingly, a crystalline form having a lower IDR is desirable for achieving superior stability.

[0261] Another comparison of the relative stabilities of the various crystal forms that compound (I) may adopt may be determined by investigating whether the integrity of the crystal lattice may be perturbed from prolonged exposure to water. For example, following solubility studies or IDR studies, the remaining excess solids may be isolated and analyzed by XRPD to determine whether the extended exposure to water has altered the crystal form. An unchanged XRPD spectrum following exposure to water indicates that the crystal form may be stored for long-term usage without being susceptible to water-induced degradation.

[0262] Additionally, the relative stabilities of the various crystal forms that compound (I) may adopt may be assessed by comparing their susceptibility to hysteresis upon changes in humidity using dynamic vapor sorption (DVS) experiments. Hysteresis, which is a difference in water vapor uptake between sorption (increase in humidity) and desorption (decrease in humidity) cycles is indicative of structural changes upon exposure to water.

[0263] A further comparison of the relative stabilities of the various crystal forms that compound (I) may adopt may be determined through differential scanning calorimetry (DSC) experiments. DSC permits the determination of the melting point of a crystal form, with a higher melting point indicative of higher thermal stability.

[0264] Methods

[0265] XRPD

[0266] XRPD patterns were collected with a PANalytical X'Pert PRO MPD diffractometer using an incident beam of Cu radiation produced using an Optix long, fine-focus source. An elliptically graded multilayer mirror was used to focus Cu Ka X-rays through the specimen and onto the detector. Prior to the analysis, a silicon specimen (NIST SRM 640e) was analyzed to verify the observed position of the Si 111 peak is consistent with the NIST-certified position. A specimen of the sample was sandwiched between 3- -thick films and analyzed in transmission geometry. A beam-stop, short antiscatter extension, and antiscatter knife edge were used to minimize the background generated by air. Soller slits for the incident and diffracted beams were used to minimize broadening from axial divergence. Diffraction patterns were collected using a scanning position-sensitive detector (X'Celerator) located 240 mm from the specimen and Data Collector software v. 5.5.

[0267] Pellet Preparation and Compression Study

[0268] For each pellet, about 150 mg of the specified compound (I) sample was weighed into the die cavity of a woods apparatus and then compressed for ~ 1 minute using an applied load of approximately 1000 PSI to form a pellet.

[0269] An XRPD study was performed to evaluate the physical stability of the three forms of compound (I) after pelletization. Pressed pellets of crystalline Form B, crystalline Form A, and amorphous compound (I) were analyzed by XRPD before intrinsic dissolution testing. The resulting XRPD patterns showed that no form change occurred during the compression when compared to the patterns of the original materials.

[0270] XRPD patterns were collected with a PANalytical X'Pert PRO MPD diffractometer using an incident beam of Cu K radiation produced using a long, fine-focus source and a nickel filter. The diffractometer was configured using the symmetric Bragg-Brentano geometry. Prior to the analysis, a silicon specimen (NIST SRM 640e) was analyzed to verify the observed position of the Si 111 peak is consistent with the NIST-certified position. The specimen was put in at the center of a backfill holder. Antiscatter slits (SS) were used to minimize the background generated by air. Soller slits for the incident and diffracted beams were used to minimize broadening from axial divergence. Diffraction patterns were collected using a scanning position- sensitive detector (X'Celerator) located 240 mm from the sample and Data Collector software v.5.5.

[0271] Solubility

[0272] Solubility for amorphous compound (I), crystalline Form A, and crystalline Form B was evaluated in water in duplicate at 24 hours. Amounts of samples resulting in a 5 mg / mL loading were dissolved in water and stirred at 25°C. At the end of the 24-hour stirring, slurries were centrifuged and the supernatant was analyzed by LC.

[0273] Aqueous media solubility for crystalline Form B was performed in duplicate at 24 hours. Amounts of crystalline Form B resulting in a 20 mg / mL loading were dissolved in 4 different media (pH 1 , 3, 5, and 7 buffers) and stirred at 25°C. At the end of the 24 hour stirring period, slurries were filtered using a PVDF syringe filter and the supernatant was analyzed by LC.

[0274] Aqueous media solubility for crystalline Form B was also performed in duplicate at 24 hours using centrifugation instead of filtration. Crystalline Form B resulting in a 5 mg / mL loading were dissolved in 4 different media (pH 1 , 3, 5, and 7 buffers) and stirred at 25°C. At the end of the 24-hour stirring, slurries were centrifuged and the supernatant was analyzed by LC.

[0275] Solubility of crystalline Form A and amorphous compound (I) was also evaluated in pH 1 , 3, 5, and 7 buffer. Suspensions of crystalline Form A and amorphous compound (I) were prepared by adding about 30 mg material to 3 mL of media. Samples were stirred for 24 hours at 25.0 °C. At the end of 24 hours of stirring, the suspensions were centrifuged and the clear supernatants were analyzed for concentration by HPLC.

[0276] Intrinsic Dissolution

[0277] Intrinsic dissolution experiments were performed using a VanKel VK7010 dissolution tester equipped with a VK750D heater / circulator. A Wood's Apparatus (0.50 cm2sample surface area) as described in USP <1087> was used. Approximately 150 mg of AGIO HCI was compressed with an applied load of approximately 1000 pounds for 1 minute in the Wood's apparatus using a hydraulic press. The intrinsic dissolution medium was pH 7 Phosphate Buffer or water. The medium (500 mL) was equilibrated to 37 °C ± 0.5° C. The disks were rotated at 100 rpm. Sampling of 1 mL aliquots was performed at the given time points of 5, 10, 15, 30, 60, 90, 120, and 180 minutes using a 3 mL syringe equipped with a stainless steel cannula.

[0278] DVS

[0279] DVS experiments for crystalline Form A were performed according to the materials and methods described in Example 1 . DVS experiments for crystalline Form B were performed according to the materials and methods described in Example 3. DSC

[0280] DSC was performed using a Mettler-Toledo DSC3+ differential scanning calorimeter. Temperature calibration was performed using octane, phenyl salicylate, indium, tin, and zinc. The sample was placed into a hermetically sealed aluminum DSC pan, the weight was accurately recorded, the lid was pierced, and the sample was inserted into the DSC cell. A weighed aluminum pan configured as the sample pan was placed on the reference side of the cell. The sample was analyzed from -30 °C to 250 °C at 10 °C / min.

[0281] Results

[0282] Solubility in Water

[0283] Solubility for the amorphous compound (I), crystalline Form A, and crystalline Form B was evaluated in water in duplicate at 24 hours. Amounts of samples resulting in a 5 mg / mL loading were dissolved in water and stirred at 25°C. At the end of the 24-hour stirring, slurries were centrifuged and the supernatant was analyzed by LC. Results are shown in Table 10. Surprisingly, crystalline Form B was less soluble than Form A and the amorphous form of compound (I). The lower solubility observed with crystalline Form B is advantageous because it can better withstand exposure to water without loss of lattice integrity, enabling long-term shelf storage.

[0284] Table 10. Solubility observations after 24 hours of stirring in water

[0285] *Compound (I) in Crystalline Form B solubility is calculated based on an LC determination of ~84.8% compound (I) and ~15.2% Poloxamer 188 by weight in the Form B co-crystal.

[0286] Aqueous pH Solubility

[0287] Aqueous media solubility in 4 different media (pH 1 , 3, 5, and 7 buffers) after 24 hours stirring at 25°C followed by centrifugation was performed for crystalline Form B (FIG. 6, Table 11), as well as amorphous compound (I) and crystalline Form A (Table 12). Consistent with the above solubility results in water, crystalline Form B was less soluble in pH 1 , 3, 5, and 7 buffers than Form A and the amorphous form of compound (I), indicating that Form B is more stable.

[0288] Table 11. Aqueous solubility data for crystalline Form B (centrifugation)

[0289] *Compound (I) in Crystalline Form B solubility is calculated based on an LC determination of ~89.5% compound (I) and ~10.5% Poloxamer 188 by weight in the Form B co-crystal.

[0290] Table 12. Aqueous solubility data for amorphous compound (I) and crystalline Form A (centrifugation) Aqueous media solubility for crystalline Form B was also performed in duplicate at 24 hours using filtration. Amounts of crystalline Form B resulting in a 20 mg / mL loading were dissolved in 4 different media (pH 1 , 3, 5, and 7 buffers) and stirred at 25°C. At the end of the 24 hour stirring period, slurries were filtered using a PVDF syringe filter and the supernatant was analyzed by LC. Results are shown in Table 13.

[0291] Table 13. Aqueous Solubility Data for Crystalline Form B (Filtration)

[0292] IDR in Water

[0293] An intrinsic dissolution experiment was performed using the crystalline Form B, crystalline Form A, and amorphous compound (I) pellets to determine the intrinsic dissolution rate (IDR). The experiments were run in 500 mL of water at 37°C with a rotation speed of 100 rpm. Sampling was done by pulling a 1 mL aliquot from the vessel and transferring it to HPLC vials at each given sampling point. Following intrinsic dissolution, the pulled samples were analyzed by HPLC. The slope of the line corresponds to the IDR (Tables 14-16). Table 14. Intrinsic dissolution of crystalline Form B in water.

[0294]

[0295] Table 15. Intrinsic dissolution of crystalline Form A in water.

[0296] Table 16. Intrinsic dissolution of the amorphous form in water.

[0297] The average IDR of crystalline Form B was 0.013 mg / cm2 / min (Table 14), the IDR of the crystalline Form A was 0.034 mg / cm2 / min (Table 15), and the average IDR of the amorphous Form was 0.032 mg / cm2 / min (Table 16). The IDR of crystalline Form B is lower than the rates of the crystalline Form A and amorphous material. An overlay of another IDR experiment following the same methods set forth above is shown in FIG. 7A.

[0298] The observation that crystalline Form B exhibits a lower IDR compared to Form A and amorphous compound (I) indicates that crystalline Form B is more stable. The superior ability of Form B to withstand elevated moisture levels is highly advantageous, enabling the long-term shelf storage of compound (I) without degradation of the crystal lattice due to water.

[0299] IDR in pH 7 Phosphate Buffer

[0300] Intrinsic dissolution experiments were performed in duplicate using crystalline Form B pellets to determine the IDR. The experiment was run in 500 mL of a pH 7 0.2 M phosphate buffer at 37°C with a rotation speed of 100 rpm. Sampling was done by pulling a 1 mL aliquot from the vessel and transferring it to HPLC vials at each given sampling point. Following intrinsic dissolution, the pulled samples were analyzed by HPLC. The average IDR of the crystalline Form B replicates was 0.011 mg / cm2 / min (Table 17). A similar experiment was performed for crystalline Form A and amorphous compound (I) (FIG.

[0301] 7B). The IDR of crystalline Form B was lower than the IDR of crystalline Form A (0.027 mg / cm2 / min) and amorphous material (0.031 mg / cm2 / min). As discussed above, the lower IDR of crystalline Form B imparts the crystal form with important benefits for shelf-life storage.

[0302] Table 17. IDR of crystalline Form B in Buffer at pH 7.

[0303] *Compound (I) is calculated based on an LC determination of ~87.8% compound (I) and ~12.2% Poloxamer 188 by weight in the Form B co-crystal.

[0304] **Compound (I) is calculated based on an LC determination of ~84.8% compound (I) and ~15.2% Poloxamer 188 by weight in the Form B co-crystal.

[0305] Form change following extended exposure to water

[0306] As another measure of stability, remaining excess solids following completion of the above solubility and IDR experiments were isolated and analyzed by XRPD to determine whether the extended exposure to water compromised the integrity of the crystal forms.

[0307] Post solubility and IDR solids did not show a form change for the amorphous material (FIGS. 8A- 8C). Exposure to water triggered a form change in crystalline Form A, as evidenced by the post solubility and post IDR diffractogram for the crystalline Form A, which showed a mixture of crystalline Form A and amorphous compound (I) (FIGS. 8D-8G).

[0308] Notably, no form change was observed for crystalline Form B following solubility experiments or IDR experiments (FIGS. 8H-8J). The surprising ability of crystalline Form B to resist degradation in water demonstrates that Form B has superior stability compared to Form A.

[0309] Hysteresis under vapor sorption / desorption cycles

[0310] The DVS isotherm of crystalline Form B is provided in FIG. 5H, and the DVS isotherm of crystalline Form A is provided in FIG. 1 F. Crystalline Form A exhibited hysteresis, as evidenced by the non-overlapping sorption and desorption curves. Surprisingly, no significant hysteresis was observed for crystalline Form B. These results show that, although crystalline Form B appears to absorb water to a higher degree than Form A, Form B has a superior ability to resist water-induced degradation of the crystal lattice, which is indicative of greater stability.

[0311] Thermal stability

[0312] The DSC curve exhibits a likely melt endotherm with an onset of 99 °C, followed by a decomposition exotherm at 237 °C (peak maximum). The melting point of Form B (99 °C) is higher than the melting point of Form A (75°C) (FIG. 9), demonstrating that crystalline Form B is more stable than Form A since more heat is required to compromise the integrity of the crystal lattice.

[0313] Conclusion

[0314] The solubility, IDR, form change, and hysteresis results described above demonstrate that crystalline Form B has a superior ability to withstand exposure to water for prolonged periods of time compared to crystalline Form A and amorphous compound (I). Form B’s surprising stability enables the crystal form to be stored for long-term usage, maintaining the integrity of the crystal lattice without water- induced degradation.

[0315] The DSC results further demonstrate that crystalline Form B has a superior ability to withstand high temperatures than Form A, which is indicative of a higher stability of the crystal lattice.

[0316] Other Embodiments

[0317] All publications, patents, and patent applications mentioned in this specification are incorporated herein by reference to the same extent as if each independent publication or patent application was specifically and individually indicated to be incorporated by reference.

[0318] While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the invention that come within known or customary practice within the art to which the invention pertains and may be applied to the essential features hereinbefore set forth, and follows in the scope of the claims.

[0319] Some embodiments of the technology described herein can be defined according to any of the following numbered embodiments:

[0320] E1. A crystalline Form B of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) wherein the crystalline Form B of the compound exhibits characteristic X-ray powder diffraction (XRPD) peaks at 12.86 ± 0.20° 26, 16.32 ± 0.20° 26, and 22.69 ± 6.26° 26.

[0321] E2. The crystalline Form B of E1 , wherein the crystalline Form B of said compound exhibits characteristic XRPD peaks at 17.81 ± 6.26° 26, 21 .69 ± 6.26° 26, and 24.86 ± 6.26° 26. E3. The crystalline Form B of E1 or E2, wherein the crystalline Form B of said compound exhibits characteristic XRPD peaks at 23.94 ± 0.20° 20 and 27.93 ± 0.20° 20.

[0322] E4. The crystalline Form B of any one of E1 -E3, wherein the crystalline Form B of said compound exhibits characteristic XRPD peaks at 25.87 ± 0.20° 20, 26.99 ± 0.20° 20, and 28.74 ± 0.20° 20.

[0323] E5. The crystalline Form B of any one of E1 -E4, wherein the crystalline Form B of said compound exhibits characteristic XRPD peaks at 7.07 ± 0.20° 20, 12.27 ± 0.20° 20, 14.17 ± 0.20° 20, and 18.79 ± 0.20° 20 .

[0324] E6. The crystalline Form B of any one of E1 -E5, wherein the crystalline Form B of said compound exhibits characteristic XRPD peaks at 21 .34 ± 0.20° 20 and 25.72 ± 0.20° 20.

[0325] E7. The crystalline Form B of any one of E1 -E6, wherein the crystalline Form B of said compound exhibits characteristic XRPD peaks at 30.56 ± 0.20° 20, 31 .22 ± 0.20° 20, 32.56 ± 0.20° 20, 33.03 ± 0.20° 20, 33.35 ± 0.20° 20, 33.79 ± 0.20° 20, 34.67 ± 0.20° 20, 34.90 ± 0.20° 20, 35.67 ± 0.20° 20, 36.08 ± 0.20° 20, 37.42 ± 0.20° 20, 37.84 ± 0.20° 20, 38.25 ± 0.20° 20, and 39.72 ± 0.20° 20.

[0326] E8. The crystalline Form B of any one of E1-E7, having the XRPD spectrum as shown in FIG. 5A.

[0327] E9. The crystalline Form B of any one of E1-E8, wherein the XRPD peaks at diffraction angle 20 (°) are collected by irradiating with Cu Ka.

[0328] E10. The crystalline Form B of any one of E1-E9, having a differential scanning calorimetry (DSC) endotherm at about 99 °C.

[0329] E11 . The crystalline Form B of any one of E1 -E10, having the DSC curve substantially as depicted in FIG. 5F.

[0330] E12. The crystalline Form B of any one of E1-E11 , exhibiting1H nuclear magnetic resonance (NMR) peaks centered at a chemical shift (6) of about 1 .0 ppm, about 2.3 ppm, about 2.5 ppm, about 3.3 ppm, about 3.5 ppm, about 3.7 ppm, about 7.2 ppm, about 7.3 ppm, about 7.4 ppm, and about 7.6 ppm.

[0331] E13. The crystalline Form B of any one of E1-E12, wherein the crystalline Form B is characterized by a 1H NMR spectrum substantially as depicted in FIG. 5B.

[0332] E14. The crystalline Form B of any one of E1-E13, wherein the crystalline Form B exhibits a weight loss of from 0.01 % to 5% when heated from 36 °C to 160 °C as measured by thermogravimetric analysis (TGA), optionally wherein the weight loss is from 0.01 % to 0.05%, further optionally wherein the weight loss is about 0.03%.

[0333] E15. The crystalline Form B of E14, wherein the crystalline Form B exhibits a weight loss of about 0.03% when heated from 36 °C to 160 °C as measured by TGA.

[0334] E16. The crystalline Form B of any one of E1-E15, wherein the crystalline Form B exhibits a TGA curve substantially as depicted in FIG. 5G.

[0335] E17. The crystalline Form B of any one of E1-E16, wherein the crystalline Form B exhibits a weight gain of from 0.01 % to 5% when the relative humidity is increased from 5% to 95% as measured by dynamic vapor sorption (DVS) , optionally wherein the weight gain is from 1% to 5%, further optionally wherein the weight gain is about 1 .65%.

[0336] E18. The crystalline Form B of E17, wherein the crystalline Form B exhibits a weight gain of about 1 .65% when the relative humidity is increased from 5% to 95% as measured by DVS.

[0337] E19. The crystalline Form B of any one of E1-E18, wherein the crystalline Form B exhibits a weight loss of from 0.01% to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS, optionally wherein the weight loss is from 1% to 5%, further optionally wherein the weight loss is about 1 .67%.

[0338] E20. The crystalline Form B of E19, wherein the crystalline Form B exhibits a weight loss of about 1 .67% when the relative humidity is decreased from 95% to 5% as measured by DVS.

[0339] E21 . The crystalline Form B of any one of E1-E20, wherein the crystalline Form B exhibits a DVS curve substantially as depicted in FIG. 5H.

[0340] E22. The crystalline Form B of any one of E1-E21 , wherein the crystalline Form B exhibits a Raman curve substantially as depicted in FIG. 5I or FIG. 5J.

[0341] E23. The crystalline Form B of any one of E1-E22, wherein the crystalline Form B has a solubility of from 0.100 mg / mL to 0.200 mg / mL in water after 24 hours of stirring.

[0342] E24. The crystalline Form B of E23, wherein the crystalline Form B has a solubility of about 0.189 mg / mL in water after 24 hours of stirring.

[0343] E25. The crystalline Form B of any one of E1-E24, wherein the crystalline Form B has an intrinsic dissolution rate (IDR) of from 0.010 mg / cm2 / min to 0.020 mg / cm2 / min in water.

[0344] E26. The crystalline Form B of E25, wherein the crystalline Form B has an IDR of about 0.013 mg / cm2 / min in water.

[0345] E27. The crystalline Form B of any one of E1-E26, wherein the crystalline Form B is obtainable by recrystallization from a solvent.

[0346] E28. The crystalline Form B of E27, wherein the crystalline Form B is obtainable by recrystallization from ethanol, methanol, or water.

[0347] E29. Crystalline Form B of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) wherein the crystalline Form B exhibits a weight loss of from 0.01% to 5% when heated from 36 °C to 160 °C as measured by TGA, optionally wherein the weight loss is from 0.01 % to 0.05%, further optionally wherein the weight loss is about 0.03%.

[0348] E30. Crystalline Form B of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) having (a) the XRPD spectrum as shown in FIG. 5A and optionally at least one of the following properties:

[0349] (b) a DSC endotherm at about 99 °C;

[0350] (c) a DSC substantially as depicted in FIG. 5F;

[0351] (d)1H NMR peaks centered at a chemical shift (6) of about 1 .0 ppm, about 2.3 ppm, about 2.5 ppm, about 3.3 ppm, about 3.5 ppm, about 3.7 ppm, about 7.2 ppm, about 7.3 ppm, about 7.4 ppm, and about 7.6 ppm;

[0352] (e) a1H NMR spectrum substantially as depicted in FIG. 5B;

[0353] (f) a weight loss of from 0.01% to 5% when heated from 36 °C to 160 °C as measured by TGA, optionally wherein the weight loss is from 0.01 % to 0.05%, further optionally wherein the weight loss is about 0.03%;

[0354] (g) a TGA curve substantially as depicted in FIG. 5G;

[0355] (h) a weight gain of from 0.01% to 5% when the relative humidity is increased from 5% to 95% as measured by DVS, optionally wherein the weight gain is from 1% to 5%, further optionally wherein the weight gain is about 1 .65%;

[0356] (i) a weight loss of from 0.01% to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS, optionally wherein the weight loss is from 1% to 5%, further optionally wherein the weight loss is about 1 .67%;

[0357] (j) a DVS curve substantially as depicted in FIG. 5H;

[0358] (k) a Raman curve substantially as depicted in FIG. 5I or FIG. 5J;

[0359] (l) a solubility of from 0.100 mg / mL to 0.200 mg / mL in water after 24 hours of stirring;

[0360] (m) an IDR of from 0.010 mg / cm2 / min to 0.020 mg / cm2 / min in water; or

[0361] (n) combinations thereof.

[0362] E31 . A crystalline Form B of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)

[0363] characterized by unit cell parameters substantially equal to the following cell dimensions: a = 14.410 A; b = 14.410 A; c = 8.247 A; a = 90 degrees; p = 90 degrees; y = 120 degrees; and Space group = P3i. E32. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 80% (e.g., about 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0364] E33. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 84% (e.g., about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0365] E34. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 87% (e.g., about 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0366] E35. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I) comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 89% (e.g., about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

[0367] E36. A pharmaceutical composition comprising the crystalline Form B of any one of E1-E31 or the solid form of any one of E32-E35, and one or more carriers, diluents, or excipients.

[0368] E37. A method of treating a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline Form B of any one of E1 -E31 , the solid form of any one of E32-E35, or the pharmaceutical composition of E36.

[0369] E38. A method of reducing the likelihood of embryo implantation failure in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline Form B of any one of E1-E31 , the solid form of any one of E32-E35, or the pharmaceutical composition of E36. E39. A method of improving endometrial receptivity in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline Form B of any one of E1-E31 , the solid form of any one of E32-E35, or the pharmaceutical composition of E36.

[0370] E40. A method of reducing uterine contractility in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline Form B of any one of E1 -E31 , the solid form of any one of E32-E35, or the pharmaceutical composition of E36.

[0371] E41 . The method of any one of E37-E40, wherein the method comprises transferring the one or more embryos to the uterus of the subject.

[0372] E42. The method of any one of E37-E41 , wherein the method further comprises inducing follicular maturation in the subject, optionally wherein the inducing of follicular maturation comprises administering human chorionic gonadotropin (hCG) to the subject.

[0373] E43. The method of any one of E37-E42, wherein from 1 to 2 embryos are transferred to the subject.

[0374] E44. The method of E43, wherein 1 embryo is transferred to the subject.

[0375] E45. The method of E43, wherein 2 embryos are transferred to the subject.

[0376] E46. The method of any one of E37-E45, wherein the subject is a mammal and the one or more embryos are mammalian embryos.

[0377] E47. The method of E46, wherein the mammal is a human and the one or more mammalian embryos are human embryos.

[0378] E48. The method of any one of E37-E47, wherein the one or more embryos are produced ex vivo by in vitro fertilization (IVF).

[0379] E49. The method of E48, wherein the one or more embryos are produced ex vivo by IVF of one or more ova derived from the subject.

[0380] E50. The method of any one of E37-E47, wherein the one or more embryos are produced ex vivo by intracytoplasmic sperm injection (ICSI).

[0381] E51 . The method of E50, wherein the one or more embryos are produced ex vivo by ICSI into one or more ova derived from the subject.

[0382] E52. The method of E49 or E51 , wherein the one or more ova are derived from one or more oocytes isolated from the subject.

[0383] E53. The method of E52, wherein the one or more oocytes comprise from 1 to 4 mature oocytes.

[0384] E54. The method of E52 or E53, wherein a gonadotropin-releasing hormone (GnRH) antagonist is administered to the subject prior to isolation of the one or more oocytes from the subject.

[0385] E55. The method of any one of E52-E54, wherein hCG is administered to the subject prior to isolation of the one or more oocytes from the subject.

[0386] E56. The method of any one of E52-E55, wherein progesterone is administered to the subject following isolation of the one or more oocytes from the subject. E57. The method of E49 or E51 , wherein the one or more ova are isolated directly from the subject.

[0387] E58. The method of E57, wherein a GnRH antagonist is administered to the subject prior to isolation of the one or more ova from the subject.

[0388] E59. The method of E57 or E58, wherein hCG is administered to the subject prior to isolation of the one or more ova from the subject.

[0389] E60. The method of any one of E57-E59, wherein progesterone is administered to the subject following isolation of the one or more ova from the subject.

[0390] E61 . The method of any one of E37-E60, wherein the one or more embryos each comprise from 6 to 8 blastomeres immediately prior to the transfer of the one or more embryos to the subject.

[0391] E62. The method of E61 , wherein the blastomeres are of approximately equal sizes as assessed by visual microscopy.

[0392] E63. A kit comprising the crystalline Form B of any one of E1 -E31 , the solid form of any one of E32- E35, or the pharmaceutical composition of E36, wherein the kit further comprises a package insert instructing a user of the kit to administer the crystalline Form B to a subject in accordance with the method of any one of E37-E62.

[0393] Other embodiments are within the claims.

Claims

CLAIMS1. A crystalline form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)wherein the crystalline form of the compound exhibits characteristic X-ray powder diffraction (XRPD) peaks at 12.86 ± 0.20° 26, 16.32 ± 0.20° 26, and 22.69 ± 6.26° 26.

2. The crystalline form of claim 1 , wherein the crystalline form of said compound exhibits characteristic XRPD peaks at 17.81 ± 6.26° 26, 21 .69 ± 6.26° 26, and 24.86 ± 6.26° 26.

3. The crystalline form of claim 1 or 2, wherein the crystalline form of said compound exhibits characteristic XRPD peaks at 23.94 ± 6.26° 26 and 27.93 ± 6.26° 26.

4. The crystalline form of any one of claims 1 -3, wherein the crystalline form of said compound exhibits characteristic XRPD peaks at 25.87 ± 6.26° 26, 26.99 ± 6.26° 26, and 28.74 ± 6.26° 26.

5. The crystalline form of any one of claims 1 -4, wherein the crystalline form of said compound exhibits characteristic XRPD peaks at 7.67 ± 6.26° 26, 12.27 ± 6.26° 26, 14.17 ± 6.26° 26, and 18.79 ± 6.26° 26 .

6. The crystalline form of any one of claims 1 -5, wherein the crystalline form of said compound exhibits characteristic XRPD peaks at 21 .34 ± 6.26° 26 and 25.72 ± 6.26° 26.

7. The crystalline form of any one of claims 1 -6, wherein the crystalline form of said compound exhibits characteristic XRPD peaks at 36.56 ± 6.26° 26, 31 .22 ± 6.26° 26, 32.56 ± 6.26° 26, 33.63 ± 6.26° 26, 33.35 ± 6.26° 26, 33.79 ± 6.26° 26, 34.67 ± 6.26° 26, 34.96 ± 6.26° 26, 35.67 ± 6.26° 26, 36.68 ± 6.26° 26, 37.42 ± 6.26° 26, 37.84 ± 6.26° 26, 38.25 ± 6.26° 26, and 39.72 ± 6.26° 26.

8. The crystalline form of any one of claims 1-7, having the XRPD spectrum as shown in FIG. 5A.

9. The crystalline form of any one of claims 1-8, wherein the XRPD peaks at diffraction angle 26 (°) are collected by irradiating with Cu Ka.

10. The crystalline form of any one of claims 1-9, having a differential scanning calorimetry (DSC) endotherm at about 99 °C.11 . The crystalline form of any one of claims 1-10, having the DSC curve substantially as depicted in FIG. 5F.

12. The crystalline form of any one of claims 1-11 , exhibiting1H nuclear magnetic resonance (NMR) peaks centered at a chemical shift (6) of about 1 .0 ppm, about 2.3 ppm, about 2.5 ppm, about 3.3 ppm, about 3.5 ppm, about 3.7 ppm, about 7.2 ppm, about 7.3 ppm, about 7.4 ppm, and about 7.6 ppm.

13. The crystalline form of any one of claims 1-12, wherein the crystalline form is characterized by a1H NMR spectrum substantially as depicted in FIG. 5B.

14. The crystalline form of any one of claims 1-13, wherein the crystalline form exhibits a weight loss of from 0.01 % to 5% when heated from 36 °C to 160 °C as measured by thermogravimetric analysis (TGA), optionally wherein the weight loss is from 0.01 % to 0.05%, further optionally wherein the weight loss is about 0.03%.

15. The crystalline form of claim 14, wherein the crystalline form exhibits a weight loss of about 0.03% when heated from 36 °C to 160 °C as measured by TGA.

16. The crystalline form of any one of claims 1-15, wherein the crystalline form exhibits a TGA curve substantially as depicted in FIG. 5G.

17. The crystalline form of any one of claims 1-16, wherein the crystalline form exhibits a weight gain of from 0.01 % to 5% when the relative humidity is increased from 5% to 95% as measured by dynamic vapor sorption (DVS), optionally wherein the weight gain is from 1 % to 5%, further optionally wherein the weight gain is about 1 .65%.

18. The crystalline form of claim 17, wherein the crystalline form exhibits a weight gain of about 1 .65% when the relative humidity is increased from 5% to 95% as measured by DVS.

19. The crystalline form of any one of claims 1-18, wherein the crystalline form exhibits a weight loss of from 0.01 % to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS, optionally wherein the weight loss is from 1 % to 5%, further optionally wherein the weight loss is about 1.67%.

20. The crystalline form of claim 19, wherein the crystalline form exhibits a weight loss of about 1 .67% when the relative humidity is decreased from 95% to 5% as measured by DVS.21 . The crystalline form of any one of claims 1 -20, wherein the crystalline form exhibits a DVS curve substantially as depicted in FIG. 5H.

22. The crystalline form of any one of claims 1 -21 , wherein the crystalline form exhibits a Raman curve substantially as depicted in FIG. 5I or FIG. 5J.

23. The crystalline form of any one of claims 1-22, wherein the crystalline form has a solubility of from 0.100 mg / mL to 0.200 mg / mL in water after 24 hours of stirring.

24. The crystalline form of claim 23, wherein the crystalline form has a solubility of about 0.189 mg / mL in water after 24 hours of stirring.

25. The crystalline form of any one of claims 1-24, wherein the crystalline form has an intrinsic dissolution rate (IDR) of from 0.010 mg / cm2 / min to 0.020 mg / cm2 / min in water.

26. The crystalline form of claim 25, wherein the crystalline form has an IDR of about 0.013 mg / cm2 / min in water.

27. The crystalline form of any one of claims 1 -26, wherein the crystalline form is obtainable by recrystallization from a solvent.

28. The crystalline form of claim 27, wherein the crystalline form is obtainable by recrystallization from ethanol, methanol, or water.

29. Crystalline form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)wherein the crystalline form exhibits a weight loss of from 0.01% to 5% when heated from 36 °C to 160 °C as measured by TGA, optionally wherein the weight loss is from 0.01% to 0.05%, further optionally wherein the weight loss is about 0.03% when heated from 36 °C to 160 °C as measured by TGA.

30. Crystalline form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)having (a) the XRPD spectrum as shown in FIG. 5A and optionally at least one of the following properties:(b) a DSC endotherm at about 99 °C;(c) a DSC substantially as depicted in FIG. 5F;(d)1H NMR peaks centered at a chemical shift (6) of about 1 .0 ppm, about 2.3 ppm, about 2.5 ppm, about 3.3 ppm, about 3.5 ppm, about 3.7 ppm, about 7.2 ppm, about 7.3 ppm, about 7.4 ppm, and about 7.6 ppm;(e) a1H NMR spectrum substantially as depicted in FIG. 5B;(f) a weight loss of from 0.01 % to 5% when heated from 36 °C to 160 °C as measured by TGA, optionally wherein the weight loss is from 0.01 % to 0.05%, further optionally wherein the weight loss is about 0.03%;(g) a TGA curve substantially as depicted in FIG. 5G;(h) a weight gain of from 0.01 % to 5% when the relative humidity is increased from 5% to 95% as measured by DVS, optionally wherein the weight gain is from 1 % to 5%, further optionally wherein the weight gain is about 1 .65%;(i) a weight loss of from 0.01 % to 5% when the relative humidity is decreased from 95% to 5% as measured by DVS, optionally wherein the weight loss is from 1 % to 5%, further optionally wherein the weight loss is about 1 .67%;(j) a DVS curve substantially as depicted in FIG. 5H;(k) a Raman curve substantially as depicted in FIG. 5I or FIG. 5J;(l) a solubility of from 0.100 mg / mL to 0.200 mg / mL in water after 24 hours of stirring;(m) an IDR of from 0.010 mg / cm2 / min to 0.020 mg / cm2 / min in water; or(n) combinations thereof.

31. A crystalline form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)characterized by unit cell parameters substantially equal to the following cell dimensions: a = 14.410 A; b = 14.410 A; c = 8.247 A; a = 90 degrees; p = 90 degrees; y = 120 degrees; and Space group = P3i.

32. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 80% (e.g., about 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

33. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 84% (e.g., about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

34. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 87% (e.g., about 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

35. A solid form of a compound, (3Z,5S)-5-(hydroxymethyl)-1-[(2'-methyl-1 , 1 '-biphenyl-4- yl)carbonyl]pyrrolidin-3-one O-methyloxime, represented by formula (I)comprising crystalline Form B of compound (I), wherein the crystalline Form B of compound (I) is present at greater than about 89% (e.g., about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) by weight of the solid form.

36. A pharmaceutical composition comprising the crystalline form of any one of claims 1-31 or the solid form of any one of claims 32-35, and one or more carriers, diluents, or excipients.

37. A method of treating a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form of any one of claims 1-31 , the solid form of any one of claims 32-35, or the pharmaceutical composition of claim 36.

38. A method of reducing the likelihood of embryo implantation failure in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form of any one of claims 1-31 , the solid form of any one of claims 32-35, or the pharmaceutical composition of claim 36.

39. A method of improving endometrial receptivity in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form of any one of claims 1-31 , the solid form of any one of claims 32-35, or the pharmaceutical composition of claim 36.

40. A method of reducing uterine contractility in a subject undergoing an embryo transfer procedure in which one or more embryos are transferred to the uterus of the subject, the method comprising administering to the subject the crystalline form of any one of claims 1-31 , the solid form of any one of claims 32-35, or the pharmaceutical composition of claim 36.41 . The method of any one of claims 37-40, wherein the method comprises transferring the one or more embryos to the uterus of the subject.

42. The method of any one of claims 37-41 , wherein the method further comprises inducing follicular maturation in the subject, optionally wherein the inducing of follicular maturation comprises administering human chorionic gonadotropin (hCG) to the subject.

43. The method of any one of claims 37-42, wherein from 1 to 2 embryos are transferred to the subject.

44. The method of claim 43, wherein 1 embryo is transferred to the subject.

45. The method of claim 43, wherein 2 embryos are transferred to the subject.

46. The method of any one of claims 37-45, wherein the subject is a mammal and the one or more embryos are mammalian embryos.

47. The method of claim 46, wherein the mammal is a human and the one or more mammalian embryos are human embryos.

48. The method of any one of claims 37-47, wherein the one or more embryos are produced ex vivo by in vitro fertilization (IVF).

49. The method of claim 48, wherein the one or more embryos are produced ex vivo by IVF of one or more ova derived from the subject.

50. The method of any one of claims 37-47, wherein the one or more embryos are produced ex vivo by intracytoplasmic sperm injection (ICSI).51 . The method of claim 50, wherein the one or more embryos are produced ex vivo by ICSI into one or more ova derived from the subject.

52. The method of claim 49 or 51 , wherein the one or more ova are derived from one or more oocytes isolated from the subject.

53. The method of claim 52, wherein the one or more oocytes comprise from 1 to 4 mature oocytes.

54. The method of claim 52 or 53, wherein a gonadotropin-releasing hormone (GnRH) antagonist is administered to the subject prior to isolation of the one or more oocytes from the subject.

55. The method of any one of claims 52-54, wherein hCG is administered to the subject prior to isolation of the one or more oocytes from the subject.

56. The method of any one of claims 52-55, wherein progesterone is administered to the subject following isolation of the one or more oocytes from the subject.

57. The method of claim 49 or 51 , wherein the one or more ova are isolated directly from the subject.

58. The method of claim 57, wherein a GnRH antagonist is administered to the subject prior to isolation of the one or more ova from the subject.

59. The method of claim 57 or 58, wherein hCG is administered to the subject prior to isolation of the one or more ova from the subject.

60. The method of any one of claims 57-59, wherein progesterone is administered to the subject following isolation of the one or more ova from the subject.61 . The method of any one of claims 37-60, wherein the one or more embryos each comprise from 6 to 8 blastomeres immediately prior to the transfer of the one or more embryos to the subject.

62. The method of claim 61 , wherein the blastomeres are of approximately equal sizes as assessed by visual microscopy.

63. A kit comprising the crystalline form of any one of claims 1 -31 , the solid form of any one of laims 32-35, or the pharmaceutical composition of claim 36, wherein the kit further comprises a package insert instructing a user of the kit to administer the crystalline form to a subject in accordance with the method of any one of claims 37-62.

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