WO2023111817A1

WO2023111817A1 - Crystalline forms of [(1r,5s,6r)-3-{2-[(2s)-2-methylazetidin-1-yl]-6-(trifluoromethyl) pyrimidin-4-yl}-3-azabicyclo[3.1.0]hex-6-yl]acetic acid

Info

Publication number: WO2023111817A1
Application number: PCT/IB2022/062077
Authority: WO
Inventors: Kevin Richard Back; Emily Kathleen CORLETT; Thomas William DANE; Cheryl Louise DOHERTY; Adrian Paul HUTCHINSON; Paul Thomas WHITESIDE
Original assignee: Pfizer Inc.
Priority date: 2021-12-17
Filing date: 2022-12-12
Publication date: 2023-06-22

Abstract

The invention provides solid forms of 2-amino-2-(hydroxymethyl)propane-1,3-diol salt of [(1R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1.0]hex-6-yl]acetic acid, for example, anhydrous tris salt Form 1 and monohydrate tris salt Form 2; as well as pharmaceutical compositions, and the uses thereof in treating diseases, conditions, or disorders, including hereditary fructose intolerance (HFI).

Description

Crystalline Forms of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl) pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid

FIELD OF INVENTION

The invention provides crystalline forms of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid, including the tris salt and other polymorphs, pharmaceutical compositions, dosage forms, and the use thereof in treating diseases, conditions or disorders modulated by the inhibition of ketohexokinase (KHK) enzyme(s) in an subject.

BACKGROUND OF THE INVENTION

[(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (referred to herein as “free acid”) having the following structural formula:

is an inhibitor of ketohexokinase enzyme and was prepared as the crystalline free acid in Example 4 of U.S. Patent No. 9,809,579, which claims the benefit of U.S. Provisional Patent Application No. 62/272,598, filed on December 29, 2015, and Provisional Patent Application No. 62/423,549, filed on November 17, 2016, all of which are hereby incorporated herein by reference in their entireties for all purposes. The crystalline free acid has been shown to be an experimentally acceptable form, however, it possesses a high plasticity. Further experimental salt work has identified additional physically and chemically stable salt solid forms.

Nonalcoholic fatty liver disease (NAFLD) is the hepatic manifestation of metabolic syndrome, and is a spectrum of hepatic conditions encompassing steatosis, non-alcoholic steatohepatitis (NASH), fibrosis, cirrhosis and ultimately hepatocellular carcinoma. NAFLD and NASH are considered the primary fatty liver diseases as they account for the greatest proportion of individuals with elevated hepatic lipids. The severity of NAFLD/NASH is based on the presence of lipid, inflammatory cell infiltrate, hepatocyte ballooning, and the degree of fibrosis. Although not all individuals with steatosis to NASH, a substantial portion do.

Recent human data suggests that fructose consumption may contribute to the development of NAFLD/NASH (Vos, M. B„ and Lavine, J. E. (2013, Hepatology 57 , 2525-2531). Compared to glucose, fructose significantly elevates de novo lipid synthesis (Stanhope, K. L., Schwarz, et al., (2009), J Clin Invest 119, 1322-1334), a distinct characteristic of patients with NAFLD (Lambert, J. E., et al., (2014), Gastroenterology 146, 726-735). Studies in humans have demonstrated that short term fructose feeding causes increases in hepatic triglycerides and that removal of fructose consumption can reverse hepatic triglyceride accumulation (Schwarz, J. M., Noworolski, et al., (2015), J Clin Endocrinol Metab 100, 2434-2442). Moreover, in adolescents with NAFLD, 50% reduction of sugar intake for 10 days reduced hepatic triglyceride by 20% (Schwarz, J. M., Noworolski, et al., (2015) PP07-3: Isocaloric Fructose Restriction for 10 Days Reduces Hepatic De Novo Lipogenesis and Liver Fat in Obese Latino and African American Children, http://press.endocrine.org. proxyl .athensams.net/doi/abs/10.1210/endo- meetings.2015.OABA.6.PP07-3).

The high prevalence of T2D, obesity and NAFLD/NASH and associated co-morbidities, such as cardiovascular disease and stroke, has led to increased desire for both preventive care and therapeutic interventions. Inhibiting KHK metabolism of fructose offers a novel alternative to current treatment strategies.

Ketohexokinase (KHK) is the principle enzyme in fructose metabolism and catalyzes the conversion of fructose to fructose-1 -phosphate (F1 P). KHK is expressed as two alternative mRNA splice variants, denoted KHKa and KHKc, resulting from alternative splicing of the third exon. The affinity and capacity of KHKc for fructose phosphorylation is much greater than KHKa as evidenced by a much lower Km (Ishimoto, Lanaspa et al., PNAS 109, 4320-4325, 2012). While KHKa is ubiquitously expressed, the expression of KHKc is highest in the liver, kidney and intestines, the primary sites of fructose metabolism in the body (Diggle CP, et al. (2009) J Histochem Cytochem 57.7Q3-774-, Ishimoto, Lanaspa, et al., PNAS 109, 4320-4325, 2012). Additionally, loss of function mutations has been reported in humans with no adverse effects except the appearance of fructose in the urine after ingestion of the sugar.

A more severe condition involved in fructose metabolism is Hereditary Fructose Intolerance (HFI, OMIM #229600) which is caused by defects in aldolase B (GENE: ALDOB) which is the enzyme responsible for breaking down F1 P and is immediately downstream of the KHK step in the pathway (Bouteldja N, et. al, J. Inherit. Metab. Dis. 2010 Apr;33(2):105-12; Tolan, DR, Hum Mutat. 1995;6(3):210-8; http://www.omim.org/entry/229600). It is a rare disorder which affects an estimated 1 in 20,000 people, and mutations result in accumulation of F1 P, depletion of ATP, and increase in uric acid, the combination of which causes hypoglycemia, hyperuricemia, and lactic acidosis, among other metabolic derangements. HFI impairs the body’s ability to metabolize dietary fructose resulting in acute symptoms such as vomiting, severe hypoglycemia, diarrhea, and abdominal distress, leading to long term growth defects, liver and kidney damage and potentially death (Ali M et al, J. Med. Genet. 1998 May:35(5):353-65). Patients generally suffer through the first years of life prior to diagnosis, and the only course of treatment is avoiding fructose in the diet. This is made challenging by the presence of this macronutrient in a majority of food items. In addition to physical symptoms, many patients experience emotional and social isolation as a consequence of their unusual diet, and constantly struggle to adhere to strict dietary limitations (HFI-INFO Discussion Board, http://hfiinfo.proboards.com. Accessed 14 December 2015). Even when they appear non-symptomatic, some patients develop NAFLD and kidney disease, which underscores the inadequacy of self-imposed dietary restriction as the only treatment option, and the high unmet medical need for this condition.

In hyperglycemic conditions, endogenous fructose production occurs through the polyol pathway, a pathway by which glucose is converted to fructose with sorbitol as an intermediate. The activity of this pathway increases with hyperglycemia. In these studies, the authors demonstrated that the KHK null mice were protected from glucose induced weight gain, insulin resistance and hepatic steatosis suggesting that under hyperglycemic conditions, endogenously produced fructose may contribute to insulin resistance and hepatic steatosis (Lanaspa, M.A., et al., Nature Comm. 4, 2434, 2013). Therefore, the inhibition of KHK is anticipated to benefit many diseases where alterations of either or both of endogenous and ingested fructose are involved.

There remains a need for oral medicaments containing KHK inhibitors to treat diseases including fructose intolerance (e.g., hereditary fructose intolerance, intestinal fructose intolerance (also referred to as fructose non-absorption or fructose-malabsorption), and fructose-1 ,6- diphosphatase deficiency)), NAFLD, NASH, and T2D.

It is well known that a solid form, for example a crystalline form of a particular drug (including, e.g., anhydrate, hydrate, solvate, etc.) is often an important determinant of the drug’s ease of preparation, stability, solubility, storage stability, ease of formulation, ease of handling, and in vivo pharmacology and/or efficacy. Different crystalline forms occur where the same composition of matter crystallizes in a different lattice arrangement resulting in different thermodynamic properties and stabilities specific to the polymorph form. In cases where two or more solid forms (e.g., two or more crystalline forms, or an amorphous form and one or more crystalline forms) can be produced, it is desirable to have a method to make each of the solid forms in pure form. In deciding which solid form is preferable, the numerous properties of the solid forms must be compared and the preferred solid (e.g., crystalline) form chosen based on the many physical property variables. It is entirely possible that one crystalline form can be preferable in some circumstances where certain aspects such as ease of preparation, stability, etc. are deemed to be critical. In other situations, a different crystalline form maybe preferred for greater solubility and/or superior pharmacokinetics. Moreover, because of the potential advantages associated with one pure crystalline form, it is desirable to prevent or minimize polymorphic conversion (i.e., conversion of one crystal form to another; or conversion between one crystal form and amorphous form) when two or more solid forms of one substance can exist. Such polymorphic conversion can occur during both the preparation of formulations containing a solid form (e.g., a crystalline form), and during storage of a pharmaceutical dosage form containing a solid form (e.g., a crystalline form). Because improved drug formulations showing, for example, better bioavailability or better stability are consistently sought, there is an ongoing need for new or purer solid (e.g., crystalline) forms of existing drug molecules.

Salt formation provides a means of altering the physicochemical and resultant biological characteristics of a drug without modifying its chemical structure. A salt form can have a dramatic influence on the properties of the drug. The selection of a suitable salt form involves evaluation of many factors, including whether any salt can be formed. Other factors included in this selection include hygroscopicity, stability, solubility, and the process profile of any salt form that might be discovered.

The novel solid forms (e.g., tris salt) of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid described herein are directed toward this and other important ends.

SUMMARY OF THE INVENTION

The present invention is directed to crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3- diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (triflu oromethy I) pyrimid in-4-y l}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid.

The present invention is also directed to anhydrous crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid, having:

(a) an X-ray powder diffraction pattern, measured using copper wavelength radiation, comprising peaks expressed in degrees 20 at 5.4 ± 0.1 , and 21 .8 ± 0.1 ;

(b) a Raman spectrum comprising one or more wavenumber (cm ¹) values selected from the group consisting of 1559 ± 2 cm^-1 and 1597 ± 2 cm'¹;

(c) a ¹³C solid state NMR spectrum comprising one or more peaks in terms of chemical shifts selected from the group consisting of 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm;

(d) a ¹⁵N ssNMR spectrum comprising one or more peaks in terms of chemical shifts selected -285.9 ± 0.2 ppm, and -278.0 ± 0.2 ppm; or a combination of two or more of (a), (b), (c), and (d).

The present invention is also directed to pharmaceutical composition comprising a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3- diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (triflu oromethy I) pyrimid in-4-y l}-3- azabicyclo [3.1 ,0]hex-6-yl]acetic acid and a pharmaceutically acceptable carrier, diluent, and/or excipient.

The present invention is also directed to a method of treating or reducing the effects of fructose intolerance in a subject in need thereof, wherein the method provides for the administration to the subject (e.g., human) a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid.

The present invention is also directed to a method of treating fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, nonalcoholic steatohepatitis with cirrhosis and hepatocellular carcinoma, alcoholic fatty liver disease, alcoholic steatohepatitis, hepatitis B, hepatitis C, and biliary cirrhosis in a subject in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid.

The present invention is also directed to 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid monohydrate (tris salt Form 2) having:

(a) an X-ray powder diffraction pattern, measured using copper wavelength radiation, comprising peaks expressed in degrees 20 at 10.2 ± 0.1 , and 11 .2 ± 0.1 ;

(b) a Raman spectrum comprising one or more peaks having wavenumber (cm ¹) values selected from the group consisting of 889 ± 2 cm ¹, 906 ± 2 cm ¹, and 1552 ± 2 cm ¹;

(c) a ¹³C solid state NMR spectrum comprising one or more peaks, in terms of chemical shifts, selected from the group consisting of 42.4 ± 0.2 ppm, 62.4 ± 0.2 ppm, and164.6 ± 0.2 ppm;

(d) a ¹⁵N ssNMR spectrum comprising one or more peaks, in terms of chemical shifts selected -174.3.0 ± 0.2 ppm, and -172.4 ± 0.2 ppm;

(e) a ¹⁹F ssNMR spectrum comprising a peak in terms of chemical shifts at -70.4 ± 0.2 ppm; or a combination of two or more of (a), (b), (c), (d) and (e). The present invention is also directed to pharmaceutical composition comprising a therapeutically effective amount of 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid monohydrate (tris salt Form 2) and a pharmaceutically acceptable carrier, diluent, and/or excipient.

The present invention is also directed to a method of treating or reducing the effects of fructose intolerance in a subject in need thereof, wherein the method provides for the administration to the subject (e.g., human) a therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid monohydrate (tris salt Form 2).

The present invention is also directed to a method of treating fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, nonalcoholic steatohepatitis with cirrhosis and hepatocellular carcinoma, alcoholic fatty liver disease, alcoholic steatohepatitis, hepatitis B, hepatitis C, and biliary cirrhosis in a subject in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid monohydrate (tris salt Form 2).

The crystalline forms of the invention may be characterized according to the powder X- ray diffraction (PXRD) data, solid state Nuclear Magnetic Resonance (ssNMR) data (e.g., ¹³C ssNMR data), and/or FT-Raman spectroscopy data provided herein.

BRIEF DESCRIPTION OF FIGURES

FIG. 1 shows an illustrative PXRD pattern of anhydrous crystalline tris salt Form 1 carried out on a Bruker AXS D8 Endeavor diffractometer equipped with a Cu radiation source (Vertical Axis: Intensity (CPS); Horizontal Axis: Two theta (degrees)).

FIG. 2 shows an illustrative Raman spectra of anhydrous crystalline tris salt Form 1 collected using a RAM II FT-Raman module attached to a Vertex 70 spectrometer (Bruker Optik GmbH). The instrument is equipped with a 1064 nm solid-state (Nd:YAG) laser and a liquid nitrogen cooled germanium detector.

FIG. 3 shows an illustrative ¹³C ssNMR pattern of anhydrous crystalline tris salt Form 1 conducted on a Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹³C spectra were collected on a 4 mm MAS probe at a magic angle spinning rate of 10 kHz. FIG. 4 shows illustrative ¹⁵N ssNMR pattern of anhydrous crystalline tris salt Form 1 conducted on a Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹⁵N spectra were collected on a 4 mm MAS probe at a magic angle spinning rate of 8 kHz.

FIG. 5 shows illustrative ¹⁹F ssNMR pattern of anhydrous crystalline tris salt Form 1 Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹⁹F spectra were recorded with a 3.2 mm MAS pro beat a spin rate of 20 kHz.

FIG. 6 shows an illustrative PXRD pattern of crystalline monohydrate tris salt Form 2 carried out on a Bruker AXS D8 Endeavor diffractometer equipped with a Cu radiation source (Vertical Axis: Intensity (CPS); Horizontal Axis: Two theta (degrees)).

FIG. 7 shows an illustrative Raman spectra of crystalline monohydrate tris salt Form 2 collected using a RAM II FT-Raman module attached to a Vertex 70 spectrometer (Bruker Optik GmbH). The instrument is equipped with a 1064 nm solid-state (Nd:YAG) laser and a liquid nitrogen cooled germanium detector.

FIG. 8 shows an illustrative ¹³C ssNMR pattern of crystalline monohydrate tris salt Form 2 conducted on a Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹³C spectra were collected on a 4 mm MAS probe at a magic angle spinning rate of 10 kHz.

FIG. 9 shows illustrative ¹⁵N ssNMR pattern of crystalline monohydrate tris salt Form 2 conducted on a Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹⁵N spectra were collected on a 4 mm MAS probe at a magic angle spinning rate of 8 kHz.

FIG. 10 shows illustrative ¹⁹F ssNMR pattern of crystalline monohydrate tris salt Form 2 Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹⁹F spectra were recorded with a 3.2 mm MAS pro beat a spin rate of 20 kHz.

DETAILED DESCRIPTION OF THE INVENTION

The present invention may be understood more readily by reference to the following detailed description of exemplary embodiments of the invention and the examples included therein.

It is to be understood that this invention is not limited to specific preparation methods that may of course vary. It is also to be understood that the terminology used herein is for the purpose of describing embodiments only and is not intended to be limiting.

The term “tris” means 2-amino-2-(hydroxymethyl)propane-1 ,3-diol, also known as THAM and tromethamine.

The term “tris salt Form 1 ” means the anhydrous crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol (tris) salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid as the mono-tris salt. It is intended that tris salt Form 1 is free of water, but residual solvent, including water, could be present if the material is not dried completely. The tris salt Form 1 is made using 2-amino-2- (hydroxymethyl)propane-l ,3-diol. Tris is associated with the carboxylic acid moiety of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid. Unless otherwise stated, when referencing the tris salt of the compound, the counterion and the [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid are in a stoichiometric ratio of about 1 :1 .

The term “tris salt Form 2” means crystalline monohydrate 2-amino-2- (hydroxymethyl)propane-l ,3-diol (tris) salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid as the mono-tris salt.

The term “monohydrate” as used herein refers to the inclusion of about one water molecule.

The term “peak(s)” means a signal that is discernable from noise”.

The term “chemical shifts” as used herein with regard to ¹³C solid state NMR spectrum, ¹⁵N ssNMR spectrum, and/or ¹⁹F solid state NMR spectrum means a signal in the solid state NMR spectrum that is discernable from the noise.

Any solid form of the present invention can be substantially pure. As used herein, the term "substantially pure" with reference to a particular solid form (e.g. a crystalline form) means that the particular solid form (e.g. the crystalline form) includes less than 15%, less than 10%, less than 5%, less than 3%, or less than 1 % by weight of any other physical form of Compound 1 or impurity.

The term “substantially the same” when used to describe X-ray powder diffraction patterns is meant to include patterns in which peaks are within a standard deviation of +/- 0.2° 2Q.

As used herein, the term "substantially pure" with reference to a particular crystalline form means that the crystalline form includes less than 10%, preferably less than 5%, preferably less than 3%, preferably less than 1 % by weight of any other physical form of the crystalline free acid.

The term “substantially the same” when used to describe powder X-ray diffraction (PXRD) patterns is meant to include patterns in which peaks (in terms of 20) are within the deviations specified herein.

The term “substantially the same” when used to describe an ssNMR spectrum meant to include ssNMR spectra in which peaks (in terms of chemical shifts) are within the deviations specified herein. The term “substantially the same” when used to describe an FT-Raman spectrum meant to include FT-Raman spectra in which peaks (in terms of wavenumber) are within the deviations specified herein.

The term "a" or "an" may mean one or more. As used herein in the claim(s), when used in conjunction with the word "comprising", the words "a" or "an" may mean one or more than one. As used herein "another" may mean at least a second or more.

The term “about” generally means within 10%, preferably within 5%, and more preferably within 1% of a given value or range. Alternatively, the term “about” means within an acceptable standard error of the mean, when considered by one skilled in the art.

The term “subject” includes humans, mammals. Human subjects may be of either gender or at any stage of development.

“Therapeutically effective amount” means an amount of a compound described herein that treats the disease, condition, or disorder described herein. “Therapeutically effective amount” can also mean an amount of a compound described herein, optionally, in combination with an amount of another compound(s), that treats the disease, condition, or disorder described herein.

The term "treating", "treat" or "treatment" as used herein embraces preventative, i.e., prophylactic; palliative treatment, i.e., relieve, alleviate, or slow the progression of the patient’s disease (or condition) or any tissue damage associated with the disease (or condition); and reversal where the patient’s disease (or condition) is not only alleviated but any tissue damage associated with the disease (or condition) is placed in a better state then when treatment was initiated.

Those skilled in the art would readily understand that multiple nomenclatures can be used to name a same compound (including a same salt).

Every example or embodiment of solid forms of the invention may be claimed individually or grouped together in any combination with any number of each and every embodiment described herein.

Solid Forms

The invention concerns crystalline forms of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid. Specifically, the invention concerns the mono-tris salt of ([(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid). There are two crystalline forms of the mono-tris salt: Form 1 is an anhydrous crystalline solid (tris salt Form 1) and Form 2 is a crystalline monohydrate solid (tris salt Form 2). The crystalline forms of the present invention have properties which are particularly suitable for use as a drug, including improved solubility, and bioavailability.

One embodiment concerns a crystalline salt form that is anhydrous crystalline 2-amino- 2-(hydroxymethyl)propane-1 , 3-d io I salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

Another embodiment concerns anhydrous crystalline 2-amino-2-(hydroxymethyl) propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) having a purity of greater than 90%.

Another embodiment concerns anhydrous crystalline 2-amino-2-(hydroxymethyl) propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) having a purity of greater than 95%.

Another embodiment concerns anhydrous crystalline 2-amino-2-(hydroxymethyl) propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) having a purity of greater than 97%.

Another embodiment concerns anhydrous crystalline 2-amino-2-(hydroxymethyl) propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) having a purity of greater than 99%.

In any of the above-mentioned embodiments the anhydrous crystaline form is designated as tris salt Form 1 that can be identified by its unique solid-state signatures with respect to, for example, powder X-ray diffraction (PXRD) data, solid state Nuclear Magnetic Resonance (ssNMR) data (e.g., ¹³C ssNMR data, ¹⁵N ssNMR data, and/or ¹⁹F ssNMR), and/or FT-Raman Spectroscopy data provided herein. Tris salt Form 1 contains a 1 :1 ratio of [(1 R,5S,6R)-3-{2-[(2S)- 2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid to 2-amino-2-(hydroxymethyl) propane-1 ,3-diol (tromethamine).

In any of the above-mentioned embodiments the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least one characteristic peak expressed in degrees 20 selected from 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21.8 ± 0.1. In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 5.4+ 0.1.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 11.8 + 0.1.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 16.3 + 0.1.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 21 .8 + 0.1.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least two characteristic peaks expressed in degrees 20 selected from 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21.8 ± 0.1.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least three characteristic peaks expressed in degrees 20 selected from 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21.8 ± 0.1.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having characteristic peaks expressed in degrees 20 at 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21 .8 ± 0.1 .

In another embodiment, the anhydrous crystalline tris salt Form 1 has a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, substantially the same as Figure 1 .

A further embodiment concerns the anhydrous crystalline tris salt Form 1 of any of the above-mentioned embodiments, wherein the crystalline form exhibits a ¹³C solid state NMR spectrum comprising at least one peak, in terms of chemical shifts selected from 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 55.8 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 58.3 ± 0.2 ppm. In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 63.7 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 64.2 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 183.6 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a ¹³C solid state NMR spectrum comprising at least two peaks, in terms of chemical shifts selected 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a ¹³C solid state NMR spectrum comprising at least three peaks, in terms of chemical shifts 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a ¹³C solid state NMR spectrum comprising at least four peaks, in terms of chemical shifts 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a ¹³C solid state NMR spectrum comprising peaks, in terms of chemical shifts at 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹³C ssNMR spectrum substantially the same as Figure 3.

A further embodiment concerns the anhydrous crystalline tris salt Form 1 of any of the above-mentioned embodiments, wherein the crystalline form has a ¹⁵N ssNMR spectrum comprising at least one peak, in terms of chemical shifts, selected from -342.1 ± 0.2 ppm, -285.9

± 0.2 ppm, -278.0 ± 0.2 ppm, and -177.5 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁵N ssNMR spectrum comprising one peak, in terms of chemical shifts, at -342.1 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁵N ssNMR spectrum comprising one peak, in terms of chemical shifts, at -285.9 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁵N ssNMR spectrum comprising one peak, in terms of chemical shifts, at -177.5 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁵N ssNMR spectrum comprising at least two peaks, in terms of chemical shifts selected from -342.1 ± 0.2 ppm, -285.9 ± 0.2 ppm, -278.0 ± 0.2 ppm, and -177.5 ± 0.2 ppm In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁵N ssNMR spectrum comprising peaks, in terms of chemical shifts, at -342.1 ± 0.2 ppm, -285.9 ± 0.2 ppm, -278.0 ± 0.2 ppm, and -177.5 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁵N ssNMR spectrum substantially the same as Figure 4.

A further embodiment concerns the anhydrous crystalline tris salt Form 1 of any of the above-mentioned embodiments, wherein the crystalline form has a ¹⁹F ssNMR spectrum comprising at least one peak, in terms of chemical shifts, at -68.5 ± 0.2 ppm.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a ¹⁹F ssNMR spectrum substantially the same as Figure 5.

A further embodiment concerns the anhydrous crystalline tris salt Form 1 of any of the above-mentioned embodiments, wherein the crystalline form has a Raman spectrum comprising at least one peak having a wavenumber (cm'¹) value selected from the group consisting of 304 ± 2 cm'¹ , 315 ± 2 cm'¹ , 935 ± 2 cm'¹ , 1559 ± 2 cm'¹ and 1597 ± 2 cm'¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 304 ± 2 cm ¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a Raman spectrum comprising one peak having a wavenumber (cm^-1) value at 315 ± 2 cm ¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 935 ± 2 cm ¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 1559 ± 2 cm'¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 1597 ± 2 cm'¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a Raman spectrum having at least two characteristic peaks having wavenumber (cm'¹) values selected from the group consisting of 304 ± 2 cm ¹, 315 ± 2 cm'¹, 935 ± 2 cm'¹, 1559 ± 2 cm'¹ and 1597 ± 2 cm ¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a Raman spectrum having at least three characteristic peaks comprising wavenumber (cm ¹) values selected from the group consisting 304 ± 2 cm ¹, 315 ± 2 cm ¹, 935 ± 2 cm ¹, 1559 ± 2 cm'¹ and 1597 ± 2 cm'¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a Raman spectrum having at least four characteristic peaks comprising wavenumber (cm'¹) values selected from the group consisting 304 ± 2 cm ¹, 315 ± 2 cm ¹, 935 ± 2 cm ¹, 1559 ± 2 cm ¹ and 1597 ± 2 cm ¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 exhibits a Raman spectrum comprising wavenumber (cm'¹) values at 304 ± 2 cm^-1 , 315 ± 2 cm^-1 , 935 ± 2 cm^-1 , 1559 ± 2 cm'¹ and 1597 ± 2 cm'¹.

In another embodiment, the anhydrous crystalline tris salt Form 1 has a Raman spectrum substantially the same as Figure 2.

A further embodiment concerns anhydrous crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (Form 1) having:

(b) a Raman spectrum comprising one or more wavenumber (cm'¹) values selected from the group consisting of 1559 ± 2 cm^-1 and 1597 ± 2 cm'¹;

In a further embodiment, the anhydrous crystalline tris salt Form 1 of any of the above- mentioned embodiments has a purity greater than 97%.

In a further embodiment, the anhydrous crystalline tris salt Form 1 of any of the above- mentioned embodiments has a purity greater than 99%.

A further embodiment concerns crystalline monohydrate 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt monohydrate Form 2).

In another embodiment, the crystalline monohydrate 2-amino-2-(hydroxymethyl)propane- 1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt monohydrate Form 2) has a purity of greater than 90%.

In another embodiment, the crystalline monohydrate 2-amino-2-(hydroxymethyl)propane- 1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt monohydrate Form 2)has a purity of greater than 95%. In another embodiment, the crystalline monohydrate 2-amino-2-(hydroxymethyl)propane- 1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt monohydrate Form 2)has a purity of greater than 97%.

In another embodiment, the crystalline monohydrate 2-amino-2-(hydroxymethyl)propane- 1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt monohydrate Form 2) has a purity of greater than 99%.

In any of the above-mentioned embodiments, the tris salt monohydrate Form 2 can be identified by its unique solid-state signatures with respect to, for example, powder X-ray diffraction (PXRD) data, solid state Nuclear Magnetic Resonance (ssNMR) data (e.g., ¹³C ssNMR data, ¹⁵N ssNMR data, and/or ¹⁹F ssNMR), and/or FT-Raman Spectroscopy data provided herein. Tris salt monohydrate Form 2 contains a 1 :1 ratio of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid to 2-amino-2-(hydroxymethyl)propane-1 ,3-diol (tromethamine).

In any of the above-mentioned embodiments the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least one characteristic peak expressed in degrees 20 selected from 5.6 ± 0.1 , 10.2 ± 0.1 , 11 .2 ± 0.1 , and 15.3 ± 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 5.6+ 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 10.2 + 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 11.2 + 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, of one peak, in terms of 20, at 15.3 + 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least two characteristic peaks expressed in degrees 20 selected from 5.6 ± 0.1 , 10.2 ± 0.1 , 11 .2 ± 0.1 , and 15.3 ± 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least three characteristic peaks expressed in degrees 20 selected from 5.6 ± 0.1 , 10.2 ± 0.1 , 11 .2 ± 0.1 , and 15.3 ± 0.1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having characteristic peaks expressed in degrees 20 at 5.6 ± 0.1 , 10.2 ± 0.1 , 11.2 ± 0.1 , and 15.3 ± 0.1.

In another embodiment, the tris salt monohydrate Form 2 has a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, substantially the same as Figure 6.

A further embodiment concerns the tris salt monohydrate Form 2 of any of the above- mentioned embodiments, wherein the crystalline form exhibits a ¹³C solid state NMR spectrum comprising at least one peak, in terms of chemical shifts selected from 42.4 ± 0.2 ppm, 46.7 ± 0.2 ppm, 62.4 ± 0.2 ppm, 91 .2 ± 0.2 ppm, and 164.6 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 42.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 46.7 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 62.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 91.2 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹³C ssNMR spectrum comprising one peak, in terms of chemical shifts, at 164.6 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 exhibits a ¹³C solid state NMR spectrum comprising at least two peaks, in terms of chemical shifts selected 42.4 ± 0.2 ppm, 46.7 ± 0.2 ppm, 62.4 ± 0.2 ppm, 91.2 ± 0.2 ppm, and 164.6 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 exhibits a ¹³C solid state NMR spectrum comprising at least three peaks, in terms of chemical shifts 42.4 ± 0.2 ppm, 46.7 ± 0.2 ppm, 62.4 ± 0.2 ppm, 91 .2 ± 0.2 ppm, and 164.6 ± 0.2 ppm. In another embodiment, the tris salt monohydrate Form 2 exhibits a ¹³C solid state NMR spectrum comprising at least four peaks, in terms of chemical shifts 42.4 ± 0.2 ppm, 46.7 ± 0.2 ppm, 62.4 ± 0.2 ppm, 91 .2 ± 0.2 ppm, and 164.6 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 exhibits a ¹³C solid state NMR spectrum comprising peaks, in terms of chemical shifts at 42.4 ± 0.2 ppm, 46.7 ± 0.2 ppm, 62.4 ± 0.2 ppm, 91 .2 ± 0.2 ppm, and 164.6 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹³C ssNMR spectrum substantially the same as Figure 8.

A further embodiment concerns the tris salt monohydrate Form 2 of any of the above- mentioned embodiments, wherein the crystalline form has a ¹⁵N ssNMR spectrum comprising at least one peak, in terms of chemical shifts, selected from -342.8 ± 0.2 ppm, -174.3 ± 0.2 ppm, and -172.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁵N ssNMR spectrum comprising one peak, in terms of chemical shifts, at -342.8 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁵N ssNMR spectrum comprising one peak, in terms of chemical shifts, at -174.3 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁵N ssNMR spectrum comprising one peak, in terms of chemical shifts, at -172.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁵N ssNMR spectrum comprising at least two peaks, in terms of chemical shifts selected from -342.8 ± 0.2 ppm,

-174.3 ± 0.2 ppm, and -172.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁵N ssNMR spectrum comprising peaks, in terms of chemical shifts, at -342.8 ± 0.2 ppm, -174.3 ± 0.2 ppm, and -172.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁵N ssNMR spectrum substantially the same as Figure 9.

A further embodiment concerns the tris salt monohydrate Form 2 of any of the above- mentioned embodiments, wherein the crystalline form has a ¹⁹F ssNMR spectrum comprising at least one peak, in terms of chemical shifts, at -70.4 ± 0.2 ppm.

In another embodiment, the tris salt monohydrate Form 2 has a ¹⁹F ssNMR spectrum substantially the same as Figure 10.

A further embodiment concerns the tris salt monohydrate Form 2 of any of the above- mentioned embodiments, wherein the crystalline form has a Raman spectrum comprising at least one peak having a wavenumber (cm'¹) value selected from the group consisting of 889 ± 2 cm ¹, 906 ± 2 cm ¹ , 1 162 ± 2 cm ¹ , 1552 ± 2 cm ¹ and 2928 ± 2 cm ¹.

In another embodiment, the tris salt monohydrate Form 2 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 889 ± 2 cm ¹.

In another embodiment, the tris salt monohydrate Form 2 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 906 ± 2 cm ¹.

In another embodiment, the tris salt monohydrate Form 2 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 1162 ± 2 cm'¹.

In another embodiment, the tris salt monohydrate Form 2 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 1552 ± 2 cm'¹.

In another embodiment, the tris salt monohydrate Form 2 has a Raman spectrum comprising one peak having a wavenumber (cm'¹) value at 2928 ± 2 cm'¹.

In another embodiment, the tris salt monohydrate Form 2 exhibits a Raman spectrum having at least two characteristic peaks having wavenumber (cm'¹) values selected from the group consisting of 889 ± 2 cm^-1 , 906 ± 2 cm^-1 , 1 162 ± 2 cm^-1 , 1552 ± 2 cm^-1 and 2928 ± 2 cm^-1.

In another embodiment, the tris salt monohydrate Form 2 exhibits a Raman spectrum having at least three characteristic peaks comprising wavenumber (cm^-1) values selected from the group consisting 889 ± 2 cm'¹, 906 ± 2 cm'¹, 1 162 ± 2 cm'¹, 1552 ± 2 cm'¹ and 2928 ± 2 cm'¹.

In another embodiment, the tris salt monohydrate Form 2 exhibits a Raman spectrum having at least four characteristic peaks comprising wavenumber (cm ¹) values selected from the group consisting 889 ± 2 cm'¹, 906 ± 2 cm'¹, 1 162 ± 2 cm'¹, 1552 ± 2 cm'¹ and 2928 ± 2 cm'¹.

In another embodiment, the tris salt monohydrate Form 2 has a Raman spectrum substantially the same as Figure 7.

A further embodiment concerns 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid monohydrate ( tris salt Form 2) having:

(a) an X-ray powder diffraction pattern, measured using copper wavelength radiation, comprising peaks expressed in degrees 20 at 10.2 ± 0.1 , and 1 1 .2 ± 0.1 ;

(b) a Raman spectrum comprising one or more peaks having wavenumber (cm'¹) values selected from the group consisting of 889 ± 2 cm ¹, 906 ± 2 cm ¹, and 1552 ± 2 cm ¹;

(d) a ¹⁵N ssNMR spectrum comprising one or more peaks, in terms of chemical shifts selected -174.3.0 ± 0.2 ppm, and -172.4 ± 0.2 ppm; (e) a ¹⁹F ssNMR spectrum comprising a peak in terms of chemical shifts at -70.4 ± 0.2 ppm; or a combination of two or more of (a), (b), (c), (d) and (e).

In a further embodiment, the tris salt monohydrate Form 2 of any of the above- mentioned embodiments has a purity greater than 97%.

In a further embodiment, the tris salt monohydrate Form 2 of any of the above-mentioned embodiments has a purity greater than 99%.

Another embodiment concerns amorphous form (Form 3) of the crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid.

Pharmaceutical Compositions and Routes of Administration

Another embodiment concerns a pharmaceutical composition comprising a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1), and a pharmaceutically acceptable carrier, diluent, and/or excipient. The anhydrous tris salt Form 1 utilized in the pharmaceutical composition can be the tris salt Form 1 as described in any of the above-mentioned embodiments. In one embodiment, the pharmaceutical composition comprises at least 10% of anhydrous tris salt Form 1. In another embodiment, the pharmaceutical composition comprises at least 20% of anhydrous tris salt Form 1 . In another embodiment the pharmaceutical composition comprises at least 30% of anhydrous tris salt Form 1 . In another embodiment, the pharmaceutical composition comprises at least 40% of tris salt Form 1. In another embodiment the pharmaceutical composition comprises at least 50% of anhydrous tris salt Form 1 . In another embodiment, the pharmaceutical composition comprises at least 60% of anhydrous tris salt Form 1. In another embodiment the pharmaceutical composition comprises at least 70% of anhydrous tris salt Form 1 . In another embodiment, the pharmaceutical composition comprises at least 80% of anhydrous tris salt Form 1 . In another embodiment the pharmaceutical composition comprises at least 90% of anhydrous tris salt Form 1 . In another embodiment, the pharmaceutical composition comprises at least 95% of anhydrous tris salt Form 1 . In another embodiment the pharmaceutical composition comprises at least 97% of anhydrous tris salt Form 1 . In another embodiment, the pharmaceutical composition comprises at least 99% of anhydrous tris salt Form 1 .

Another embodiment concerns a pharmaceutical composition comprising a therapeutically effective amount of crystalline monohydrate 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo

[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2), and a pharmaceutically acceptable carrier, diluent, and/or excipient.

Another embodiment concerns a pharmaceutical composition comprising a therapeutically effective amount of 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

In any one of the above-mentioned embodiments, the "pharmaceutically acceptable carrier, diluent or excipient" includes all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. Examples of pharmaceutically acceptable carrier, diluent or excipients include one or more of water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, and the like, as well as combinations thereof, and may include isotonic agents, for example, sugars, sodium chloride, or polyalcohols such as mannitol, or sorbitol in the composition. Pharmaceutically acceptable substances such as wetting agents or minor amounts of auxiliary substances such as wetting or emulsifying agents, preservatives, or buffers, which enhance the shelf life or effectiveness of the active pharmaceutical ingredient (e.g., anhydrous tris salt Form 1 , monohydrate tris salt Form 2, amorphous Form 3).

The compositions of this invention may be in a variety of forms. These include, for example, liquid, semi-solid and solid dosage forms, such as liquid solutions (e.g., oral, injectable and infusible solutions), dispersions or suspensions, tablets, pills, powders, liposomes, topical formulations, intranasal formulations, and suppositories. The form depends on the intended mode of administration and therapeutic application.

Typical compositions are in the form of parenteral (e.g., injectable, or infusible solutions). "Parenteral administration" includes, for example, subcutaneous injections, intravenous injections, intraperitoneally, intramuscular injections, intrasternal injections, and infusion. Injectable preparations (i.e., sterile injectable aqueous or oleaginous suspensions) may be formulated according to the known art using suitable dispersing, wetting agents, and/or suspending agents.

Oral administration of a solid dosage form may be, for example, presented in discrete units, such as hard or soft capsules, pills, cachets, lozenges, or tablets, each containing a predetermined amount of at least one compound of the invention. Oral administration may also be in a powder or granule form. Oral dose forms may be sub-lingual, such as, for example, a lozenge. In such solid dosage forms, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be combined with one or more adjuvants. In the case of capsules, tablets, and pills, the dosage forms also may comprise buffering agents or may be prepared with enteric coatings. Capsules or tablets may be in the form of a controlled release formulation.

Oral administration may also be in a liquid dose form. Liquid dosage forms for oral administration include, for example, pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs containing inert diluents commonly used in the art (e.g., water). Such compositions also may comprise adjuvants, such as wetting, emulsifying, suspending, flavoring (e.g., sweetening), and/or perfuming agents.

"Topical administration" includes, for example, transdermal administration, such as via transdermal patches or iontophoresis devices, intraocular administration, or intranasal or inhalation administration. Compositions for topical administration also include, for example, topical gels, sprays, ointments, and creams. Atopical formulation may include a compound which enhances absorption or penetration of the active ingredient through the skin or other affected areas. When the compounds of this invention are administered by a transdermal device, administration will be accomplished using a patch either of the reservoir and porous membrane type or of a solid matrix variety. Typical formulations for this purpose include gels, hydrogels, lotions, solutions, creams, ointments, dusting powders, dressings, foams, films, skin patches, wafers, implants, sponges, fibres, bandages and microemulsions. Liposomes may also be used. Typical carriers include alcohol, water, mineral oil, liquid petrolatum, white petrolatum, glycerin, polyethylene glycol and propylene glycol. Penetration enhancers may be incorporated - see, for example, B. C. Finnin and T. M. Morgan, J. Pharm. Sci., vol. 88, pp. 955-958, 1999.

Formulations suitable for topical administration to the eye include, for example, eye drops wherein the compound of this invention is dissolved or suspended in a suitable carrier. A typical formulation suitable for ocular or aural administration may be in the form of drops of a micronized suspension or solution in isotonic, pH-adjusted, sterile saline. Other formulations suitable for ocular and aural administration include ointments, biodegradable (i.e., absorbable gel sponges, collagen) and non-biodegradable (i.e., silicone) implants, wafers, lenses and particulate or vesicular systems, such as niosomes or liposomes. A polymer such as crossed linked polyacrylic acid, polyvinyl alcohol, hyaluronic acid, a cellulosic polymer, for example, hydroxypropylmethylcellulose, hydroxyethylcellulose, or methylcellulose, or a heteropolysaccharide polymer, for example, gelan gum, may be incorporated together with a preservative, such as benzalkonium chloride. Such formulations may also be delivered by iontophoresis.

“Intranasal administration” or administration by “inhalation”, are conveniently delivered in the form of a solution or suspension from a pump spray container that is squeezed or pumped by the patient or as an aerosol spray presentation from a pressurized container or a nebulizer, with the use of a suitable propellant. Formulations suitable for intranasal administration are typically administered in the form of a dry powder (either alone, as a mixture, for example, in a dry blend with lactose, or as a mixed component particle, for example, mixed with phospholipids, such as phosphatidylcholine) from a dry powder inhaler or as an aerosol spray from a pressurized container, pump, spray, atomizer (preferably an atomizer using electrohydrodynamics to produce a fine mist), or nebulizer, with or without the use of a suitable propellant, such as 1 , 1 ,1 ,2- tetrafluoroethane or 1 ,1 ,1 ,2,3,3, 3-heptafluoropropane. For intranasal use, the powder may comprise a bioadhesive agent, for example, chitosan or cyclodextrin.

“Rectal” dosage forms may be in the form of, for example, a suppository or enema. Cocoa butter is a traditional suppository base, but various alternatives may be used as appropriate.

Multiple carriers, diluent or excipient materials and modes of administration known in the pharmaceutical art may also be used. Pharmaceutical compositions of the invention may be prepared by any of the well-known techniques of pharmacy, such as effective formulation and administration procedures. The above considerations regarding effective formulations and administration procedures are well known in the art and are described in standard textbooks. Formulation of drugs is discussed in, for example, Hoover, John E., Remington’s Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania, 1975; Liberman et al., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y., 1980; and Kibbe et al., Eds., Handbook of Pharmaceutical Excipients (3rd Ed.), American Pharmaceutical Association, Washington, 1999.

Another embodiment concerns anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3, or compositions containing anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 with a pharmaceutically acceptable carrier, diluent and/or excipient can be administered in an amount effective to treat a condition as described herein.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be administered by any suitable route in the form of a pharmaceutical composition adapted to such a route, and in a dose effective for the treatment intended. Anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be administered orally, rectally, vaginally, intranasally, parenterally, or topically.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be administered orally. Oral administration may involve swallowing, so that the compound enters the gastrointestinal tract, or buccal or sublingual administration may be employed by which the compound enters the bloodstream directly from the mouth.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may also be administered directly into the bloodstream, into muscle, or into an internal organ via parenteral administration. Suitable means for parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, intrasternal, intracranial, intramuscular, and subcutaneous. Suitable devices for parenteral administration include needle (including microneedle) injectors, needle-free injectors, and infusion techniques.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may also be administered topically to the skin or mucosa, that is, dermally or transdermally.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 can also be administered intranasally or by inhalation.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be administered rectally.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may also be administered topically directly to the eye or ear.

In another embodiment, anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be administered vaginally.

In any one of the above-mentioned embodiments, the pharmaceutically acceptable carriers, diluents, excipients, or stabilizers are nontoxic to recipients at the dosages and concentrations employed, and may comprise buffers such as phosphate, citrate, and other organic acids; salts such as sodium chloride; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens, such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or Igs; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g., Zn-protein complexes); and/or non-ionic surfactants such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG).

Liposomes containing these agents and/or anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 are prepared by methods known in the art, such as described in U.S. Pat. Nos. 4,485,045 and 4,544,545. Liposomes with enhanced circulation time are disclosed in U.S. Patent No. 5,013,556. Particularly useful liposomes can be generated by the reverse phase evaporation method with a lipid composition comprising phosphatidylcholine, cholesterol and PEG-derivatized phosphatidylethanolamine (PEG-PE). Liposomes are extruded through filters of defined pore size to yield liposomes with the desired diameter.

These agents and/or anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may also be entrapped in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxymethylcellulose or gelatin-microcapsules and poly-(methylmethacrylate) microcapsules, respectively, in colloidal drug delivery systems (for example, liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules) or in macroemulsions. Such techniques are disclosed in Remington, The Science and Practice of Pharmacy, 20th Ed., Mack Publishing (2000).

Sustained-release preparations may be used. Suitable examples of sustained-release preparations include semi-permeable matrices of solid hydrophobic polymers containing anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 which matrices are in the form of shaped articles, e.g., films, or microcapsules. Examples of sustained-release matrices include polyesters, hydrogels (for example, poly(2-hydroxyethyl-methacrylate), or 'poly(vinylalcohol)), polylactides (U.S. Pat. No. 3,773,919), copolymers of L-glutamic acid and 7 ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as those used in LUPRON DEPOT™ (injectable microspheres composed of lactic acid-glycolic acid copolymer and leuprolide acetate), sucrose acetate isobutyrate, and poly- D-(-)-3-hydroxybutyric acid.

The compositions to be used for intravenous administration must be sterile. This is readily accomplished by, for example, filtration through sterile filtration membranes. Anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 is generally placed into a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper pierceable by a hypodermic injection needle.

Suitable emulsions may be prepared using commercially available fat emulsions, such as Intralipid™, Liposyn™, Infonutrol™, Lipofundin™ and Lipiphysan™. The active ingredient may be either dissolved in a pre-mixed emulsion composition or alternatively it may be dissolved in an oil (e.g., soybean oil, safflower oil, cottonseed oil, sesame oil, corn oil or almond oil) and an emulsion formed upon mixing with a phospholipid (e.g., egg phospholipids, soybean phospholipids or soybean lecithin) and water. It will be appreciated that other ingredients may be added, for example glycerol or glucose, to adjust the tonicity of the emulsion. Suitable emulsions will typically contain up to 20% oil, for example, between 5 and 20%. The fat emulsion can comprise fat droplets between 0.1 and 1.0 pm, particularly 0.1 and 0.5 pm, and have a pH in the range of 5.5 to 8.0. The emulsion compositions can be those prepared by mixing anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 with Intralipid™ or the components thereof (soybean oil, egg phospholipids, glycerol, and water).

Compositions for inhalation or insufflation include solutions and suspensions in pharmaceutically acceptable, aqueous, or organic solvents, or mixtures thereof, and powders. The liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as set out above. In some embodiments, the compositions are administered by the oral or nasal respiratory route for local or systemic effect. Compositions in preferably sterile pharmaceutically acceptable solvents may be nebulized by use of gases. Nebulized solutions may be breathed directly from the nebulizing device or the nebulizing device may be attached to a face mask, tent or intermittent positive pressure breathing machine. Solution, suspension, or powder compositions may be administered, preferably orally or nasally, from devices which deliver the formulation in an appropriate manner.

Dosing

Another embodiment concerns the dosage regimen for anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3. The dosage regimen for anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 is based on a variety of factors, including the type, age, weight, sex, and medical condition of the patient; the severity of the condition; the route of administration; and the activity of the compound employed. Thus, the dosage regimen may vary widely. For example, the total daily dose of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may typically range from about 0.001 to about 100 mg/kg (i.e., mg compound of the invention per kg body weight) for the treatment of the indicated conditions discussed herein. The total daily dose of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may also range from about 0.01 to about 30 mg/kg; from about 0.03 to about 10 mg/kg; and from about 0.1 to about 3. It is not uncommon that the administration of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 will be repeated a plurality of times in a day (typically no greater than 4 times). Multiple doses per day typically may be used to increase the total daily dose, if desired.

Another embodiment concerns the oral administration of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be provided in the form of tablets containing 0.1 , 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0, 30.0 50.0, 75.0, 100, 125, 150, 175, 200, 250, 300, 400, 500, and 600 milligrams of the active ingredient for the symptomatic adjustment of the dosage to the subject. In certain embodiments, the tablets may contain from about 100 mg to about 400 mg of the active agent. Another embodiment concerns the intravenous administration, of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may range from about 0.01 to about 10 mg/kg/minute during a constant rate infusion.

Methods of Treatment

Anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3, by virtue of their pharmacologic action, are useful for fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, and nonalcoholic steatohepatitis with cirrhosis and with hepatocellular carcinoma or with a metabolic-related disease, hypertriglyceridemia, atherosclerosis, angina, stroke, myocardial infarction, aortic vascular disease, renal vascular disease, cerebral vascular disease, heart failure, atrial fibrillation, coronary heart disease, obesity, dyslipidemia, hyper apo B lipoproteinemia, type 2 diabetes mellitus, glycemic control in patients with type 2 diabetes mellitus, fructose intolerance (e.g., hereditary fructose intolerance, intestinal fructose intolerance (also referred to as fructose non-absorption or fructose-malabsorption), and fructose-1 ,6-diphosphatase deficiency)), conditions of impaired glucose tolerance (IGT), conditions of impaired fasting plasma glucose, metabolic syndrome, syndrome X, hyperglycemia, hyperinsulinemia, insulin resistance, and/or impaired glucose metabolism.

One embodiment concerns a method of treating fructose intolerance or reducing the effects of fructose intolerance in a subject (e.g., human) in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

In certain embodiments, the fructose intolerance is hereditary fructose intolerance. In other embodiments, the fructose intolerance is intestinal fructose intolerance (fructose non-absorption or fructose malabsorption). In other embodiments the fructose intolerance is fructose-1 ,6- diphosphatase deficiency.

Another embodiment concerns a method of treating fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, and nonalcoholic steatohepatitis with cirrhosis and with hepatocellular carcinoma in a subject (e.g., human) in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1). Another embodiment concerns a method of treating hypertriglyceridemia, atherosclerosis, angina, stroke, myocardial infarction, aortic vascular disease, renal vascular disease, cerebral vascular disease, heart failure, atrial fibrillation, and coronary heart disease, wherein the method comprises administering to a subject (e.g., human) in need thereof a therapeutically effective amount of is anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

Another embodiment concerns a method of treating obesity, dyslipidemia, hyper apo B lipoproteinemia, type 2 diabetes mellitus, glycemic control in patients with type 2 diabetes mellitus, conditions of impaired glucose tolerance (IGT), conditions of impaired fasting plasma glucose, metabolic syndrome, syndrome X, hyperglycemia, hyperinsulinemia, insulin resistance, and/or impaired glucose metabolism, wherein the method comprises administering to the subject a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3- diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (triflu oromethy I) pyrimid in-4-y l}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

Another embodiment concerns a method of treating nonalcoholic fatty liver disease, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of anhydrous crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

Another embodiment concerns a method of treating nonalcoholic steatohepatitis, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of anhydrous crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

Another embodiment concerns a method of treating nonalcoholic steatohepatitis with liver fibrosis, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1).

In any of the above-mentioned embodiments, the anhydrous crystalline tris salt (Form 1) can be identified by its unique solid-state signatures with respect to, for example, powder X-ray diffraction (PXRD) data, solid state Nuclear Magnetic Resonance (ssNMR) data (e.g. ¹³C ssNMR data, ¹⁵N ssNMR data, and/or ¹⁹F ssNMR), and/or FT-Raman Spectroscopy data provided in any one of the above-mentioned embodiments. Another embodiment concerns the therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) is administered to a subject in need thereof in an amount from about 25mg to about 1000mg administered twice daily.

In another embodiment, the therapeutically effective amount of anhydrous crystalline 2- amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) is administered to a subject in need thereof in an amount from about 10Omg twice daily.

In another embodiment, the therapeutically effective amount of anhydrous crystalline 2- amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) is administered to a subject in need thereof in an amount from about 200mg twice daily.

In another embodiment, the therapeutically effective amount of anhydrous crystalline 2- amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) is administered to a subject in need thereof in an amount from about 300mg twice daily.

In another embodiment, the therapeutically effective amount of anhydrous crystalline 2- amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) is administered to a subject in need thereof in an amount from about 400mg twice daily.

In another embodiment, the therapeutically effective amount of anhydrous crystalline 2- amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (tris salt Form 1) is administered to a subject in need thereof in an amount from about 600mg twice daily.

Another embodiment concerns a method of treating fructose intolerance or reducing the effects of fructose intolerance, the method comprising in a subject (e.g., human) in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2- [(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

In certain embodiments, the fructose intolerance is hereditary fructose intolerance. In other embodiments, the fructose intolerance is intestinal fructose intolerance (fructose non-absorption or fructose malabsorption). In other embodiments the fructose intolerance is fructose-1 ,6- diphosphatase deficiency. Another embodiment concerns a method of treating treating fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, and nonalcoholic steatohepatitis with cirrhosis and with hepatocellular carcinoma in a subject (e.g., human) in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

Another embodiment concerns a method of treating hypertriglyceridemia, atherosclerosis, angina, stroke, myocardial infarction, aortic vascular disease, renal vascular disease, cerebral vascular disease, heart failure, atrial fibrillation, and coronary heart disease, wherein the method comprises administering to a subject (e.g., human) in need thereof a therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

Another embodiment concerns a method of treating obesity, dyslipidemia, hyper apo B lipoproteinemia, type 2 diabetes mellitus, glycemic control in patients with type 2 diabetes mellitus, conditions of impaired glucose tolerance (IGT), conditions of impaired fasting plasma glucose, metabolic syndrome, syndrome X, hyperglycemia, hyperinsulinemia, insulin resistance, and/or impaired glucose metabolism, wherein the method comprises administering to the subject a therapeutically effective amount of monohydrate crystalline 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

Another embodiment concerns a method of treating nonalcoholic fatty liver disease, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of monohydrate crystalline 2-amino- 2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

Another embodiment concerns a method of treating nonalcoholic steatohepatitis, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of monohydrate crystalline 2-amino- 2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

Another embodiment concerns a method of treating nonalcoholic steatohepatitis with liver fibrosis, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2).

In any of the above-mentioned embodiments, the crystalline monohydrate form (tris salt Form 2) administered to the subject can be identified by its unique powder X-ray diffraction (PXRD) data, and/or FT-Raman Spectroscopy data provided in Figure 6 and/or Figure 7.

In any of the above-mentioned embodiments, the therapeutically effective amount of crystalline monohydrate 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2- [(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2) is administered to a subject in need thereof in an amount from about 25mg to about 1000mg administered twice daily.

In another embodiment, the therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2) is administered to a subject in need thereof in an amount from about 10Omg twice daily.

In another embodiment, the therapeutically effective amount of mononhydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2) is administered to a subject in need thereof in an amount from about 200mg twice daily.

In another embodiment, the therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2) is administered to a subject in need thereof in an amount from about 300mg twice daily.

In another embodiment, the therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2) is administered to a subject in need thereof in an amount from about 400mg twice daily.

In another embodiment, the therapeutically effective amount of monohydrate crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]- 6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid (tris salt Form 2) is administered to a subject in need thereof in an amount from about 600mg twice daily.

Another embodiment concerns a method of treating fructose intolerance or reducing the effects of fructose intolerance, in a subject (e.g., human) in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

Another embodiment concerns a method of treating treating fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, and nonalcoholic steatohepatitis with cirrhosis and with hepatocellular carcinoma in a subject (e.g., human) in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

Another embodiment concerns a method of treating hypertriglyceridemia, atherosclerosis, angina, stroke, myocardial infarction, aortic vascular disease, renal vascular disease, cerebral vascular disease, heart failure, atrial fibrillation, and coronary heart disease, wherein the method comprises administering to a subject (e.g., human) in need thereof a therapeutically effective amount of 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

Another embodiment concerns a method of treating obesity, dyslipidemia, hyper apo B lipoproteinemia, type 2 diabetes mellitus, glycemic control in patients with type 2 diabetes mellitus, conditions of impaired glucose tolerance (IGT), conditions of impaired fasting plasma glucose, metabolic syndrome, syndrome X, hyperglycemia, hyperinsulinemia, insulin resistance, and/or impaired glucose metabolism, wherein the method comprises administering to the subject a therapeutically effective amount of 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

Another embodiment concerns a method of treating nonalcoholic fatty liver disease, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of 2-amino-2-(hydroxymethyl) propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl) pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3). Another embodiment concerns a method of treating nonalcoholic steatohepatitis, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of 2-amino-2-(hydroxymethyl) propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl) pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

Another embodiment concerns a method of treating nonalcoholic steatohepatitis with liver fibrosis, in a subject in need thereof, wherein the method provides for the administration to a subject (e.g., human) in need thereof a therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3).

In any of the above-mentioned embodiments, the therapeutically effective amount of 2- amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3) is administered to a subject in need thereof in an amount from about 25mg to about 1000mg administered twice daily.

In another embodiment, the therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3) is administered to a subject in need thereof in an amount from about 100mg twice daily.

In another embodiment, the therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3) is administered to a subject in need thereof in an amount from about 200mg twice daily.

In another embodiment, the therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3) is administered to a subject in need thereof in an amount from about 300mg twice daily.

In another embodiment, the therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form 3) is administered to a subject in need thereof in an amount from about 400mg twice daily.

In another embodiment, the therapeutically effective amount of 2-amino-2- (hydroxymethyl)propane-l ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo [3.1 ,0]hex-6-yl]acetic acid in amorphous form (Form

3) is administered to a subject in need thereof in an amount from about 600mg twice daily.

Co-administration and Combination Therapy

Anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 can be used alone, or in combination with other therapeutic agents. The invention provides any of the uses, methods or compositions as defined herein wherein anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 of any embodiment herein is used in combination with one or more other therapeutic agent discussed herein. This would include a pharmaceutical combination (e.g. a pharmaceutical composition) for the treatment of a disease or condition for which a KHK inhibitor is indicated, comprising anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3, as defined in any of the embodiments described herein, and one or more other therapeutic agent discussed herein.

The administration of two or more compounds “in combination” means that all the compounds are administered closely enough in time that each may generate a biological effect in the same time frame. The presence of one agent may alter the biological effects of the other compound(s). The two or more compounds may be administered simultaneously, concurrently sequentially, or separately (optionally with different dosing cycles). Additionally, simultaneous administration may be carried out by mixing the compounds prior to administration (or using a fixed dose combination) or by administering the compounds at the same point in time but as separate dosage forms (and optionally by different administration routes) at the same or different site of administration.

The phrases “concurrent administration,” “co-administration,” “simultaneous administration,” and “administered simultaneously” mean that the compounds are administered in combination.

Another embodiment concerns methods of treatment that include administering anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 in combination with one or more other pharmaceutical agents, wherein the one or more other pharmaceutical agents may be selected from the agents discussed herein.

Anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3, and the additional pharmaceutical agents can be combined with pharmaceutically acceptable vehicles such as saline, Ringer’s solution, dextrose solution, and the like. The dosage regimen, i.e., dose, timing, and repetition, will depend on the individual and that individual’s medical history.

Anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 may be coadministered with at least one other pharmaceutical agent, wherein the agent is selected from the group consisting of an acetyl-CoA carboxylase- (ACC) inhibitor, a farnesoid X receptor agonist (e.g., tropifexor, obeticholic acid), a diacylglycerol O-acyltransferase 1 (DGAT-1) inhibitor, monoacylglycerol O-acyltransferase inhibitors, a phosphodiesterase (PDE)-10 inhibitor, an AMPK activator, a sulfonylurea, a meglitinide, an a-amylase inhibitor, an a-glucoside hydrolase inhibitor, an a-glucosidase inhibitor, a PPARy agonist, a PPAR a/y agonist (e.g., elafibranor), a biguanide, a glucagon-like peptide 1 (GLP-1) modulator, liraglutide, albiglutide, exenatide, albiglutide, lixisenatide, dulaglutide, semaglutide, a protein tyrosine phosphatase-1 B (PTP-1 B) inhibitor, SIRT-1 activator, a dipeptidyl peptidase IV (DPP-IV) inhibitor, an insulin secreatagogue, a fatty acid oxidation inhibitor, an A2 antagonist, a c-jun amino-terminal kinase (JNK) inhibitor, glucokinase activators (GKa), insulin, an insulin mimetic, a glycogen phosphorylase inhibitor, a VPAC2 receptor agonist, SGLT2 inhibitors, a glucagon receptor modulator, GPR119 modulators, FGF21 derivatives or analogs, TGR5 receptor modulators, GPBAR1 receptor modulators, GPR40 agonists, GPR120 modulators, high affinity nicotinic acid receptor (HM74A) activators, SGLT1 inhibitors, inhibitors or modulators of carnitine palmitoyl transferase enzymes, inhibitors of fructose 1 ,6-diphosphatase, inhibitors of aldose reductase, mineralocorticoid receptor inhibitors, inhibitors of TORC2, inhibitors of CCR2 and/or CCR5, inhibitors of PKC isoforms (e.g. PKCa, PKCp, PKCY), inhibitors of fatty acid synthetase, inhibitors of serine palmitoyl transferase, modulators of GPR81 , GPR39, GPR43, GPR41 , GPR105, Kv1.3, retinol binding protein 4, glucocorticoid receptor, somatostatin receptors, inhibitors or modulators of PDHK2 or PDHK4, inhibitors of MAP4K4, modulators of IL1 family including ILI beta, HMG-CoA reductase inhibitors, squalene synthetase inhibitors, fibrates, bile acid sequestrants, ACAT inhibitors, MTP inhibitors, lipooxygenase inhibitors, cholesterol absorption inhibitors, PCSK9 modulators, cholesteryl ester transfer protein inhibitors and modulators of RXRalpha.

Additional pharmaceutical agents, includes at least one other pharmaceutical agent, wherein the agent is selected from the group consisting of cysteamine or a pharmaceutically acceptable salt thereof, cystamine or a pharmaceutically acceptable salt thereof, an anti-oxidant compound, lecithin, vitamin B complex, a bile salt preparations, an antagonists of Cannabinoid-1 (CB1) receptor, an inverse agonists of Cannabinoid-1 (CB1) receptor, a peroxisome proliferator- activated receptor) activity regulators, a benzothiazepine or benzothiepine compound, an RNA antisense construct to inhibit protein tyrosine phosphatase PTPRU, a heteroatom-linked substituted piperidine and derivatives thereof, an azacyclopentane derivative capable of inhibiting stearoyl-coenzyme alpha delta-9 desaturase, acylamide compound having secretagogue or inducer activity of adiponectin, a quaternary ammonium compound, Glatiramer acetate, pentraxin proteins, a HMG-CoA reductase inhibitor, n-acetyl cysteine, isoflavone compound, a macrolide antibiotic, a galectin inhibitor, an antibody, or any combination of thereof. When any one of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 according to any one of the above-mentioned embodiments are co-administered with an additional pharmaceutically active agent, the combination may be a fixed-dose combination.

When any one of anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 according to any one of the above-mentioned embodiments are co-administered with an additional pharmaceutically active agent, the combination may not be a fixed-dose combination, such that each pharmaceutically active ingredient is administered separately.

KITS

Another aspect of the invention provides kits comprising anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3. A kit may include, in addition to anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 or pharmaceutical composition thereof, diagnostic, or therapeutic agents. A kit may also include instructions for use in a diagnostic or therapeutic method. In some embodiments, the kit includes anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 and a diagnostic agent. In other embodiments, the kit includes anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3, or a pharmaceutical composition thereof.

In yet another embodiment, the invention comprises kits that are suitable for use in performing the methods of treatment described herein. In one embodiment, the kit contains a first dosage form comprising anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 in quantities sufficient to carry out the methods of the invention. In another embodiment, the kit comprises anhydrous tris salt Form 1 , monohydrate tris salt Form 2, or amorphous Form 3 in quantities sufficient to carry out the methods of the invention and a container for the dosage and a container for the dosage.

PREPARATION

The compounds exemplified below may be prepared by the general and specific methods described below, coupled with the common general knowledge of one skilled in the art of synthetic organic chemistry and/or solid forms of pharmaceutical compounds. Such common general knowledge can be found in standard reference books such as Comprehensive Organic Chemistry, Ed. Barton and Ollis, Elsevier; Comprehensive Organic Transformations: A Guide to Functional Group Preparations, Larock, John Wiley and Sons; and Compendium of Organic Synthetic Methods, Vol. I-XII (published by Wiley-lnterscience). The starting materials used herein are commercially available or may be prepared by routine methods known in the art. In the preparation of the compounds exemplified below, it is noted that some of the preparation methods described herein may require protection of remote functionality (e.g., primary amine, secondary amine, carboxyl in precursors). The need for such protection will vary depending on the nature of the remote functionality and the conditions of the preparation methods. The need for such protection is readily determined by one skilled in the art. The use of such protection/deprotection methods is also within the skill in the art. For a general description of protecting groups and their use, see T.W. Greene, Protective Groups in Organic Synthesis, John Wiley & Sons, New York, 1991.

For example, certain compounds contain primary amines or carboxylic acid functionalities which may interfere with reactions at other sites of the molecule if left unprotected. Accordingly, such functionalities may be protected by an appropriate protecting group which may be removed in a subsequent step. Suitable protecting groups for amine and carboxylic acid protection include those protecting groups commonly used in peptide synthesis (such as N-t-butoxycarbonyl (Boc), benzyloxycarbonyl (Cbz), and 9-fluorenylmethylenoxycarbonyl (Fmoc) for amines and lower alkyl or benzyl esters for carboxylic acids) which are generally not chemically reactive under the reaction conditions described and can typically be removed without chemically altering other functionality in the compounds.

The descriptions below are intended to provide a general description of the methodology employed in the preparation of anhydrous tris salt (Form 1) and tris salt monohydrate (Form 2). The compounds may contain single or multiple chiral centers with the stereochemical designation (R) or (S). It will be apparent to one skilled in the art that all the synthetic transformations can be conducted in a similar manner whether the materials are enantioenriched or racemic. Moreover, the resolution to the desired optically active material may take place at any desired point in the sequence using well known methods such as described herein and in the chemistry literature. For example, intermediates and finals may be separated using chiral chromatographic methods. Alternatively, chiral salts may be utilized to isolate enantiomerically enriched intermediates and final compounds.

EXAMPLES

The following illustrate the synthesis of free acid, crystalline anhydrous tris salt Form 1 , crystalline tris salt monohydrate and amorphous tris salt of the present invention.

G or g is gram, and mg means milligram.

H or h means hour.

IPA means isopropyl alcohol.

L is liter. mL is milliliter.

MCC means microcrystalline cellulose.

RT or rt means room temperature which is the same as ambient temperature (about 20 to 25 °C).

¹H Nuclear magnetic resonance (NMR) spectra were in all cases consistent with the proposed structures. Characteristic chemical shifts (8) are given in parts-per-million (ppm) relative to the residual proton signal in the deuterated solvent (CHCI₃ at 7.27 ppm; CD₂HOD at 3.31 ppm) and are reported using conventional abbreviations for designation of major peaks: e.g. s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; br, broad. ssNMR means solid-state NMR.

PXRD means Powder X-ray Diffraction.

RH means relative humidity.

Crystalline Free Acid Form 1

Example 1. Preparation of crystalline free acid of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1.0]hex-6-yl]acetic acid.

To a stirred solution of unpurified methyl [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6- (trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetate in methanol (650 mL) was added a solution of sodium hydroxide (35.1 g, 877 mmol) in water (70 mL) in small portions under stirring at 5 °C to 15 °C. The mixture became clear in 30 min. The clear solution was stirred at RT for 3 h, then concentrated to ~1/3 of the initial volume and the residue was diluted with water (750 mL) and brine (250 mL), then washed with a mixture of MTBE (260 mL) and heptane (130 mL). The organic wash was discarded. The aqueous phase was washed with MTBE-heptane (2:1) mixture (2 x 300 mL) and the organic layers discarded. The aqueous layer was then combined with MTBE (250 mL) and heptane (250 mL) and cooled to 0 °C. Slowly under stirring at 0 °C to 4 °C, 6 M aq. HCI (130 mL) was added, followed by 1 M aq. KHSO₄ (150 mL), and the obtained mixture was stirred for 15 min. The organic phase was separated, and the aqueous phase was additionally extracted with a mixture of MTBE (170 mL) and heptane (170 mL). The combined organic extract was washed with water-brine (1 :1) mixture (150 mL), dried over anhydrous MgSO₄ (60 g) and SiO₂ (60 g), filtered, and concentrated to give a colorless oil. It was combined (as a concentrated solution in MTBE) with another batch, which was prepared using identical conditions on the same scale. The combined MTBE solution was concentrated in vacuo, then heptane (2000 mL) was added, and the suspension was concentrated again, with gradual increase of vacuum to obtain the desired product (406.0 g). A portion of this material (196 g) was dissolved in MTBE (220 mL) at 60 °C to 63 °C, stirred slowly, and heptane (1500 mL) was added at 55 °C to 60 °C. The mixture was seeded with crystalline title compound (50 mg). The mixture was stirred at 60 °C for 30 min, then additional heptane (1700 mL) was added during 20 min. The heterogeneous mixture was stirred at 60 °C for 2 h and then slowly cooled to 25 °C and stirred for 20 h. A small amount of solid was stuck on the flask walls and easily moved into the liquid phase with a spatula and the mixture was further stirred at 25 °C for 24 h. The solids were filtered off, washed with 5% MTBE in heptane, and dried in vacuo at 50 °C for 48 h to obtain Example 4 as a white crystalline solid (178.2 g, 73% over 3 steps). Crystalline solid of Example 4 has also been obtained using similar purification conditions without seeding.

MP: 122-123 °C, [a]_D +86.3° (CDCI₃, c = 1 .37). MS(ES+): 357.3 (M+H). ¹H NMR (400 MHz, CDCI₃) 5: 10.84 (br. s, 1 H), 5.92 (s, 1 H), 4.38-4.51 (m, 1 H), 3.89-4.10 (m, 3H), 3.53-3.66 (m, 1 H), 3.41- 3.53 (m, 2H), 2.30-2.46 (m, 3H), 1 .94 (ddt, 1 H), 1 .55-1 .63 (m, 2H), 1 .50 (d, 3H), 0.94 (m, 1 H).

Anhydrous Crystalline Tris Salt Form 1

Example 2. Preparation of anhydrous tris salt Form 1 of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1.0]hex-6-yl]acetic acid.

Crystalline free acid of Example 1 (528.9mg; 1.48mmol) was dissolved in nitromethane (12.5mL; 24vol). One equivalent of solid tris (tromethamine) (181 mg; 1.49mmol) was added to the solution followed by seed crystals of tris salt. The suspension was heated to 60°C for 1 hour, cooled at 0.1 °C/min to 20°C, and stirred at 20°C overnight (~16 hours). The solids were isolated by filtration under vacuum and air-dried for 1 hour under a nitrogen blanket. PXRD analysis of the wet cake was consistent with a non-solvated form of the tris salt. The solids were dried in a vacuum oven at 40°C with nitrogen bleed for 3 hours. The yield was 97.0% (687mg; 1 .44mmol) of the anhydrous tris salt Form 1 .

Example 3. Powder X-Ray Diffraction Analysis of anhydrous tris salt Form 1 of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid.

The powder X-ray diffraction pattern was generated using a Bruker AXS D8 Endeavor diffractometer equipped with a Cu radiation source. The tube voltage and amperage were set to 40 kV and 40 mA, respectively. The motorized divergence slits were set at constant illumination of 11 mm. Diffracted radiation was detected using a LYNXEYE XE-T energy dispersive X-ray detector, with the position sensitive detector (PSD) opening set at 4.00°. Data was collected on the theta-theta goniometer at the Cu wavelength from 2.0 to 55.0 degrees 2-theta (°20) using a step size of 0.019 °20 and a time per step of 0.2 seconds. Samples were prepared for analysis by placing them in a silicon low background cavity holder and rotated at 15 rpm during data collection. Data were analyzed in DIFFRAC.EVA V5.0 software. Peak lists were prepared using reflections with a relative intensity > 5 % of the most intense band in each respective diffraction pattern. A typical error of ± 0.1 °20 in peak positions applies to this data. The minor error associated with this measurement can occur because of a variety of factors including: (a) sample preparation (e.g.sample height), (b) instrument characteristics, (c) instrument calibration, (d) operator input (e.g. in determining the peak locations), and (e) the nature of the material (e.g. preferred orientation and transparency effects).

To obtain the absolute peak positions, the powder pattern should be aligned against a reference. This could either be the simulated powder pattern from the crystal structure of the same form solved at room temperature, or an internal standard e.g. silica or corundum. The collected powder pattern of anhydrous tris salt Form 1 was aligned to the powder pattern of the same material containing internal standard, Si (SRM 640e).

For PXRD, the relative intensities of the peaks can vary, depending upon, for example, the sample preparation technique and the sample mounting procedure. Moreover, instrument variation and other factors can often affect the 2-theta values.

Anhydrous tris salt Form 1 is a highly crystalline, high melting, non-hygroscopic solid form with aqueous solubility. Anhydrous tris salt Form 1 is thermodynamically stable at a water activity (aw) of about 0.7 and below. Anhydrous tris salt Form 1 has a PXRD pattern substantially the same as that shown in FIG. 1. Peak locations and intensities for the PXRD pattern in FIG. 1 are provided in Table 1 .

Table 1 - PXRD Peaks and Relative Intensities of Anhydrous Tris Salt Form 1

Example 4. Raman spectroscopy Analysis of Anhydrous Tris Salt Form 1

Raman spectra were collected using a RAM II FT-Raman module attached to a Vertex 70 spectrometer (Bruker Optik GmbH). The instrument is equipped with a 1064 nm solid-state (Nd:YAG) laser and a liquid nitrogen cooled germanium detector. Prior to data acquisition, instrument performance and calibration verifications were conducted using a white light source, and polystyrene and naphthalene references. Samples were prepared and analysed in truncated NMR tubes. A sample rotator (Ventacon, UK) was used during measurement to maximise the volume of material exposed to the laser during data collection. The backscattered Raman signal from the sample was optimized and data were collected at a spectral resolution of 2 cm ¹ using a laser power of 750 mW. A Blackmann-Harris 4-term apodization function was applied to minimise spectral aberrations. Spectra were generated between 3500 and 50 cm ¹ with the number of scans adjusted accordingly to ensure adequate signal to noise.

Spectra were normalised by setting the intensity of the most intense peak to 2.00. Peaks were then identified using the automatic peak picking function in the OPUS v8.2 software (Bruker Optik GmbH) with the sensitivity set to 2.5%. Peak positions and relative peak intensities were extracted and tabulated. The variability in the peak positions with this experimental configuration is within ±2 cur¹.

It is expected that, since FT-Raman and dispersive Raman are similar techniques, peak positions reported in this document for FT-Raman spectra would be consistent with those which would be observed using a dispersive Raman measurement, assuming appropriate instrument calibration. Peak locations and intensities for the Raman spectra in FIG. 2 are provided in Table 2.

Table 2 - Raman Peaks and Normalized Intensity of Anhydrous Tris Salt Form 1

Example 5. Solid State NMR Analysis of Anhydrous Tris Salt Form 1

Solid state NMR (ssNMR) analysis was conducted on a Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹³C and ¹⁵N ssNMR spectra were collected on a 4 mm MAS probe at a magic angle spinning rate of 10 or 8 kHz, respectively, and ¹⁹F spectra were recorded with a 3.2 mm MAS probe at a spin rate of 20 kHz. The temperature regulated to 20°C in each case. ¹³C cross-polarization (CP) spectra (see Figure 3 and Table 3) with TOSS spinning sideband suppression were recorded with a 10 ms CP contact time and recycle delay of 5 seconds. Carbon spectral referencing is relative to neat tetramethylsilane, carried out by setting the high-frequency signal from an external sample of adamantane to 38.5 ppm.

¹⁵N CP spectra (see Figure 4 and Table 4) were recorded with a 10 ms CP contact time and a recycle delay of 5 seconds. Nitrogen spectral referencing is relative to neat nitromethane, carried out by setting the signal from an external sample of glycine to -346.8 ppm.

¹⁹F spectra (see Figure 5 and Table 5) were collected using direct excitation with proton decoupling and a 30 s recycle delay. Fluorine spectral referencing is relative to CFCI₃, carried out by setting the signal from an external sample of 50% v/v trifluoroacetic acid in H₂O to -76.54 ppm.

Automatic peak picking was performed using ACD Labs 2019 Spectrus Processor software with a threshold value of 3% relative intensity used for preliminary peak selection. The output of the automated peak picking was visually checked to ensure validity and adjustments were manually made if necessary. Although specific ¹³C, ¹⁵N and ¹⁹F ssNMR peak values are reported herein there does exist a range for these peak values due to differences in instruments, samples, and sample preparation. A typical variability for ¹³C, ¹⁵N and ¹⁹F chemical shift x-axis values is on the order of plus or minus 0.2 ppm for a crystalline solid. The ssNMR peak heights reported herein are relative intensities. The ssNMR intensities can vary depending on the actual setup of the experimental parameters and the thermal history of the sample.

Anhydrous tris salt Form 1 has a ¹³C ssNMR spectrum substantially the same as that shown in FIG. 3. Form 1 has characteristic ¹³C ssNMR chemical shifts, expressed as ppm, at 55.8, 58.3, 63.7, 64.2, and 183.6, + 0.2 ppm. ¹³C chemical shifts (+ 0.2 ppm) of Form 1 as shown in FIG. 3 are listed in Table 3.

Table 3 - ¹³C chemical shifts and Intensity of Anhydrous Tris Salt Form 1

Anhydrous tris salt Form 1 has a ¹⁵N ssNMR spectrum substantially the same as that shown in FIG. 4. Form 1 has characteristic ¹⁵N ssNMR chemical shifts, expressed as ppm, at 177.5, -278, -285.9, and -342.1 , + 0.2 ppm. ¹⁵N chemical shifts (+ 0.2 ppm) of Form 1 as shown in FIG. 4 are listed in Table 4.

Table 4 - ¹⁵N chemical shifts and Intensity of Anhydrous Tris Salt Form 1

Table 5. ¹⁹F ssNMR peak list for Anhydrous Tris Salt Form 1

Example 6. Scale Up Preparation of anhydrous tris salt Form 1 of [(1 R,5S,6R)-3- {2-[(2S)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1.0]hex-6- yl]acetic acid.

Crystalline free acid of Example 1 (3999 mg; 11 .2 mmol) and one equivalent of solid tris (tromethamine) (1360 mg; 11.2 mmol) was slurried in nitromethane (94.5 mL; 24 vol) and mixed for 10 minutes. Seeds of the anhydrous tris salt Form 1 of Example 2 were added. The suspension was heated to 60 °C and mixed for 15 min and then seeded again. The suspension was mixed for a total of 2 hours and then allowed to cool to RT. Mixing was continued at RT for 2 hours. The batch was filtered for 1 hour (5276mg). The batch solids were dried in a vacuum oven at 40 °C with nitrogen bleed for 17 hours (5.24 g; 97.8% yield relative to salt). The batch was determined to be 99.9% area (234 nm) pure by HPLC. Example 7. Alternative Preparation of anhydrous tris salt Form 1 of [(1 R,5S,6R)- 3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1.0]hex- 6-yl]acetic acid.

800mg of crystalline free acid of Example 1 was added to an 8-dram vial with 300mg tris (tromethamine). 14.8ml heptane and 7.4ml ethanol was added. The mixture was stirred on a whirly mixer and split into two 4-dram vials and magnetic stirrers were added. The vials were decapped and left to evaporate at room temperature for a week. The resulting material was a clear film with a small amount solid white material. Water was added (5ml) to both vials and the material was slurried at room temperature on a roller mixer for an hour. The two slurries were combined into one lot and evaporated in a vacuum oven at 60°C over the course of 24 hours.

Analytical Data Anhydrous Tris Salt Form 1

Anhydrous crystalline tris salt Form 1 is a highly crystalline, anhydrous, non-hygroscopic solid form that exhibits favorable aqueous stability, and is both physically and chemically stable. Anhydrous crystalline tris salt Form 1 has a high melting point (139.5 °C).

Anhydrous tris salt Form 1 is kinetically stable with respect to hydration at high humidity, as the sorption value at 90% RH shows it has not hydrated (See: Table 6 below).

Table 6. Water Sorption of anhydrous tris salt Form 1

In addition to water sorption testing, Anhydrous tris salt Form 1 chemical stability was also tested. The material was exposed to 70°C/75% RH for 1 week, without any significant degradation being observed, though a colour change from white to off-white was seen. The material was also subjected to 1 xlCH light exposure, which did result in some degradation, with purity dropping from 99.9% to 99.4%. Chemical stability of anhydrous tris salt Form 1 was tested against and found to be more stable than the free acid Form 1 (Example 1) under light, as the free acid purity dropped to 99.1 % under the same conditions.

An ASAP stability study of anhydrous tris salt Form 1 was carried out, with 50°C/75% RH, 60°C/40% RH, 70°C/10% RH, 70°C/75% RH and 80°C/40% RH conditions. One degradant was observed at the harshest conditions (70°C/75% RH and 80°C/40% RH) but only grew to 0.14%, so the shelf life is predicted to be greater than 2 years at 25°C/60% RH.

Crystalline Monohydrate Tris Salt Form 2

Example 8. Preparation of crystalline monohydrate tris salt Form 2 of [(1 R,5S,6R)- 3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1.0]hex- 6-yl]acetic acid.

Crystalline free acid of Example 1 (1000 mg; 2.8 mmol) was mixed with nitromethane (23.1 mL; 23 vol). One equivalent of tris (tromethamine) from a 3 M solution in water (0.936 mL; 2.8 mmol) was added to the solution followed by seed crystals (3-5 mg) of crystalline monohydrate tris salt Form 2. The suspension was heated to 60 °C and mixed for 15 min and then seeded again. The suspension was mixed for a total of 2 hours and then then cooled to 20 °C at a rate of 0.1 °C/min and held at this temperature overnight. The next day, the suspension was vortexed to produce a free-flowing slurry. The slurry (Batch #1) was saved to be mixed with a second slurry (Batch #3) to prepare the final monohydrate tris salt Form 2.

Crystalline free acid of Example 1 (506.7 mg; 1.4 mmol) was mixed with nitromethane (11 .5 mL; 23 vol). One equivalent of tris (tromethamine) from a 3 M solution in water (0.474 mL; 1 .4 mmol) was added to the solution followed by seed crystals (2-3 mg) of crystalline monohydrate tris tris salt Form 2. The suspension was heated to 60 °C and mixed for 5 min and then seeded again. The suspension was mixed for a total of 2 hours and then then cooled to 20 °C at a rate of 0.1 °C/min and held at this temperature overnight. The next day, the suspension was vortexed to produce a free-flowing slurry (Batch #3).

The slurry Batch #3 was mixed with slurry Batch #1 and filtered for 5 hours and isolated (1 .88 g; 89.8% yield relative to salt). The isolate was determined to be 99.9% area (234 nm) pure by HPLC. Proton NMR confirmed that the isolate was a mono-tris salt Form 2. The PXRD pattern of Form 2 is shown in Figure 6. The Raman spectra for Form 2 is shown in Figure 7. Example 9. Alternative Preparation of crystalline monohydrate tris salt Form 2 of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 .0]hex-6-yl]acetic acid.

1 g of anhydrous tris salt Form 1 was weighed into a 4-dram vial, to which was added 1 ml of purified water. A magnetic stirrer was added to the vial and it was placed on a temperature- controlled stirrer block, and set to stir at 500 RPM. The block was set to heat to 25°C at 1 °C/min, cool to 15°C at 0.1 °C/min, hold for 10 minutes at 15°C, and then continue to repeat this cycle. After 17 hours the solid material in the vial had converted to tris salt monohydrate Form 2.

Example 10. Powder X-Ray Diffraction Analysis of monohydrate tris salt Form 2 of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo[3.1 .0]hex-6-yl]acetic acid.

To obtain the absolute peak positions, the powder pattern should be aligned against a reference. This could either be the simulated powder pattern from the crystal structure of the same form solved at room temperature, or an internal standard e.g. silica or corundum. The collected powder pattern of monohydrate tris salt Form 2 was aligned to the powder pattern of the same material containing internal standard, Si (SRM 640e). For PXRD, the relative intensities of the peaks can vary, depending upon, for example, the sample preparation technique and the sample mounting procedure. Moreover, instrument variation and other factors can often affect the 2-theta values.

Monohydrate tris salt Form 2 has a PXRD pattern substantially the same as that shown in FIG. 6. Peak locations and intensities for the PXRD pattern in FIG. 6 are provided in Table 7.

Table 7 - PXRD Peaks and Relative Intensities of Monohydrate Tris Salt Form 2

Example 11. Raman spectroscopy Analysis of Monohyrate Tris Salt Form 2

Raman spectra were collected using a RAM II FT-Raman module attached to a Vertex 70 spectrometer (Bruker Optik GmbH). The instrument is equipped with a 1064 nm solid-state (Nd:YAG) laser and a liquid nitrogen cooled germanium detector. Prior to data acquisition, instrument performance and calibration verifications were conducted using a white light source, and polystyrene and naphthalene references. Samples were prepared and analyzed in truncated NMR tubes. A sample rotator (Ventacon, UK) was used during measurement to maximize the volume of material exposed to the laser during data collection. The backscattered Raman signal from the sample was optimized and data were collected at a spectral resolution of 2 cm ¹ using a laser power of 1000 mW. A Blackmann-Harris 4-term apodization function was applied to minimize spectral aberrations. Spectra were generated between 3500 and 50 cm ¹ with the number of scans adjusted accordingly to ensure adequate signal to noise.

Spectra were normalized by setting the intensity of the most intense peak to 2.00. Peaks were then identified using the automatic peak picking function in the OPUS v8.2 software (Bruker Optik GmbH) with the sensitivity set to 2.5%. Peak positions and relative peak intensities were extracted and tabulated. The variability in the peak positions with this experimental configuration is within ±2 cur¹.

It is expected that, since FT-Raman and dispersive Raman are similar techniques, peak positions reported in this document for FT-Raman spectra would be consistent with those which would be observed using a dispersive Raman measurement, assuming appropriate instrument calibration. Peak locations and intensities for the Raman spectra in FIG. 7 are provided in Table 8.

Table 8 - Raman Peaks and Normalized Intensity of Monohydrate Tris Salt Form 2

Example 12. Solid State NMR Analysis of Monohydrate Tris Salt Form 2

Solid state NMR (ssNMR) analysis was conducted on a Bruker Avance III HD 400 MHz (¹H frequency) NMR spectrometer. ¹³C and ¹⁵N ssNMR spectra were collected on a 4 mm MAS probe at a magic angle spinning rate of 10 or 8 kHz, respectively, and ¹⁹F spectra were recorded with a 3.2 mm MAS probe at a spin rate of 20 kHz. The temperature regulated to 20°C in each case. ¹³C cross-polarization (CP) spectra (see Figure 8 and Table 9) with TOSS spinning sideband suppression were recorded with a 4 ms CP contact time and recycle delay of 1.5 seconds. Carbon spectral referencing is relative to neat tetramethylsilane, carried out by setting the high-frequency signal from an external sample of adamantane to 38.5 ppm.

¹⁵N CP spectra (see Figure 9 and Table 10) were recorded with a 10 ms CP contact time and a recycle delay of 1 .5 seconds. Nitrogen spectral referencing is relative to neat nitromethane, carried out by setting the signal from an external sample of glycine to -346.8 ppm.

¹⁹F spectra (see Figure 10 and Table 11) were collected using direct excitation with proton decoupling and a 4 s recycle delay. Fluorine spectral referencing is relative to CFCI₃, carried out by setting the signal from an external sample of 50% v/v trifluoroacetic acid in H₂O to -76.54 ppm.

Monohydrate tris salt Form 2 has a ¹⁵N ssNMR spectrum substantially the same as that shown in FIG. 8. Form 2 has characteristic ¹³C ssNMR chemical shifts, expressed as ppm, at 42.4, 46.7, 62.4,91.2, and 164.6, + 0.2 ppm. ¹³C chemical shifts (+ 0.2 ppm) of tris salt Form 2 as shown in FIG. 8 are listed in Table 9. Table 9 - ¹³C chemical shifts and Intensity of Monohydrate Tris Salt Form 2

Monohydrate tris salt Form 2 has a ¹⁵N ssNMR spectrum substantially the same as that shown in FIG. 9. Form 2 has characteristic ¹⁵N ssNMR chemical shifts, expressed as ppm, at 172.4, -174.3, and -342.8, + 0.2 ppm. ¹⁵N chemical shifts (+ 0.2 ppm) of Form 2 as shown in FIG. 9 are listed in Table 10.

Table 10 - ¹⁵N chemical shifts and Intensity of Monohydrate Tris Salt Form 2

Monohydrate tris salt Form 2 has a ¹⁹F ssNMR spectrum substantially the same as that shown in FIG. 10. Form 2 has a characteristic ¹⁹F ssNMR chemical shift, expressed as ppm, at -70.4. ¹⁹F chemical shifts (+ 0.2 ppm) of Form 2 as shown in FIG. 10 are listed in Table 11 . Table 11. ¹⁹F ssNMR peak list for Monohydrate Tris Salt Form 2

Analytical Data Crystalline Monohydrate Tris Salt Form 2

Crystalline Monohydrate Tris Salt Form 2 is a metastable with respect to crystalline anhydrous tris salt Form 1 at water activities of 0.7 aw and below, but more stable at activities of 0.8 aw and above at room temperature.

Throughout this application, various publications are referenced. The disclosures of these publications in their entireties are hereby incorporated by reference into this application for all purposes. It will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the scope or spirit of the invention. Other embodiments of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the invention being indicated by the following claims.

Claims

52 What is claimed is:

1. A crystalline form that is crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo

[3.1 ,0]hex-6-yl]acetic acid.

2. The crystalline form of claim 1 , wherein the crystalline form is an anhydrous crystalline salt.

3. The anhydrous crystalline salt of claim 2, wherein the ratio of [(1 R,5S,6R)-3-{2-[(2S)-2- methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid to 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt is 1 :1.

4. The anhydrous crystalline salt of claim 3 having a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, with at least one characteristic peak expressed in degrees 20 selected from 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21 .8 ± 0.1 .

5. The anhydrous crystalline form of claim 4 having a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least two characteristic peaks expressed in degrees 20 selected from 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21 .8 ± 0.1 .

6. The anhydrous crystalline form of claim 5 having a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having at least three characteristic peaks expressed in degrees 20 selected from 5.4 ± 0.1 , 11 .8 ± 0.1 , 16.3 ± 0.1 , and 21 .8 ± 0.1 .

7. The anhydrous crystalline form of claim 6 having a powder X-ray diffraction pattern (PXRD), measured using copper wavelength radiation, having characteristic peaks expressed in degrees 20 at 5.4 ± 0.1 , 11.8 ± 0.1 , 16.3 ± 0.1 , and 21 .8 ± 0.1.

8. An anhydrous crystalline form that is crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3- diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3- azabicyclo [3.1 ,0]hex-6-yl]acetic acid having a ¹³C solid state NMR spectrum comprising at least one peak in terms of chemical shifts selected from 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm. 53

9. The anhydrous crystalline form of claim 8 having a ¹³C solid state NMR spectrum comprising at least two peaks in terms of chemical shifts selected from 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

10. The anhydrous crystalline form of claim 9 having a ¹³C solid state NMR spectrum comprising at least three peaks in terms of chemical shifts selected from 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

11 . The anhydrous crystalline form of claim 10 having a ¹³C solid state NMR spectrum comprising at least four peaks in terms of chemical shifts selected from 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

12. The anhydrous crystalline form of claim 11 having a ¹³C solid state NMR spectrum comprising peaks in terms of chemical shifts at 55.8 ± 0.2 ppm, 58.3 ± 0.2 ppm, 63.7 ± 0.2 ppm, 64.2 ± 0.2 ppm, and 183.6 ± 0.2 ppm.

13. The anhydrous crystalline form of any one of claims 8-12 having a ¹⁵N ssNMR spectrum comprising at least one peak in terms of chemical shifts selected from -342.1 ± 0.2 ppm, -285.9 ± 0.2 ppm, -278.0 ± 0.2 ppm, and -177.5 ± 0.2 ppm.

14. The anhydrous crystalline form of claim 13 having a ¹⁵N ssNMR spectrum comprising at least two peaks in terms of chemical shifts selected -342.1 ± 0.2 ppm, -285.9 ± 0.2 ppm, -278.0 ± 0.2 ppm, and -177.5 ± 0.2 ppm.

15. The anhydrous crystalline form of claim 13 having a ¹⁵N ssNMR spectrum comprising at least three peaks in terms of chemical shifts selected -342.1 ± 0.2 ppm, -285.9 ± 0.2 ppm, 278.0 ± 0.2 ppm, and -177.5 ± 0.2 ppm.

16. The anhydrous crystalline form of claim 15 having a ¹⁵N ssNMR spectrum comprising peaks in terms of chemical shifts at -342.1 ± 0.2 ppm, -285.9 ± 0.2 ppm, -278.0 ± 0.2 ppm, and - 177.5 ± 0.2 ppm.

17. The crystalline form of any one of claims 3-16 having a Raman spectrum comprising at least one wavenumber (cm ¹) value selected from the group consisting of and 304 ± 2 cm ¹, 315 ± 2 cm ¹ , 935 ± 2 cm ¹ , 1559 ± 2 cm ¹ and 1597 ± 2 cm ¹. 54

18. The anhydrous crystalline form of claim 17 having a Raman spectrum having at least two characteristic wavenumber (cm ¹) values selected from the group consisting of 304 ± 2 cm ¹, 315 ± 2 cm'¹ , 935 ± 2 cm'¹ , 1559 ± 2 cm'¹ and 1597 ± 2 cm'¹.

19. The anhydrous crystalline form of claim 18 having a Raman spectrum having at least three characteristic wavenumber (cm'¹) values selected from the group consisting 304 ± 2 cm ¹, 315 ± 2 cm'¹ , 935 ± 2 cm'¹ , 1559 ± 2 cm^-1 and 1597 ± 2 cm'¹.

20. The anhydrous crystalline form of claim 19 having a Raman spectrum having at least four characteristic wavenumber (cm'¹) values selected from the group consisting 304 ± 2 cm ¹, 315 ± 2 cm'¹ , 935 ± 2 cm'¹ , 1559 ± 2 cm^-1 and 1597 ± 2 cm'¹.

21 . The anhydrous crystalline form of claim 20 having a Raman spectrum comprising wavenumber (cm'¹) values at 304 ± 2 cm'¹, 315 ± 2 cm'¹, 935 ± 2 cm ¹, 1559 ± 2 cm'¹ and 1597 ± 2 cm ¹.

22. Anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3- {2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6- yl]acetic acid, having:

23. The crystalline salt of claim 1 , wherein the crystalline form is a monohydrate.

24. The crystalline salt of claim 23, wherein the crystalline form exhibits an X-ray powder diffraction pattern as shown in Figure 6. 55

25. The crystalline salt of claim 24, wherein the crystalline form exhibits a Raman spectrum pattern as shown in Figure 7.

26. The crystalline form of any one of claims 2-25, wherein the crystalline form has a purity of greater than 97%.

27. The crystalline form of claim 26, wherein the crystalline form has a purity of greater than 99%.

28. A pharmaceutical composition comprising a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methyl azetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6-yl]acetic acid (Form 1) according to any one of claims 2-23, and a pharmaceutically acceptable carrier, diluent, and/or excipient

29. A method of treating or reducing the effects of fructose intolerance in a subject in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of anhydrous crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3-diol salt of [(1 R,5S,6R)- 3-{2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl}-3-azabicyclo[3.1 ,0]hex-6- yl]acetic acid (Form 1) according to any one of claims 2-27.

30. The method of claim 28, wherein the fructose intolerance is hereditary fructose intolerance.

31 . A method of treating a disease selected from treating fatty liver, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, nonalcoholic steatohepatitis with liver fibrosis, nonalcoholic steatohepatitis with cirrhosis, nonalcoholic steatohepatitis with cirrhosis and hepatocellular carcinoma, alcoholic fatty liver disease, alcoholic steatohepatitis, hepatitis B, hepatitis C, and biliary cirrhosis in a subject in need thereof, wherein the method comprises administering to the subject a therapeutically effective amount of crystalline 2-amino-2-(hydroxymethyl)propane-1 ,3- diol salt of [(1 R,5S,6R)-3-{2-[(2S)-2-methylazetidin-1 -yl]-6- (triflu oromethy I) pyrimid in-4-y l}-3- azabicyclo[3.1 ,0]hex-6-yl]acetic acid according to any one of claims 2-27.