WO2016085941A1 - Processes for the preparation of cannabinoid receptor modulators - Google Patents

Processes for the preparation of cannabinoid receptor modulators Download PDF

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WO2016085941A1
WO2016085941A1 PCT/US2015/062345 US2015062345W WO2016085941A1 WO 2016085941 A1 WO2016085941 A1 WO 2016085941A1 US 2015062345 W US2015062345 W US 2015062345W WO 2016085941 A1 WO2016085941 A1 WO 2016085941A1
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cyclopropa
tetrahydro
formula
cyclopenta
pyrazin
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Biman B. PAL
Antonio Garrido Montalban
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Arena Pharmaceuticals Inc
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Arena Pharmaceuticals Inc
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing three or more hetero rings

Definitions

  • the present invention relates to processes, and process intermediates useful in the preparation of (la l S',5a l S r )-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide (Compound 1) and crystalline forms thereof.
  • the compound (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide has been identified as a modulator of the cannabinoid 2 (CB2) receptor and useful in the treatment of CB2 receptor-mediated disorders, for example, osteoarthritis; pain, for example bone and joint pain, muscle pain, dental pain, migraine and other headache pain, inflammatory pain, neuropathic pain, pain that occurs as an adverse effect of therapeutics, and pain associated with a disorder selected from:
  • osteoarthritis cancer, multiple sclerosis, allergic reactions, nephritic syndrome, scleroderma, thyroiditis, diabetic neuropathy, fibromyalgia, HIV related-neuropathy, sciatica, and autoimmune conditions; hyperalgesia; allodynia; inflammatory hyperalgesia; neuropathic hyperalgesia; acute nociception; osteoporosis; multiple sclerosis-associated spasticity; autoimmune disorders, for example an autoimmune disorder selected from the group consisting of: multiple sclerosis, Guillan-Barre syndrome, polyradiculoneuropathy, chronic inflammatory demyelination, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylarthritis, and reactive arthritis; allergic reactions, for example, an allergic reaction associated with a disorder selected from: atopic dermatitis, pruritus, urticaria, asthma, conjunctivitis, allergic rhinitis, and anaphylaxi
  • Cannabinoids are a group of extracellular signaling molecules that are found in both plants and animals. Signals from these molecules are mediated in animals by two G-protein coupled receptors, CB l and CB2.
  • CBl is expressed most abundantly in the neurons of the CNS but is also present at lower concentrations in a variety of peripheral tissues and cells (Matsuda, L. A. et al. (1990) Nature 346:561- 564).
  • CB2 is expressed predominantly, although not exclusively, in non-neural tissues, e.g. in hematopoietic cells, endothelial cells, osteoblasts, osteoclasts, the endocrine pancreas, and cancerous cell lines (Munro, S. et al.
  • CB 1 is believed to be primarily responsible for mediating the psychotropic effects of cannabinoids on the body
  • CB2 is believed to be primarily responsible for most of their non-neural effects.
  • Compound 1 was shown to be a potent and selective CB2 agonist and demonstrated a robust and sustained analgesic efficacy over 6 hours in the streptozotocin-induced Painful Diabetic Peripheral Neuropathy (PDPN) model and also showed therapeutic efficacy for osteoarthritis pain in the monosodium iodoacetate-induced (MIA) model of osteoarthritis; see PCT application
  • PCT/US2010/002360 filed 27 August 2010 (International Publication Number WO2011/025541), and PCT application PCT/US2012/026506, filed 24 February 2012 (International Publication Number WO2012/116276), both are incorporated by reference in their entirety.
  • the present invention provides, inter alia, processes for the preparation (la l S',5a l S r )-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide of Formula (I) (Compound 1) and intermediates thereof.
  • the processes and intermediates of the present invention are useful in preparing (laS,5aS)-2-(4 oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((5)- l- hydroxymethyl-2,2-dimethyl-propyl)-amide, crystalline forms, and pharmaceutical compositions that are useful in the treatment of CB2 receptor-mediated disorders.
  • One aspect of the present invention pertains to processes for preparing
  • One aspect of the present invention pertains to one or more of the processes as described above in Steps a) through e), either provided separately or together, that are useful in the preparation of an intermediate for use directly or indirectly in the preparation of (la l S',5aS')-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl-2,2- dimethyl-propyl) -amide of Formula (I), salts, and/or crystalline forms thereof.
  • One aspect of the present invention pertains to processes for preparing (la l S',5a l S r )-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide of Formula (I) further comprising the step of formulating (la l S',5a l S')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((3 ⁇ 4- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition.
  • the intermediate is (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He) and crystalline forms thereof.
  • the intermediate is sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf) and crystalline forms thereof.
  • Each of the intermediates is useful in the preparation of Compound 1.
  • Figure 1 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the crystalline form of the intermediate (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He).
  • PXRD powder X-ray diffraction
  • Figure 2 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the crystalline form of the intermediate sodium (4aS,5a3 ⁇ 4-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf)-
  • Figure 3 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the crystalline form of 3-((4a l S',5a l S r )-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg).
  • Figure 4 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the anhydrous crystalline form of Compound 1 as described in Example 4, PCT WO2012/ 116276.
  • PXRD powder X-ray diffraction
  • FIG. 5 shows a differential scanning calorimetry (DSC) thermogram and a
  • thermogravimetric analysis (TGA) thermogram for a sample containing the anhydrous crystalline form of Compound 1 as described in Example 4, PCT WO2012/116276.
  • FIG. 6 shows an adsorption and desorption isotherm, Dynamic Moisture Sorption (DMS), for a sample containing the anhydrous crystalline form of Compound 1 as described in Example 4, PCT WO2012/116276.
  • DMS Dynamic Moisture Sorption
  • Figure 7 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the anhydrous crystalline form of Compound 1 obtained from the process described in Example 6 herein.
  • PXRD powder X-ray diffraction
  • Figure 8 shows the effect of Compound 1 in the monosodium iodoacetate (MIA) model of osteoarthritis in rats, see Example 11.
  • agonist refers to a moiety that interacts with and activates a G-protein-coupled receptor, for instance a cannabinoid receptor, and can thereby initiate a physiological or
  • Ci-Ce alkyl refers to a straight or branched carbon radical containing 1 to 6 carbons. Some embodiments are 1 to 5 carbons, some embodiments are 1 to 4 carbons, some embodiments are 1 to 3 carbons and some embodiments are 1 or 2 carbons.
  • alkyl examples include, but are not limited to, methyl, ethyl, w-propyl, isopropyl, w-butyl, sec-butyl, isobutyl, ieri-butyl, pentyl, isopentyl, ieri-pentyl, weo-pentyl, 1-methylbutyl [i.e., -CH(CH 3 )CH 2 CH 2 CH 3 ], 2-methylbutyl [i.e., -CH 2 CH(CH 3 )CH 2 CH 3 ], w-hexyl and the like.
  • in need of treatment and the term “in need thereof” when referring to treatment are used interchangeably to mean a judgment made by a caregiver (e.g. physician, nurse, nurse practitioner, etc. in the case of humans; veterinarian in the case of animals, including non-human mammals) that an individual or animal requires or will benefit from treatment. This judgment is made based on a variety of factors that are in the realm of a caregiver's expertise, but that includes the knowledge that the individual or animal is ill, or will become ill, as the result of a disease, condition or disorder that is treatable by the compounds of the invention. Accordingly, the compounds of the invention can be used in a protective or preventive manner; or compounds of the invention can be used to alleviate, inhibit or ameliorate the disease, condition or disorder.
  • a caregiver e.g. physician, nurse, nurse practitioner, etc. in the case of humans; veterinarian in the case of animals, including non-human mammals
  • mice refers to any animal, including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, swine, cattle, sheep, horses, or primates, and most preferably humans.
  • modulate or modulating refers to an increase or decrease in the amount, quality, response or effect of a particular activity, function or molecule.
  • composition refers to a compound or crystalline form thereof, including but not limited to, salts, solvates, and hydrates of a compound of the present invention, in combination with at least one additional component, such as, a composition obtained/prepared during synthesis, preformulation, in-process testing (i.e., TLC, HPLC, NMR samples), and the like
  • composition refers to a specific composition comprising at least one active ingredient; including but not limited to, salts, solvates, and hydrates of compounds of the present invention, whereby the composition is amenable to investigation for a specified, efficacious outcome in a mammal (for example, without limitation, a human).
  • a mammal for example, without limitation, a human
  • Those of ordinary skill in the art will understand and appreciate the techniques appropriate for determining whether an active ingredient has a desired efficacious outcome based upon the needs of the artisan.
  • phrases "pharmaceutically acceptable salts, solvates, and hydrates" when referring to a compound/compounds as described herein embraces pharmaceutically acceptable solvates and/or hydrates of the compound/compounds, pharmaceutically acceptable salts of the compound/compounds, as well as pharmaceutically acceptable solvates and/or hydrates of pharmaceutically acceptable salts of the compound/compounds. It is also understood that when the phrase “pharmaceutically acceptable solvates and hydrates" or the phrase “pharmaceutically acceptable solvate or hydrate” is used when referring to a compound/compounds as described herein that are salts, it embraces
  • solvates and/or hydrates of such salts. It is also understood by a person of ordinary skill in the art that hydrates are a subgenus of solvates.
  • therapeutically effective amount refers to the amount of active compound or pharmaceutical agent that elicits the biological or medicinal response in a tissue, system, animal, individual or human that is being sought by a researcher, veterinarian, medical doctor or other clinician or caregiver or by an individual, which includes one or more of the following:
  • preventing the disease for example, preventing a disease, condition or disorder in an individual that may be predisposed to the disease, condition or disorder but does not yet experience or display the pathology or symptomatology of the disease;
  • inhibiting the disease for example, inhibiting a disease, condition or disorder in an individual that is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., arresting further development of the pathology and/or symptomatology);
  • Ameliorating the disease for example, ameliorating a disease, condition or disorder in an individual that is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., reversing the pathology and/or symptomatology).
  • GC gas chromatography
  • headspace GC headspace GC
  • HPLC high-pressure liquid chromatography
  • a compound is present in the composition of less than or equal to 1.0% as determined by HPLC
  • the phrase "a compound is present in the composition of less than or equal to 1.0% as determined by HPLC” refers to the use of HPLC and a method as described herein to determine that the compound is present in a quantity of less than or equal to 1.0%.
  • C 1 -C6 alkyl phosphonic anhydride refers to the following formula wherein each C 1 -C6 alkyl group can be the same or different and has the same definition as described herein:
  • each Ci-C6 alkyl group is the same. In some embodiments, each Ci-C6 alkyl group is w-propyl (i.e., 1-propanephosphonic anhydride).
  • thermogravimetric analysis TGA
  • DSC thermogravimetric analysis
  • the temperatures observed for thermal events will depend upon sample purity, and may also depend on the rate of temperature change, as well as sample preparation technique and the particular instrument employed.
  • the values reported herein relating to DSC thermograms can vary by plus or minus about 5°C (i.e., ⁇ about 5°C).
  • the values reported herein relating to DSC thermograms can also vary by plus or minus about 20 joules per gram (i.e., ⁇ about 20 joules per gram).
  • PXRD the relative intensities of the peaks can vary, depending upon the sample preparation technique, the sample mounting procedure and the particular instrument employed.
  • the peak assignments of diffraction patterns can vary by plus or minus about 0.2° (i.e., ⁇ about 0.2°).
  • the temperature features reported herein can vary by plus or minus about 5°C (i.e., ⁇ about 5°C).
  • the TGA % weight changes reported herein over a specified temperature range can vary by plus or minus about 2% weight change (i.e., ⁇ about 2% weight change) due to, for example, variations in sample quality and sample size. Further characterization with respect to hygroscopicity of the crystalline form can be gauged by, for example, dynamic moisture sorption (DMS).
  • DMS dynamic moisture sorption
  • the DMS features reported herein can vary by plus or minus about 5% relative humidity (i.e., ⁇ about 5% relative humidity).
  • the DMS features reported herein can also vary by plus or minus about 2% weight change (i.e., ⁇ about 2% weight change).
  • One aspect of the present invention relates to novel processes for the preparation of an anhydrous form of (la l S',5aS r )-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide
  • One aspect of the present invention relates to an anhydrous crystalline form of (laS,5a5)-2-(4- oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l- hydroxymethyl-2,2-dimethyl-propyl)-amide.
  • One aspect of the present invention relates to an anhydrous crystalline form of (laS,5a5)-2-(4- oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l- hydroxymethyl-2,2-dimethyl-propyl)-amide prepared according to any one of the processes described herein.
  • One aspect of the present invention relates to an anhydrous crystalline form of (laS,5a5)-2-(4- oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide, wherein the anhydrous crystalline form has a powder X- ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°.
  • the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, and 10.7° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, and 16.9° + 0.2°.
  • the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, and 1 1.1° + 0.2°.
  • the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2(9, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, and 17.4° + 0.2°.
  • the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, and 16.5° + 0.2°.
  • the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, 16.5° + 0.2°, 14.5° + 0.2°, 11.8° + 0.2°, and 18.9° + 0.2°.
  • the anhydrous crystalline form has a powder X-ray diffraction pattern substantially as shown in Figure 4, wherein by "substantially” is meant that the reported peaks can vary by about + 0.2 °2 ⁇ .
  • the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 159.6°C and about 169.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 160.6°C and about 168.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 162.6°C and about 166.6°C.
  • the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 163.6°C and about 165.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature at about 164.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram substantially as shown in Figure 5, wherein by “substantially” is meant that the reported DSC features can vary by about + 5°C and that the reported DSC features can vary by about + 20 joules per gram.
  • the anhydrous crystalline form has a thermogravimetric analysis profile showing about 0.5% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile showing about 0.25% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile substantially as shown in Figure 5, wherein by “substantially” is meant that the reported TGA features can vary by about + 5°C, and that that the reported TGA features can vary by about + 2% weight change.
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, and 10.7° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 159.6°C and about 169.6°C; and 3) a thermogravimetric analysis profile showing about 0.5% weight loss below about 135°C.
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, and 16.9° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 160.6°C and about 168.6°C; and 3) a thermogravimetric analysis profile showing about 0.25% weight loss below about 135°C.
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, and 11.1° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 162.6°C and about 166.6°C; and 3) a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, and 17.4° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 163.6°C and about 165.6°C; and 3) a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° +
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, 16.5° + 0.2°, 14.5° + 0.2°, 11.8°
  • thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
  • the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern substantially as shown in Figure 4; 2) a differential scanning calorimetry thermogram substantially as shown in Figure 5; and 3) a thermogravimetric analysis profile substantially as shown in Figure 5.
  • One aspect of the present invention relates to a novel crystalline form of (4a l S',5aS')-methyl 1-
  • Forma (He) One aspect of the present invention relates to a novel crystalline form of (4aS,5aS)-mefhyl 1- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of (Formula (He)) prepared according to any one of the processes described herein.
  • One aspect of the present invention relates to a crystalline form of (4aS,5aS)-mefhyl l-(pyrazin- 2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He)), wherein the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2° and 11.0° + 0.2°.
  • the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, and 17.9° + 0.2°. In some embodiments, the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, and 17.9° + 0.2°.
  • the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.9° + 0.2°, and 25.3° + 0.2°.
  • the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.3° + 0.2°, 17.9° + 0.2°, 21.0° + 0.2°, 24.3° + 0.2°, 25.3° + 0.2°, and 25.9° + 0.2°.
  • the crystalline form of Formula (He) has a powder X-ray diffraction pattern substantially as shown in Figure 1, wherein by "substantially” is meant that the reported peaks can vary by about + 0.2 °2 ⁇ .
  • One aspect of the present invention relates to a novel crystalline form of sodium (4aS,5a5)-l- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (Ilf)).
  • One aspect of the present invention relates to a novel crystalline form of sodium
  • One aspect of the present invention relates to a crystalline form of sodium
  • the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 1 ⁇ , at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 2(9, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°.
  • the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°.
  • the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 2(9, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 11.5° + 0.2°, 12.3° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°.
  • the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern substantially as shown in Figure 2, wherein by “substantially” is meant that the reported peaks can vary by about + 0.2 °2 ⁇ .
  • One aspect of the present invention relates to a novel crystalline form of
  • One aspect of the present invention relates to a novel crystalline form of
  • One aspect of the present invention relates to a crystalline form of 3-((4a l S',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Formula (Ilg)), wherein the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 1 ⁇ , at 12.6° + 0.2° and 16.8° + 0.2°.
  • the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 1 ⁇ , at 12.6° + 0.2°, 16.8° + 0.2°, and 27.1° + 0.2°. In some embodiments, the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 12.6° + 0.2°, 16.8° + 0.2°, 22.0° + 0.2°, 26.5° + 0.2°, and 27.1° + 0.2°.
  • the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 12.6° + 0.2°, 16.8° + 0.2°, 21.6° + 0.2°, 22.0° + 0.2°, 22.6° + 0.2°, 26.5° + 0.2°, and 27.1° + 0.2°.
  • Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 1 ⁇ , at 12.6° + 0.2°, 12.9° + 0.2°, 16.8° + 0.2°, 21.0° + 0.2°, 21.6° + 0.2°, 22.0° + 0.2°, 22.2° + 0.2°, 22.6° + 0.2°, 26.5° + 0.2°, 27.1° + 0.2°, and 27.6° + 0.2°.
  • the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern substantially as shown in Figure 3, wherein by “substantially” is meant that the reported peaks can vary by about + 0.2 °2 ⁇ .
  • the crystalline forms described herein can be prepared by any of the suitable procedures known in the art for preparing crystalline polymorphs. In some embodiments the crystalline forms described herein are prepared according to the Examples. In some embodiments, the crystalline forms described herein can be prepared by heating crystalline forms other than the crystalline forms described herein. In some embodiments, the crystalline forms described herein can be prepared by recrystahizing crystalline forms other than the crystalline forms described herein.
  • the present invention is directed, inter alia, to processes useful in the preparation of (laS ⁇ aS")- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimethyl-propyl)-amide and crystalline forms thereof.
  • spectroscopic means such as nuclear magnetic resonance spectroscopy ⁇ e.g., 3 ⁇ 4 or 13 C), infrared spectroscopy, spectrophotometry ⁇ e.g., UV- visible), mass spectrometry, gas chromatography (GC), or by chromatography, such as, high performance liquid chromatography (HPLC) or thin layer chromatography.
  • HPLC high performance liquid chromatography
  • the usual isolation and purification operations such as concentration, filtration, extraction, solid-phase extraction, recrystallization, enantiomeric-enrichment via recrystallization, chromatography, and the like may be used, to isolate the desired product.
  • ketone forming step utilizing the intermediate of Formula (lib) in the presence of an oxidizing agent (e.g., TEMPO/NaOCl), ketone-forming solvent (e.g., MTBE), and a ketone- forming aqueous buffer (e.g., potassium phosphate buffer) is provided below in Scheme II.
  • an oxidizing agent e.g., TEMPO/NaOCl
  • ketone-forming solvent e.g., MTBE
  • a ketone-forming aqueous buffer e.g., potassium phosphate buffer
  • the sodium salt intermediates i.e., sodium (£ ' )-2-methoxy-2-oxo-l- ((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-ylidene)efhanolate of Formula (IIc-i) and sodium (Z)-2-methoxy- 2-oxo-l-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of Formula (IIc-ii)) depicted inside the brackets above can be used directly in the next step (i.e., condensing-step), or can be isolated, or the sodium salt intermediates could be subsequently neutralized to provide the diketo-ester intermediate (i.e., methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii)) as represented below:
  • the representative reacting step and condensing step can be successively employed in a two-step process using the intermediate of Formula (lie) and dimethyl oxalate in the presence of sodium methoxide and methanol without isolation of the intermediates of Formulae (IIc-i), (Ilc-ii), and/or (Ilc-iii) followed by using the intermediate of Formula (lid) in the presence of a condensing-step solvent as provided below in Scheme V.
  • One aspect of the present invention includes every combination of one or more process step and intermediates related thereto used in the preparation of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a- tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimefhyl- propyl)-amide of Formula (I) and crystalline forms thereof, such as those processes exemplified by Schemes I, II, III, IV, V, VI, VII, and VIII (supra) and compounds of Formulae (I), (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Ilf), (Hf-i), (Ilg), (Ilh), (Ilia), (Illb), and (IIIc).
  • One aspect of the present invention pertains to one or more of the intermediates, such as Compounds of Formulae (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Ilf), (Ilf-i), (Ilg), and (Ilh), as exemplified in Schemes I, II, III, IV, V, VI, VII, and VIII (supra), useful in the preparation of the Compound of Formula (I) and crystalline forms thereof.
  • the intermediates such as Compounds of Formulae (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Ilf), (Ilf-i), (Ilg), and (Ilh), as exemplified in Schemes I, II, III, IV, V, VI, VII, and VIII (supra), useful in the preparation of the Compound of Formula (
  • Example 1, Step C, WO2012/116276 Replacing absolute ethanol in the previously reported method with methanol improved the volume efficiency but provided an undesirable mixture of esters. Using dimethyl oxalate in methanol and replacing THF/potassium ieri-butoxide with sodium methoxide provided unexpected improvements over the previously reported methods, such as those cited above. Further, the Compound of Formulae (He) and the crystalline form of the Compound of Formulae (He) were not previously disclosed. Another improvement is the isolation of the sodium salt of Formula (Ilf) instead of the free acid of Formula (Ilf-i) as previously disclosed (see, Example 1.107, Step D, WO2011/025541 and Example 1, Step D, WO2012/116276).
  • the crystal form of the sodium salt of Formula (Ilf) showed improved manufacturing properties compared to the free acid, such as, processing (e.g., crystal size), processability (e.g. improved filterability on scale), and ease of drying while retaining purity and yield.
  • processing e.g., crystal size
  • processability e.g. improved filterability on scale
  • ease of drying while retaining purity and yield.
  • WO2011/025541 and WO2012/116276 the crystalline form of the sodium salt of Formula (Ilf) was not isolated and was not previously disclosed.
  • Another improvement relates to the oxidation processes for preparing the Compound of Formula (Ilg). It was previously reported that two separate oxidation procedures were required when using formic acid and hydrogen peroxide to provide the Compound of Formula (Ilg) (see, Example 1.108, WO2011/025541 and Example 1, Method 2, Step E, WO2012/116276). However, it was discovered that when the oxidation was conducted in the presence of trifluoroacetic acid (i.e., TFA) the second separate oxidation step was not necessary and the present oxidation proceeds to completion (i.e., less than 0.6% of non-oxidized free acid, see Example 5).
  • TFA trifluoroacetic acid
  • Another improvement relates to the coupling processes for preparing the Compound of Formula (I) from Compounds of Formula (Ilg) and (Ilh) in the presence of acetonitrile as the coupling- step solvent. It was previously reported that coupling Compounds of Formula (Ilg) and (Ilh) in DMF followed by recrystallization with CH 2 Cl 2 /hexanes and a lengthy drying step (i.e., ⁇ 65°C for 10 days) provided the Compound of Formula (I) in a 67.2% yield (see, Example 1, Method 2, Step F,
  • Tautomeric forms can be in equilibrium or sterically locked into one form by appropriate substitution. It is understood that the various tautomeric forms are within the scope of the compounds of the present invention.
  • Compounds of the invention also include all isotopes of atoms occurring in the intermediates and/or final compounds.
  • Isotopes include those atoms having the same atomic number but different mass numbers.
  • isotopes of hydrogen include deuterium and tritium.
  • One aspect of the present invention relates to processes comprising the following steps, (i.e. Steps a) to Steps e)), for preparing (laS,5a3 ⁇ 4-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
  • reacting in Step a) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, reacting in Step a) is carried out in the presence of a nitrogen atmosphere.
  • reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first- reacting mixture.
  • the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the first-reacting mixture is at a temperature of about 20°C to about 25°C.
  • reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture.
  • sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
  • the second-reacting mixture is maintained at a temperature of about 25°C to about 50°C. In some embodiments, wherein after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 30°C to about 45°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 35°C to about 40°C.
  • sodium methoxide is in a solution comprising methanol. In some embodiments, sodium methoxide is about 5 weight percent to about 30 weight percent in methanol. In some embodiments, sodium methoxide is about 25 weight percent in methanol solution.
  • dimethyl oxalate and sodium methoxide are present in molar excess compared to (lS,5R)-bicyclo[3.1.0]hexan-2-one.
  • sodium methoxide is present in molar excess compared to dimethyl oxalate.
  • the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1 : 1.05: 1.05 to about 1 :2.4:2.8. In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1 : 1.2: 1.4.
  • the intermediate is sodium (£)-2-methoxy-2-oxo-l-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of the formula:
  • the intermediate is not isolated.
  • the intermediate is isolated.
  • the intermediate is isolated and is methyl 2-oxo-2-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (Ilc-i):
  • condensing in Step b) further comprises the step of mixing the intermediate and the condensing-step solvent together to form a first-condensing mixture.
  • the condensing-step solvent comprises methanol.
  • the condensing is carried out in the presence of the condensing-step acid.
  • the condensing-step acid is HCl. In some embodiments, the condensing- step acid is aqueous HCl. In some embodiments, the condensing-step acid is about IN aqueous HCl to about 12N aqueous HCl. In some embodiments, the condensing-step acid is about 3N aqueous HCl to about ION aqueous HCl. In some embodiments, the condensing-step acid is about 5N aqueous HCl to about 7N aqueous HCl. In some embodiments, the condensing-step acid is about 6N aqueous HCl.
  • condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture.
  • the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing- step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about
  • the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
  • condensing in Step b) further comprises the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-condensing mixture.
  • the isolating is by filtration.
  • ⁇ aS ⁇ aS ⁇ -methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 90% or greater as determined by HPLC. In some embodiments, (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
  • hydrolyzing in Step c) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, hydrolyzing in Step c) is carried out in the presence of a nitrogen atmosphere.
  • hydrolyzing in Step c) further comprising the step of mixing (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the hydrolyzing- step solvent together to form a first-hydrolyzing mixture.
  • the hydrolyzing-step solvent comprises methanol.
  • the first-hydrolyzing mixture is at a temperature of about 20°C to about
  • hydrolyzing in Step c) further comprises the step of adding the sodium hydroxide to the first-hydrolyzing mixture to form a second-hydrolyzing mixture.
  • the temperature of the second-hydrolyzing mixture is maintained at about 25°C to about 60°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 35°C to about 55°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 40°C to about 50°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 45°C.
  • hydrolyzing in Step c) further comprises the step of isolating sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-hydrolyzing mixture.
  • isolating is by filtration.
  • sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • crystalline sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
  • oxidizing in Step d) further comprises the step of mixing sodium
  • the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2.
  • the molar ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
  • the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C.
  • oxidizing in Step d) further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
  • hydrogen peroxide is added to the first-oxidizing mixture in multiple portions. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
  • oxidizing in Step d) further comprises the step of isolating 3-((4aS,5aS)- 3-carboxy-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5] cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide from the second-oxidizing mixture.
  • the isolating is by filtration.
  • 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
  • the coupling in Step e) further comprises the step of mixing 3- ((4a l S',5a l S')-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, the coupling-step base, and the coupling-step solvent together to form a first-coupling mixture.
  • the first-coupling mixture is at a temperature of about -15°C to about
  • the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
  • the coupling in Step e) further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
  • the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about
  • the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C.
  • the coupling in Step e) further comprises the step of adding (S)-tert- leucinol to the second-coupling mixture.
  • (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
  • (S)-ieri-leucinol is added substantially all at once. It was found that adding (S)-ieri-leucinol all at once instead of over a longer period of time minimized the amount of a side product.
  • the coupling-step agent is a tetramethyluronium-based coupling agent.
  • the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro-l/i-benzotriazole-l-yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and A ⁇ A ⁇ A ⁇ -tetramefhyl-CKS ⁇ -dihydro- 4-oxo-l, 2, 3-benzoxo-l
  • HATU hexafluorophosphate
  • the coupling-step agent is a phosphonic acid anhydride.
  • the phosphonic acid anhydride is a Ci-C 6 alkyl phosphonic anhydride.
  • the phosphonic acid anhydride is 1-propanephosphonic anhydride.
  • the coupling-step base comprises a tertiary amine. In some embodiments, the coupling-step base comprises a tertiary amine.
  • the coupling-step base is selected from: A ⁇ N-diisopropylefhylamine (DIEA), triethylamine (TEA), N-mefhylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimethylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4- (dimethylamino)pyridine (DBDMAP).
  • DIEA A ⁇ N-diisopropylefhylamine
  • TAA triethylamine
  • NMM N-mefhylmorpholine
  • DMAP 4-dimethylaminopyridine
  • 2,4,6- trimethylpyridine collidine
  • 2,3,5,6-tetramethylpyridine 2,3,5,6-tetramethylpyridine
  • BDMAP 2,6-di-ieri-butyl-4- (dimethylamino)pyridine
  • the coupling-step solvent is acetonitrile.
  • the coupling in Step e) further comprises the step of isolating (la ⁇ SaS")- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture.
  • the isolating is by filtration.
  • (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
  • the coupling in Step e) further comprises the step of formulating (la l S',5a l S r )-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition.
  • the formulating comprises admixing (la l S',5a l S')-2-(4-oxy-pyrazin-2- yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl- 2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
  • the formulating is admixing (la l S',5a l S')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene- 4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
  • One aspect of the present invention relates to processes comprising the following steps, i.e. Steps a) to Steps c), for preparing (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
  • hydrolyzing in Step a) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, hydrolyzing in Step a) is carried out in the presence of a nitrogen atmosphere.
  • hydrolyzing in Step a) further comprises the step of mixing (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the hydrolyzing- step solvent together to form a first-hydrolyzing mixture.
  • the hydrolyzing-step solvent comprises methanol.
  • the first-hydrolyzing mixture is at a temperature of about 20°C to about
  • hydrolyzing in Step a) further comprises the step of adding the sodium hydroxide to the first-hydrolyzing mixture to form a second-hydrolyzing mixture.
  • the temperature of the second-hydrolyzing mixture is maintained at about 25°C to about 60°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 35°C to about 55°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 40°C to about 50°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 45°C.
  • hydrolyzing in Step a) further comprises the step of isolating sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-hydrolyzing mixture.
  • isolating is by filtration.
  • sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • crystalline sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
  • oxidizing in Step b) further comprises the step of mixing sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the oxidizing-step solvent together to form a first-oxidizing mixture.
  • the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to 1.00:0.71 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2.
  • the molar ratio of sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
  • the ratio of sodium (4aS,5a3 ⁇ 4-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
  • the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C.
  • oxidizing in Step b) further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
  • hydrogen peroxide is added to the first-oxidizing mixture in multiple portions.
  • hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
  • oxidizing in Step b) further comprises the step of isolating 3-((4aS,5aS)- 3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide from the second-oxidizing mixture.
  • the isolating is by filtration.
  • 3-((4a l S',5a l S r )-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
  • the coupling in Step c) further comprises the step of mixing 3- ((4a l S',5a l S')-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, the coupling-step base, and the coupling-step solvent together to form a first-coupling mixture.
  • the first-coupling mixture is at a temperature of about -15°C to about 25°C. In some embodiments, the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
  • the coupling in Step c) further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
  • the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about -5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C. In some embodiments, the coupling in Step c) further comprises the step of adding (S)-tert- leucinol to the second-coupling mixture.
  • (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
  • (S)-ieri-leucinol is added substantially all at once.
  • the coupling-step agent is a tetramethyluronium-based coupling agent.
  • the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro-l/i-benzotriazole-l-yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and A ⁇ A ⁇ A ⁇ -tetra
  • HATU hexafluorophosphate
  • the coupling-step agent is a phosphonic acid anhydride.
  • the phosphonic acid anhydride is a Ci-C6 alkyl phosphonic anhydride.
  • the phosphonic acid anhydride is 1-propanephosphonic anhydride.
  • the coupling-step base comprises a tertiary amine. In some embodiments, the coupling-step base comprises a tertiary amine.
  • the coupling-step base is selected from: A ⁇ N-diisopropylefhylamine (DIEA), triefhylamine (TEA), N-mefhylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimefhylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4- (dimethylamino)pyridine (DBDMAP).
  • DIEA A ⁇ N-diisopropylefhylamine
  • TAA triefhylamine
  • NMM N-mefhylmorpholine
  • DMAP 4-dimethylaminopyridine
  • 2,4,6- trimefhylpyridine collidine
  • 2,3,5,6-tetramethylpyridine 2,3,5,6-tetramethylpyridine
  • BDMAP 2,6-di-ieri-butyl-4- (dimethylamin
  • the coupling-step solvent is acetonitrile.
  • the coupling in Step c) further comprises the step of isolating (laS,5aS)- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture.
  • the isolating is by filtration.
  • (la l S',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
  • the coupling in Step c) further comprises the step of formulating (la l S',5a l S r )-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition.
  • the formulating comprises admixing (laS,5aS)-2-(4-oxy-pyrazin-2- yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl- 2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
  • the formulating is admixing (la l S',5a l S')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene- 4-carboxylic acid ((S)-l -hydro xymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
  • One aspect of the present invention relates to processes comprising the following Steps a) and Steps b) for preparing (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
  • oxidizing in Step a) further comprises the step of mixing sodium
  • the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2.
  • the molar ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL.
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
  • the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C.
  • oxidizing in Step a) further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
  • hydrogen peroxide is added to the first-oxidizing mixture in multiple portions. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
  • oxidizing in Step a) further comprises the step of isolating 3-((4aS,5aS)- 3-carboxy-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5] cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide from the second-oxidizing mixture.
  • the isolating is by filtration.
  • 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
  • the coupling in Step b) further comprises the step of mixing 3- ((4a l S',5a l S')-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, the coupling-step base, and the coupling-step solvent together to form a first-coupling mixture.
  • the first-coupling mixture is at a temperature of about -15°C to about 25°C.
  • the first-coupling mixture is at a temperature of about -10°C to about 15°C.
  • the first-coupling mixture is at a temperature of about -5°C to about 10°C.
  • the first-coupling mixture is at a temperature of about 0°C to about 5°C.
  • the coupling in Step b) further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
  • the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about -5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C.
  • the coupling in Step b) further comprises the step of adding (S)-tert- leucinol to the second-coupling mixture.
  • (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
  • (S)-ieri-leucinol is added substantially all at once.
  • the coupling-step agent is a tetramethyluronium-based coupling agent.
  • the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- 1-yl)- 1 , 1 ,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro- lH-benzotriazole- 1 -yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and N,N,N,N etramethyl-0-(3,4-dihydro- 4-oxo-l, 2,3-benzotri
  • HATU hexafluorophosphate
  • the coupling-step agent is a phosphonic acid anhydride.
  • the phosphonic acid anhydride is a Ci-C6 alkyl phosphonic anhydride.
  • the phosphonic acid anhydride is 1-propanephosphonic anhydride.
  • the coupling-step base comprises a tertiary amine. In some embodiments, the coupling-step base comprises a tertiary amine.
  • the coupling-step base is selected from: A ⁇ N-diisopropylefhylamine (DIEA), triethylamine (TEA), N-methylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimethylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4-
  • the coupling-step base is triethylamine.
  • the coupling-step solvent is acetonitrile.
  • the coupling in Step b) further comprises the step of isolating (la ⁇ a ⁇ - 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture.
  • the isolating is by filtration.
  • (la l S',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
  • the coupling in Step b) further comprises the step of formulating (la l S',5a l S r )-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition.
  • the formulating comprises admixing (la l S',5a l S')-2-(4-oxy-pyrazin-2- yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl- 2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
  • the formulating is admixing (la l S',5a l S')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene- 4-carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
  • One aspect of the present invention relates to processes for preparing (la l S',5a l S')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)- (I):
  • coupling further comprises the step of mixing 3-((4a l S',5a l S r )-3-carboxy-
  • the first-coupling mixture is at a temperature of about -15°C to about 25°C. In some embodiments, the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
  • coupling further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
  • the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about -5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C.
  • coupling further comprises the step of adding (S)-ieri-leucinol to the second-coupling mixture.
  • (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
  • (S)-ieri-leucinol is added substantially all at once.
  • the coupling-step agent is a tetramethyluronium-based coupling agent.
  • the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro-l/i-benzotriazole-l-yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and A ⁇ A ⁇ A ⁇ -tetra
  • the coupling-step agent is a phosphonic acid anhydride.
  • the phosphonic acid anhydride is a C1-C6 alkyl phosphonic anhydride.
  • the phosphonic acid anhydride is 1-propanephosphonic anhydride.
  • the coupling-step base comprises a tertiary amine. In some embodiments, the coupling-step base comprises a tertiary amine.
  • the coupling-step base is selected from: A ⁇ N-diisopropylefhylamine (DIEA), triethylamine (TEA), N-mefhylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimethylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4- (dimethylamino)pyridine (DBDMAP).
  • DIEA A ⁇ N-diisopropylefhylamine
  • TAA triethylamine
  • NMM N-mefhylmorpholine
  • DMAP 4-dimethylaminopyridine
  • 2,4,6- trimethylpyridine collidine
  • 2,3,5,6-tetramethylpyridine 2,3,5,6-tetramethylpyridine
  • BDMAP 2,6-di-ieri-butyl-4- (dimethylamino)pyridine
  • the coupling-step solvent is acetonitrile.
  • coupling further comprises the step of isolating (la l S',5a l S')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture.
  • the isolating is by filtration.
  • (la l S',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
  • coupling further comprises the step of formulating (la l S',5a l S')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide to form a pharmaceutical composition.
  • the formulating comprises admixing (laS',5a l S')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- 1 /i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydro xymethyl-2,2-dimethyl- propyl)-amide with a pharmaceutical excipient.
  • the formulating is admixing (la l S',5a l S r )-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((3 ⁇ 4- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide with a pharmaceutical excipient.
  • (4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He) is prepared from methyl 2- oxo-2-((lS,53 ⁇ 4-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) that was previously prepared according to any one of the processes described herein or by an alternative process.
  • one aspect of the present invention relates to the use of methyl 2-oxo-2-((lS,5S)- 2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) in processes for preparing (4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
  • the condensing step further comprises the step of mixing the intermediate of Formula (Ilc-iii) and the condensing-step solvent together to form a first-condensing mixture.
  • the condensing-step solvent comprises methanol.
  • the condensing is carried out in the presence of the condensing-step acid.
  • the condensing-step acid is HCl.
  • the condensing- step acid is aqueous HCl.
  • the condensing-step acid is about IN aqueous HCl to about 12N aqueous HCl.
  • the condensing-step acid is about 3N aqueous HCl to about ION aqueous HCl.
  • the condensing-step acid is about 5N aqueous HCl to about 7N aqueous HCl.
  • the condensing-step acid is about 6N aqueous HCl.
  • the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture.
  • the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing- step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 20°C to about 50°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
  • the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 20°C to about 50°C.
  • the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C.
  • the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
  • the condensing step further comprises the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-condensing mixture.
  • the isolating is by filtration.
  • (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline and has a powder X-ray diffraction pattern comprising one or more peaks as described herein, in terms of 1 ⁇ .
  • (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 90% or greater as determined by HPLC. In some embodiments, (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
  • the Compound of Formula (He) can be prepared using either: 1) the Compound of Formula
  • some embodiments relate to the preparation of (4aS,5aS)-methyl l-(pyrazin-2-yl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
  • reacting in Step a) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, reacting in Step a) is carried out in the presence of a nitrogen atmosphere.
  • reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture.
  • the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the first-reacting mixture is at a temperature of about 20°C to about 25°C.
  • reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 0°C to about 35°C.
  • reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 10°C to about 35°C.
  • reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 15°C to about 30°C.
  • reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 20°C to about 25°C.
  • reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture.
  • sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
  • reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C.
  • reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C.
  • reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C.
  • reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
  • the second-reacting mixture is maintained at a temperature of about 25°C to about 50°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 30°C to about 45°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 35°C to about 40°C.
  • the sodium methoxide is in a solution comprising methanol. In some embodiments, the sodium methoxide is about 5 weight percent to about 30 weight percent in methanol. In some embodiments, the sodium methoxide is about 25 weight percent in methanol solution. In some embodiments, the dimethyl oxalate and the sodium methoxide are present in molar excess compared to (lS,5R)-bicyclo[3.1.0]hexan-2-one.
  • the sodium methoxide is present in molar excess compared to dimethyl oxalate.
  • the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.05: 1.05 to about 1 :2.4:2.8. In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.2: 1.4.
  • the intermediate is sodium (£)-2-methoxy-2-oxo-l-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of the formula:
  • the intermediate is not isolated.
  • reacting in Step a) further comprises neutralizing the intermediate. In some embodiments, reacting in Step a) further comprises neutralizing the intermediate with HC1.
  • reacting in Step a) further comprises neutralizing the intermediate comprising sodium (£)-2-methoxy-2-oxo- 1 -(( 1 S,5S)-2-oxobicyclo [3.1.0]hexan-3-ylidene)ethanolate and/or sodium (Z)-2-methoxy-2-oxo-l-((l l S',5 l S')-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate.
  • reacting in Step a) further comprises neutralizing the intermediate comprising sodium (£)-2-methoxy-2-oxo-l-((l l S',5 l S r )-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate and/or sodium (Z)-2-methoxy-2-oxo-l-((l l S',5 l S r )-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate with HC1.
  • the intermediate is isolated.
  • the intermediate is isolated and is methyl 2-oxo-2-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-yl)acetate of the formula:
  • condensing in Step b) further comprises the step of mixing the intermediate and the condensing-step solvent together to form a first-condensing mixture.
  • the condensing-step solvent comprises methanol.
  • the condensing is carried out in the presence of the condensing-step acid.
  • the condensing-step acid is HC1.
  • the condensing- step acid is aqueous HC1.
  • the condensing-step acid is about IN aqueous HC1 to about 12N aqueous HC1.
  • the condensing-step acid is about 3N aqueous HC1 to about ION aqueous HC1.
  • the condensing-step acid is about 5N aqueous HC1 to about 7N aqueous HC1.
  • the condensing-step acid is about 6N aqueous HC1.
  • condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture.
  • the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing- step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about
  • the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
  • condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 55°C or below.
  • condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second- condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 20°C to about 50°C.
  • condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C.
  • condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
  • condensing in Step b) further comprises the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-condensing mixture.
  • the isolating is by filtration.
  • (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline and has a powder X-ray diffraction pattern comprising one or more peaks as described herein, in terms of 1 ⁇ .
  • (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 90% or greater as determined by HPLC. In some embodiments, (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
  • the condensing-step solvent is substantially methanol, the same solvent used in the processes used to prepare the Compound of Formula (IIc-iii).
  • an additional solvent could be added thereby providing a condensing-step solvent comprising methanol (i.e., methanol in the presence of at least one other added solvent).
  • methanol can be substantially removed completely and an alternative condensing-step solvent used, commonly referred in the art as a "solvent exchange" step.
  • One aspect of the present invention relates to processes for preparing sodium
  • hydrolyzing is carried out in the presence of an inert anhydrous atmosphere.
  • hydrolyzing is carried out in the presence of a nitrogen atmosphere.
  • hydrolyzing further comprising the step of mixing (4aS,5aS)-methyl 1- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate and the hydrolyzing- step solvent together to form a first-hydrolyzing mixture.
  • the hydrolyzing-step solvent comprises methanol.
  • the first-hydrolyzing mixture is at a temperature of about 20°C to about
  • hydrolyzing further comprises the step of adding the sodium hydroxide to the first-hydrolyzing mixture to form a second-hydrolyzing mixture.
  • the temperature of the second-hydrolyzing mixture is maintained at about 25°C to about 60°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 35°C to about 55°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 40°C to about 50°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 45°C.
  • hydrolyzing further comprises the step of isolating sodium
  • isolating is by filtration.
  • sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • crystalline sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
  • One aspect of the present invention relates to processes for preparing 3-((4a l S',5a l S')-3-carboxy-
  • oxidizing further comprises the step of mixing sodium
  • the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to 1.00:0.71 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume.
  • the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
  • the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2.
  • the molar ratio of sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5.
  • the molar ratio of sodium (4aS,5a3 ⁇ 4-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
  • the ratio of sodium (4aS,5a3 ⁇ 4-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S r )-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL.
  • the ratio of sodium (4aS',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
  • the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL.
  • the ratio of sodium (4a l S',5a l S')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
  • the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C. In some embodiments, oxidizing further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
  • hydrogen peroxide is added to the first-oxidizing mixture in multiple portions. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
  • oxidizing further comprises the step of isolating 3-((4aS,5aS)-3- carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide from the second-oxidizing mixture.
  • the isolating is by filtration.
  • 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
  • the intermediate methyl 2-oxo-2-((lS,53 ⁇ 4-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) can be either isolated or used directly in the condensing step as described herein.
  • the intermediate methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3- yl)acetate of Formula (IIc-iii) is isolated. It is understood that one or more tautomers are possible for the Compound of Formula (IIc-iii), for example:
  • one aspect of the present invention relates to processes for preparing methyl 2- oxo-2-((lS,53 ⁇ 4-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii):
  • reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (He) with dimethyl oxalate is carried out in the presence of an inert anhydrous atmosphere.
  • the reacting step is carried out in the presence of a nitrogen atmosphere.
  • the reacting step further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture.
  • the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the first-reacting mixture is at a temperature of about 20°C to about 25°C.
  • the reacting step further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture wherein the first-reacting mixture is at a temperature of about 0°C to about 35°C.
  • the reacting step further comprises the step of mixing (lS,5R)-bicyclo[3.1.0]hexan- 2-one of Formula (He), dimethyl oxalate, and methanol together to form a first-reacting mixture wherein the first-reacting mixture is at a temperature of about 10°C to about 35°C.
  • the reacting step further comprises the step of mixing (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first-reacting mixture wherein the first-reacting mixture is at a temperature of about 15°C to about 30°C.
  • the reacting step further comprises the step of mixing (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first-reacting mixture wherein the first-reacting mixture is at a temperature of about 20°C to about 25°C.
  • the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture.
  • sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C.
  • sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C.
  • sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C.
  • sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
  • the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C.
  • the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
  • the second-reacting mixture is maintained at a temperature of about 25°C to about 50°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 30°C to about 45°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 35°C to about 40°C.
  • the sodium methoxide is in a solution comprising methanol. In some embodiments, the sodium methoxide is about 5 weight percent to about 30 weight percent in methanol. In some embodiments, the sodium methoxide is about 25 weight percent in methanol solution.
  • the dimethyl oxalate and the sodium methoxide are present in molar excess compared to (lS,5R)-bicyclo[3.1.0]hexan-2-one.
  • the sodium methoxide is present in molar excess compared to dimethyl oxalate.
  • the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.05: 1.05 to about 1 :2.4:2.8. In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.2: 1.4.
  • the reacting step further comprises the step of neutralizing with a reacting-step acid to form methyl 2-oxo-2-((lS,53 ⁇ 4-2-oxobicyclo[3.1.0]hexan-3-yl)acetate.
  • the reacting-step acid is HCl.
  • the reacting-step acid is aqueous HCl.
  • the reacting-step acid is 6N aqueous HCl.
  • the reacting step further comprises the step of isolating methyl 2-oxo-2- ((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) from the second-condensing mixture.
  • One aspect of the present invention includes every combination of one or more compounds selected from the compound of Formulae (I), (Ila), (lib), (He), (IIc-i), (IIc-ii), (IIc-iii), (lid), (He), (Hf) > (Ilf-i), (Ilg), and (Hh), and salts, solvates, and hydrates thereof.
  • One aspect of the present invention relates to a compound selected from the compound of Formula (He), and salts, solvates, and hydrates thereof:
  • the compound is (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate.
  • (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
  • the crystalline form of (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2° and 11.0° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, and 17.9° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, and 17.9° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.9° + 0.2°, and 25.3° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.3° + 0.2°, 17.9° + 0.2°, 21.0° + 0.2°, 24.3° + 0.2°, 25.3° + 0.2°, and 25.9° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern substantially as shown in Figure 1.
  • One aspect of the present invention relates to isolated sodium (4aS,5aS)-l-(pyrazin-2-yl)-
  • One aspect of the present invention relates to a crystalline form of sodium (4aS,5aS)-l- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-lH-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf).
  • the crystalline form of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-lH-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.3° + 0.2° and 25.3° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2 ⁇ , at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 11.5° + 0.2°, 12.3° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°.
  • the crystalline form has a powder X-ray diffraction pattern substantially as shown in Figure 2.
  • One aspect of the present invention relates to the Compound of Formula (Ilia), 3-((4aS,5aS)-3- ((( l S , )-3,3-dimethyl-l-(((4a,S , ,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide.
  • Compound 1 and pharmaceutical compositions comprising Compound 1 are useful in the treatment of several other diseases and disorders, and/or in the amelioration of symptoms thereof.
  • one aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of the human or animal body by therapy.
  • Another aspect of the present invention relates to methods for the treatment of a CB2 receptor- mediated disorder in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
  • the CB2 receptor-mediated disorder is any one or more of the disorders as described herein.
  • Another aspect of the present invention relates to the use of a pharmaceutical composition as described herein, in the treatment of a CB2 receptor-mediated disorder.
  • the CB2 receptor-mediated disorder is any one or more of the disorders as described herein.
  • Another aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of a CB2 receptor-mediated disorder.
  • the CB2 receptor-mediated disorder is any one or more of the disorders as described herein.
  • the analgesic properties of cannabinoids have been recognized for many years.
  • animal studies have demonstrated that the CBi/CB 2 agonists anandamide, THC, CP55,940 and WIN 55212-2 are effective against acute and chronic pain from chemical, mechanical, and thermal pain stimuli (reviewed in Walker and Huang (2002) Pharmacol. Ther. 95: 127-135; reviewed in Pacher, P et al. (2006) Pharmacol. Rev. 58(3): 389-462).
  • topical administration of the CBi/CB 2 agonist HU-210 attenuates capsaicin-induced hyperalgesia and allodynia (Rukwied, R. et al.
  • CB 2 also plays a role in mediating analgesic effects of cannabinoids (reviewed in Guindon and Hohmann (2008) Br. J. Pharmacol. 153:319-334).
  • systemic delivery of the CB 2 -selective agonist AM1241 suppresses hyperalgesia induced in the carrageenan, capsaicin, and formalin models of inflammatory pain in rodents (reviewed in Guindon and Hohmann (2008) Br. J. Pharmacol. 153:319-334).
  • Local (subcutaneous) or systemic administration of AM 1241 also reverses tactile and thermal hypersensitivity in rats following ligation of spinal nerves in the chronic constriction injury model of neuropathic pain (Malan, T.
  • CB 2 -specific agonists and/or CBi/CB 2 agonists find use in the treatment and/or prophylaxis of acute nociception and inflammatory hyperalgesia, as well as the allodynia and hyperalgesia produced by neuropathic pain.
  • these agonists are useful as an analgesic to treat pain arising from autoimmune conditions; allergic reactions; bone and joint pain; muscle pain; dental pain; nephritic syndrome; scleroderma; thyroiditis; migraine and other headache pain; pain associated with diabetic neuropathy; fibromyalgia, HIV-related neuropathy, sciatica, and neuralgias; pain arising from cancer; and pain that occurs as an adverse effect of therapeutics for the treatment of disease.
  • Another aspect of the present invention relates to methods for the treatment of pain in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
  • Some embodiments relate to methods for the treatment of pain associated with osteoarthritis in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
  • Some embodiments relate to methods for the treatment of neuropathic pain in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
  • Some embodiments relate to methods for the treatment of acute post-operative pain in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
  • Another aspect of the present invention relates to the use of a pharmaceutical composition as described herein, in the treatment of pain.
  • Some embodiments relate to the use of a pharmaceutical composition as described herein, in the treatment of pain associated with osteoarthritis.
  • Some embodiments relate to the use of a pharmaceutical composition as described herein, in the treatment of neuropathic pain.
  • Some embodiments relate to the use of a pharmaceutical composition as described herein, in the treatment of acute post-operative pain.
  • Another aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of pain.
  • Some embodiments relate to pharmaceutical compositions as described herein, for use in a method of treatment of pain associated with osteoarthritis.
  • Some embodiments relate to pharmaceutical compositions as described herein, for use in a method of treatment of neuropathic pain.
  • Some embodiments relate to pharmaceutical compositions as described herein, for use in a method of treatment of acute post-operative pain.
  • MS Multiple Sclerosis
  • EAE Experimental Autoimmune Encephalomyelitis
  • CB selective agonist WIN 55212-2 provides significant neuroprotection in the experimental allergic uveitis (EAU) model in mice (Pryce, G. et al.
  • CB 2 -selective agonist HU-308 markedly reduces the recruitment of immature myeloid cells and T cells, microglial and infiltrating myeloid cell proliferation, and axonal loss in the EAE model (Palazuelos, J. et al. (2008). J. Biol. Chem. 283(19): 13320-9).
  • the CBi/CB 2 agonist WIN 55212-2 significantly inhibits leukocyte rolling and adhesion in the brain in the EAE mouse model, an effect that is blocked by the CB 2 -selective antagonist SR144528 but not the CB selective antagonist SR141716A (Ni, X. et al. Mult. Sclerosis 10(2): 158-64).
  • CB 2 -selective agonists and/or CBi/CB 2 agonists find use in the treatment and/or prophylaxis of Multiple Sclerosis and related autoimmune demyelinating diseases, e.g. Guillan-Barre syndrome, polyradiculoneuropathy and chronic inflammatory demyelination.
  • autoimmune demyelinating diseases e.g. Guillan-Barre syndrome, polyradiculoneuropathy and chronic inflammatory demyelination.
  • the autoimmune disease Rheumatoid Arthritis is a chronic, systemic inflammatory disorder of the skeletal system that principally attacks the joints to produce an inflammatory synovitis and that often progresses to destruction of the articular cartilage and ankylosis of the joints.
  • the CBi/CBz agonists WIN 55212-2 and HU-210 significantly inhibit IL-1 alpha- stimulated proteoglycan and collagen degradation in bovine nasal cartilage explants in vitro
  • CB 2 -selective agonists and/or CBi/CB 2 agonists find use in the treatment and/or prophylaxis of autoimmune arthritic diseases, for example, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylarthritis, and reactive arthritis.
  • Type 1 Hypersensitivity and Allergic response Cannabinoid receptor agonists have been demonstrated to attenuate aberrant immune responses in allergic reactions as well.
  • type- 1, or immediate, hypersensitivity plasma cells that have been activated by an allergen secrete IgE antibodies, which bind to Fc receptors on the surface of tissue mast cells and blood basophils and eosinophils. Repeated exposure to the same allergen results in cross-linking of the bound IgE on sensitized cells, resulting in secretion of pharmacologically active mediators such as histamine, leukotriene and prostaglandin.
  • CB 2 -selective agonists and/or CBi/CBz agonists find use in the treatment of allergic reactions including atopic dermatitis (pruritis/itch), urticaria (hives), asthma, conjunctivitis, allergic rhinitis (hay fever), and anaphylaxis.
  • CB 2 agonists have been demonstrated to attenuate inflammation in the CNS. For example, administration of CB 2 agonists prevents the activation of microglia in rodent models of Alzheimer's Disease (Ashton J. C, et al. (2007) Curr. Neuropharmacol. 5(2):73-80). Likewise, administration of CB 2 agonists reduces the volume of infarcts by 30% in a rodent occlusion model of stroke (Zhang, M. et al. (2007) J. Cereb. Blood Flow Metab. 27: 1387-96). Thus, CB 2 agonists find use in the treatment and/or prophylaxis of neuropathologies associated with CNS inflammation, e.g. Alzheimer's, stroke-induced damage, dementia, ALS, and HIV.
  • CNS inflammation e.g. Alzheimer's, stroke-induced damage, dementia, ALS, and HIV.
  • CB 2 is expressed in macrophages and T cells in atherosclerotic plaques, and the CBi/CB 2 agonist THC reduces the progression of atherosclerosis in ApoE knockout mice, a well-studied mouse model of atherosclerosis.
  • the CB 2 -specific antagonist SRI 44528 completely blocks this effect in vitro and in vivo (Steffens, S. et al. (2005) Nature 434:782-786).
  • CB 2 agonists find use in treating atherosclerosis.
  • CB 2 agonists are useful for the treatment and/or prophylaxis of other disorders wherein undesired immune cell activity and/or inflammation is observed.
  • Such exemplary disorders include osteoarthritis, anaphylaxis, Behcet' s disease, graft rejection, vasculitis, gout, spondylitis, viral and bacterial diseases, e.g. AIDS, and meningitis; and other autoimmune disorders such as lupus, e.g. systemic lupus erythematosus;
  • inflammatory bowel disease e.g. Crohn' s disease, ulcerative colitis; psoriasis; autoimmune hepatitis; and type 1 diabetes mellitus.
  • Another aspect of the present invention relates to methods for the treatment of osteoarthritis in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
  • Another aspect of the present invention relates to the use of a pharmaceutical composition as described herein, in the treatment of osteoarthritis.
  • Another aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of osteoarthritis.
  • CB 2 is expressed in osteoblasts, osteocytes, and osteoclasts. Osteoblasts make new bone, whereas osteoclasts degrade it.
  • the CB 2 -specific agonist HU-308 enhances endocortical osteoblast numbers and activity while simultaneously inhibiting proliferation of osteoclast precursors in bone marrow-derived osteoblasts/stromal cells in vitro, and attenuates ovariectomy- induced bone loss and stimulates cortical thickness by stimulating endocortical bone formation and suppressing osteoclast number in vivo (Ofek, O. et al. (2006) Proc. Natl. Acad. Sci. 103(3):696-701).
  • CB 2 agonists are useful for the treatment and/or prophylaxis of disease wherein bone density is decreased, such as osteoporosis.
  • CB 2 -selective agonists and CBi/CB 2 agonists are useful for the treatment and/or prophylaxis of autoimmune arthritic diseases, for example, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylarthritis, and reactive arthritis, and for the treatment and/or prophylaxis of inflammation associated with osteoarthritis.
  • CB selective agonists and CBi/CB 2 agonists are useful for the treatment of pain associated with these arthritic disorders.
  • Retinal pigment epithelial (RPE) cells provide trophic support to photoreceptor cells in the eye, and RPE cell death has been demonstrated to be a major contributor to Age-related Macular
  • CB 2 -selective agonists find use in preventing the onset or progression of vision loss associated with AMD.
  • the cough reflex is predominantly under the control of two classes of sensory afferent nerve fibers, the myelinated A-delta fibers and the non-myelinated C-fibers, the activation of which (i.e. depolarization) elicits cough via the vagus nerve afferent pathway.
  • the CBi/CB 2 agonist CP55,940 reduces capsaicin-, PGE 2 - and hypertonic saline-induced depolarization of guinea pig and human vagus nerve preparations in vitro (Patel, H. J. et al. (2003) British J. Pharma. 140:261-8).
  • the CBi/CB 2 agonists WIN 55212-2 produced a dose-dependent inhibition of the number of capsaicin-induced coughs in mice (Morita, K. et al. (2003) Eur. J. Pharmacol. 474:269-272).
  • the CBi/CBz agonist anandamide produced a dose-dependent inhibition of the number of capsaicin-induced coughs in guinea pigs (Calignano, A. et al. (2000) Nature 408:96-101).
  • CB specific antagonist SR141716A attenuates the antitussive effects of WN 55212-2 and anandamide (Morita, K. et al. (2003) Eur. J. Pharmacol.
  • the CB 2 -selective agonist JWH133 reduces capsaicin-, PGE 2 - and hypertonic saline-induced depolarization of guinea pig and human vagus nerve preparations in vitro, and administration of CB 2 -selective agonist JWH133 prior to exposure to the tussive agent citric acid significantly reduces cough in conscious guinea-pigs (Patel, H. J. et al. (2003) British J. Pharma. 140:261-8).
  • CBi and CB 2 play an important role in mediating the antitussive effect of cannabinoids, and CB selective agonists and CBi/CB 2 agonists are useful in the treatment and/or prophylaxis of cough.
  • a number of human leukemia and lymphoma cell lines including Jurkat, Molt-4 and Sup-Tl, express CB 2 and not CBi, and agonists of CB 2 induce apoptosis in these and primary acute lymphoblastic leukemia (ALL) cells (Nagarkatti, L. C. et al. US2004/0259936).
  • ALL acute lymphoblastic leukemia
  • CB 2 is expressed on glioblastoma cell lines and treatment with agonists of CB 2 induces apoptosis of these cells in vitro (Widmer, M. (2008) /. Neurosci. Res. 86(14):3212-20).
  • CB 2 -selective agonists are useful in attenuating the growth of a malignancy of the immune system, for example, leukemias, lymphomas, and solid tumors of the glial lineage.
  • CBi-selective agonists and CBi/CB 2 agonists are useful in providing relief from pain associated with cancer (GW Pharmaceuticals press release Jan 19, 2005, Jun 19, 2007).
  • CB 2 -mediated signaling is involved in the in vivo and in vitro growth inhibition of prostate cancer cells, which suggests that CB 2 agonists have potential therapeutic interest in the management of prostate cancer.
  • CB 2 Agonists of CB 2 modulate the expansion of the progenitor pool of neurons in the CNS.
  • CB 2 antagonists inhibit the proliferation of cultured neural stem cells and the proliferation of progenitor cells in the SVZ of young animals, whereas CB 2 -selective agonists stimulate progenitor cell proliferation in vivo, with this effect being more pronounced in older animals (Goncalves, M. B. et al. (2008) Mol. Cell Neurosci. 38(4):526-36).
  • agonists of CB 2 are useful in regenerative medicine, for example to promote the expansion of progenitor cells for the replacement of neurons lost during injury or disease, such as Alzheimer's Disease, stroke-induced damage, dementia, amyotrophic lateral sclerosis (ALS) and Parkinson's Disease.
  • ALS amyotrophic lateral sclerosis
  • Polymorphism is the ability of a substance to exist as two or more crystalline phases that have different arrangements and/or conformations of the molecules in the crystal lattice. Polymorphs show the same properties in the liquid or gaseous state but they behave differently in the solid state.
  • drugs can also exist as salts and other multicomponent crystalline phases.
  • solvates and hydrates may contain an API host and either solvent or water molecules, respectively, as guests.
  • the guest compound is a solid at room temperature, the resulting form is often called a cocrystal.
  • Salts, solvates, hydrates, and cocrystals may show polymorphism as well. Crystalline phases that share the same API host, but differ with respect to their guests, may be referred to as pseudopolymorphs of one another.
  • Solvates contain molecules of the solvent of crystallization in a definite crystal lattice. Solvates, in which the solvent of crystallization is water, are termed hydrates. Because water is a constituent of the atmosphere, hydrates of drugs may be formed rather easily.
  • the present disclosure includes all isotopes of atoms occurring in the present compounds, salts, and crystalline forms thereof.
  • Isotopes include those atoms having the same atomic number but different mass numbers.
  • One aspect of the present invention includes every combination of one or more atoms in the present compounds, salts, and crystalline forms thereof that is replaced with an atom having the same atomic number but a different mass number.
  • One such example is the replacement of an atom that is the most naturally abundant isotope, such as 3 ⁇ 4 or 12 C, found in one of the present compounds, salts, and crystalline forms thereof, with a different atom that is not the most naturally
  • isotopes of hydrogen include 2 H (deuterium) and 3 H (tritium).
  • isotopes of carbon include n C, 13 C, and 14 C.
  • Isotopes of nitrogen include N and N.
  • Isotopes of oxygen include O, O, and C.
  • An isotope of fluorine includes 18 F.
  • An isotope of sulfur includes 35 S.
  • An isotope of chlorine includes 36 C1.
  • Isotopes of bromine include 75 Br, 76 Br, 77 Br, and 82 Br.
  • Isotopes of iodine include 123 I, 124 I, 125 I, and 131 I.
  • compositions such as, those prepared during synthesis, preformulation, and the like, and pharmaceutical compositions, such as, those prepared with the intent of using in a mammal for the treatment of one or more of the disorders described herein, comprising one or more of the present compounds, salts, and crystalline forms thereof, wherein the naturally occurring distribution of the isotopes in the composition is perturbed.
  • compositions and pharmaceutical compositions comprising a compound, a salt, or a crystalline form thereof as described herein wherein the compound, salt, or crystalline form is enriched at one or more positions with an isotope other than the most naturally abundant isotope.
  • compositions and Formulations are readily available to measure such isotope perturbations or enrichments, such as, mass spectrometry, and for isotopes that are radio-isotopes additional methods are available, such as, radio-detectors used in connection with HPLC or GC.
  • compositions and/or pharmaceutical compositions comprising: the anhydrous crystalline form of (la l S',5aS')-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2- dimethyl-propyl) -amide (Compound 1); and at least one compound selected from: a compound of Formulae (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Iff), (Hf-i), (Hg), (Hh), (Ilia),
  • compositions comprising:
  • compositions comprising:
  • compositions comprising:
  • compositions comprising: a. the anhydrous crystalline form of (laS',5a l S')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymefhyl-2,2-dimefhyl- propyl)-amide (Compound 1);
  • the composition is substantially free of methylene chloride.
  • substantially free of methylene chloride refers to the amount of methylene chloride present in a composition or pharmaceutical composition as being 600 ppm or less as determined by GC, such as head space GC.
  • the composition is free of methylene chloride.
  • the composition further comprises a pharmaceutically acceptable carrier.
  • the pharmaceutical composition is substantially free of methylene chloride.
  • the pharmaceutical composition is free of methylene chloride.
  • the compound is 3-((4a ⁇ ,5a5)-3-(((.S , )-3,3-dimethyl-l-(((4a l S , ,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia).
  • 1-oxide of Formula (Ilia) is present in the composition of less than or equal to 1.0% as determined by HPLC.
  • 1-oxide of Formula (IHa) is present in the composition of less than or equal to 0.5% as determined by HPLC.
  • 1-oxide of Formula (Ilia) is present in the composition of less than or equal to 0.2% as determined by HPLC.
  • the compound is any organic compound. In some embodiments for the composition and/or pharmaceutical composition, the compound is
  • (4a,S , ,5a l S , )-iV-(( l S , )-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb) is present in the composition of less than or equal to 1.0% as determined by HPLC. In some embodiments,
  • the compound is acetone.
  • acetone is present in the composition of less than or equal to 5000 ppm as determined by GC. In some embodiments, acetone is present in the composition of less than or equal to 2500 ppm as determined by GC. In some embodiments, acetone is present in the composition of less than or equal to 494 ppm as determined by GC.
  • the compound is ethyl acetate. In some embodiments, ethyl acetate is present in the composition of less than or equal to 5000 ppm as determined by GC. In some embodiments, ethyl acetate is present in the composition of less than or equal to 2500 ppm as determined by GC. In some embodiments, ethyl acetate is present in the composition of less than or equal to 5 ppm as determined by GC.
  • the pharmaceutical composition is suitable for oral, rectal, nasal, topical, buccal, sub-lingual, vaginal, parenteral, intramuscular, sub-cutaneous, or intravenous administration, or in a form suitable for administration by inhalation, insufflation, or by a transdermal patch. In some embodiments, the pharmaceutical composition is suitable for oral administration.
  • Formulations may be prepared by any suitable method, typically by uniformly mixing the active compound(s) with liquids or finely divided solid carriers, or both, in the required proportions and then, if necessary, forming the resulting mixture into a desired shape.
  • Liquid preparations for oral administration can be in the form of solutions, emulsions, aqueous or oily suspensions and syrups.
  • the oral preparations can be in the form of dry powder that can be reconstituted with water or another suitable liquid vehicle before use. Additional additives such as suspending or emulsifying agents, non-aqueous vehicles (including edible oils), preservatives and flavorings and colorants can be added to the liquid preparations.
  • Parenteral dosage forms can be prepared by dissolving the compound provided herein in a suitable liquid vehicle and filter sterilizing the solution before filling and sealing an appropriate vial or ampule. These are just a few examples of the many appropriate methods well known in the art for preparing dosage forms.
  • a compound provided herein can be formulated into pharmaceutical compositions using techniques well known to those in the art. Suitable pharmaceutically-acceptable carriers, outside those mentioned herein, are known in the art; for example, see Remington, The Science and Practice of Pharmacy, 20 th Edition, 2000, Lippincott Williams & Wilkins, (Editors: Gennaro et al).
  • a compound provided herein can, in an alternative use, be administered as a raw or pure chemical, it is preferable however to present the compound or active ingredient as a pharmaceutical formulation or composition further comprising a pharmaceutically acceptable carrier.
  • Transdermal patches dispense a drug at a controlled rate by presenting the drug for absorption in an efficient manner with minimal degradation of the drug.
  • transdermal patches comprise an impermeable backing layer, a single pressure sensitive adhesive and a removable protective layer with a release liner.
  • the compounds provided herein, together with a conventional adjuvant, carrier, or diluent, can thus be placed into the form of pharmaceutical formulations and unit dosages thereof and in such form may be employed as solids, such as tablets or filled capsules, or liquids such as solutions, suspensions, emulsions, elixirs, gels or capsules filled with the same, all for oral use, in the form of suppositories for rectal administration; or in the form of sterile injectable solutions for parenteral (including
  • Such pharmaceutical compositions and unit dosage forms thereof can comprise conventional ingredients in conventional proportions, with or without additional active compounds or principles and such unit dosage forms may contain any suitable effective amount of the active ingredient commensurate with the intended daily dosage range to be employed.
  • the pharmaceutical composition may be in the form of, for example, a tablet, capsule, suspension or liquid.
  • the pharmaceutical composition is preferably made in the form of a dosage unit containing a particular amount of the active ingredient.
  • dosage units are capsules, tablets, powders, granules or a suspension, with conventional additives such as lactose, mannitol, corn starch or potato starch; with binders such as crystalline cellulose, cellulose derivatives, acacia, corn starch or gelatins; with disintegrators such as corn starch, potato starch or sodium carboxymethyl-cellulose; and with lubricants such as talc or magnesium stearate.
  • the active ingredient may also be administered by injection as a composition wherein, for example, saline, dextrose or water may be used as a suitable pharmaceutically acceptable carrier.
  • active ingredient defined in the context of a “pharmaceutical composition”,” refers to a component of a pharmaceutical composition that provides the primary pharmacological effect, as opposed to an "inactive ingredient” which would generally be recognized as providing no pharmaceutical benefit.
  • the dose when using the compounds provided herein can vary within wide limits and as is customary and is known to the physician, it is to be tailored to the individual conditions in each individual case. It depends, for example, on the nature and severity of the illness to be treated, on the condition of the individual, such as a patient, on the compound employed, on whether an acute or chronic disease state is treated, or prophylaxis conducted, or on whether further active compounds are administered in addition to the compounds provided herein.
  • Representative doses include, but are not limited to, about 0.001 mg to about 5000 mg, about 0.001 mg to about 2500 mg, about 0.001 mg to about 1000 mg, about 0.001 mg to about 500 mg, about 0.001 mg to about 250 mg, about 0.001 mg to 100 mg, about 0.001 mg to about 50 mg and about 0.001 mg to about 25 mg.
  • Multiple doses may be administered during the day, especially when relatively large amounts are deemed to be needed, for example 2, 3 or 4 doses. Depending on the individual and as deemed appropriate from the healthcare provider it may be necessary to deviate upward or downward from the doses described herein.
  • the amount of active ingredient, or an active salt or derivative thereof, required for use in treatment will vary not only with the particular salt selected but also with the route of administration, the nature of the condition being treated and the age and condition of the individual and will ultimately be at the discretion of the attendant physician or clinician.
  • a model system typically an animal model
  • these extrapolations may merely be based on the weight of the animal model in comparison to another, such as a mammal, preferably a human, however, more often, these extrapolations are not simply based on weights, but rather incorporate a variety of factors.
  • compositions and/or compositions provided herein are selected in accordance with a variety factors as cited above.
  • the actual dosage regimen employed may vary widely and therefore may deviate from a preferred dosage regimen and one skilled in the art will recognize that dosage and dosage regimen outside these typical ranges can be tested and, where appropriate, may be used in the methods disclosed herein.
  • the desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four, or more sub-doses per day.
  • the sub-dose itself may be further divided, e.g., into a number of discrete loosely spaced administrations.
  • the daily dose can be divided, especially when relatively large amounts are administered as deemed appropriate, into several, for example 2, 3, or 4 part administrations. If appropriate, depending on individual behavior, it may be necessary to deviate upward or downward from the daily dose indicated.
  • the compounds provided herein can be administered in a wide variety of oral and parenteral dosage forms. It will be obvious to those skilled in the art that the dosage forms may comprise, as the active component, either a compound provided herein or a pharmaceutically acceptable salt, hydrate, or solvate of a compound provided herein.
  • a suitable pharmaceutically acceptable carrier can be either solid, liquid or a mixture of both.
  • Solid form preparations include powders, tablets, pills, capsules, cachets, suppositories and dispersible granules.
  • a solid carrier can be one or more substances which may also act as diluents, flavoring agents, solubilizers, lubricants, suspending agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material.
  • the carrier is a finely divided solid which is in a mixture with the finely divided active component.
  • the active component is mixed with the carrier having the necessary binding capacity in suitable proportions and compacted to the desired shape and size.
  • the powders and tablets may contain varying percentage amounts of the active compound.
  • a representative amount in a powder or tablet may contain from 0.5 to about 90 percent of the active compound; however, an artisan would know when amounts outside of this range are necessary.
  • Suitable carriers for powders and tablets are magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter and the like.
  • the term "preparation” refers to the formulation of the active compound with encapsulating material as carrier providing a capsule in which the active component, with or without carriers, is surrounded by a carrier, which is thus in association with it.
  • cachets and lozenges are included. Tablets, powders, capsules, pills, cachets and lozenges can be used as solid forms suitable for oral administration.
  • a low melting wax such as an admixture of fatty acid glycerides or cocoa butter
  • the active component is dispersed homogeneously therein, as by stirring.
  • the molten homogenous mixture is then poured into convenient sized molds, allowed to cool and thereby to solidify.
  • Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or sprays containing in addition to the active ingredient such carriers as are known in the art to be appropriate.
  • Liquid form preparations include solutions, suspensions and emulsions, for example, water or water-propylene glycol solutions.
  • parenteral injection liquid preparations can be formulated as solutions in aqueous polyethylene glycol solution.
  • injectable preparations for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents.
  • the sterile injectable preparation may also be a sterile injectable solution or suspension in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol.
  • Suitable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil may be employed including synthetic mono- or diglycerides.
  • fatty acids such as oleic acid find use in the preparation of injectables.
  • the compounds provided herein may thus be formulated for parenteral administration (e.g. by injection, for example bolus injection or continuous infusion) and may be presented in unit dose form in ampoules, pre-filled syringes, small volume infusion or in multi-dose containers with an added preservative.
  • the pharmaceutical compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
  • the active ingredient may be in powder form, obtained by aseptic isolation of sterile solid or by lyophilization from solution, for constitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use.
  • Aqueous formulations suitable for oral use can be prepared by dissolving or suspending the active component in water and adding suitable colorants, flavors, stabilizing and thickening agents, as desired.
  • Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, or other well-known suspending agents. Also included are solid form preparations which are intended to be converted, shortly before use, to liquid form preparations for oral administration. Such liquid forms include solutions, suspensions and emulsions. These preparations may contain, in addition to the active component, colorants, flavors, stabilizers, buffers, artificial and natural sweeteners, dispersants, thickeners, solubilizing agents and the like.
  • the compounds provided herein may be formulated as ointments, creams or lotions, or as a transdermal patch.
  • Ointments and creams may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening and/or gelling agents.
  • Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilizing agents, dispersing agents, suspending agents, thickening agents, or coloring agents.
  • Formulations suitable for topical administration in the mouth include lozenges comprising active agent in a flavored base, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert base such as gelatin and glycerin or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier.
  • Solutions or suspensions are applied directly to the nasal cavity by conventional means, for example with a dropper, pipette or spray.
  • the formulations may be provided in single or multi-dose form. In the latter case of a dropper or pipette, this may be achieved by the patient administering an appropriate, predetermined volume of the solution or suspension. In the case of a spray, this may be achieved for example by means of a metering atomizing spray pump.
  • Administration to the respiratory tract may also be achieved by means of an aerosol formulation in which the active ingredient is provided in a pressurized pack with a suitable propellant.
  • aerosol formulation in which the active ingredient is provided in a pressurized pack with a suitable propellant.
  • the compounds provided herein or pharmaceutical compositions comprising them are administered as aerosols, for example as nasal aerosols or by inhalation, this can be carried out, for example, using a spray, a nebulizer, a pump nebulizer, an inhalation apparatus, a metered inhaler or a dry powder inhaler.
  • Pharmaceutical forms for administration of the compounds provided herein as an aerosol can be prepared by processes well known to the person skilled in the art.
  • solutions or dispersions of the compounds provided herein in water, water/alcohol mixtures or suitable saline solutions can be employed using customary additives, for example benzyl alcohol or other suitable preservatives, absorption enhancers for increasing the bioavailability, solubilizers, dispersants and others and, if appropriate, customary propellants, for example include carbon dioxide, CFCs, such as, dichlorodifluoromethane, trichlorofluoromethane, or dichlorotetrafluoroethane; and the like.
  • the aerosol may conveniently also contain a surfactant such as lecithin.
  • the dose of drug may be controlled by provision of a metered valve.
  • the compound In formulations intended for administration to the respiratory tract, including intranasal formulations, the compound will generally have a small particle size for example of the order of 10 microns or less. Such a particle size may be obtained by means known in the art, for example by micronization. When desired, formulations adapted to give sustained release of the active ingredient may be employed.
  • the active ingredients may be provided in the form of a dry powder, for example, a powder mix of the compound in a suitable powder base such as lactose, starch, starch derivatives such as hydroxypropylmethyl cellulose and polyvinylpyrrolidone (PVP).
  • a powder mix of the compound in a suitable powder base such as lactose, starch, starch derivatives such as hydroxypropylmethyl cellulose and polyvinylpyrrolidone (PVP).
  • PVP polyvinylpyrrolidone
  • the powder carrier will form a gel in the nasal cavity.
  • the powder composition may be presented in unit dose form for example in capsules or cartridges of, e.g., gelatin, or blister packs from which the powder may be administered by means of an inhaler.
  • the pharmaceutical preparations are preferably in unit dosage forms.
  • the preparation is subdivided into unit doses containing appropriate quantities of the active component.
  • the unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packeted tablets, capsules and powders in vials or ampoules.
  • the unit dosage form can be a capsule, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form.
  • Tablets or capsules for oral administration and liquids for intravenous administration are preferred compositions.
  • the compounds provided herein may optionally exist as pharmaceutically acceptable salts including pharmaceutically acceptable acid addition salts prepared from pharmaceutically acceptable non-toxic acids including inorganic and organic acids.
  • Representative acids include, but are not limited to, acetic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethenesulfonic, dichloroacetic, formic, fumaric, gluconic, glutamic, hippuric, hydrobromic, hydrochloric, isethionic, lactic, maleic, malic, mandelic, methanesulfonic, mucic, nitric, oxalic, pamoic, pantothenic, phosphoric, succinic, sulfiric, tartaric, oxalic, j-toluenesulfonic and the like.
  • Certain compounds provided herein which contain a carboxylic acid functional group may optionally exist as pharmaceutically acceptable salts containing non-toxic, pharmaceutically acceptable metal cations and c
  • Representative metals include, but are not limited to, aluminium, calcium, lithium, magnesium, potassium, sodium, zinc and the like. In some embodiments the pharmaceutically acceptable metal is sodium.
  • Representative organic bases include, but are not limited to, benzathine (TV ⁇ A ⁇ -dibenzylethane- 1,2-diamine), chloroprocaine (2-(diethylamino)ethyl 4-(chloroamino)benzoate), choline,
  • the acid addition salts may be obtained as the direct products of compound synthesis.
  • the free base may be dissolved in a suitable solvent containing the appropriate acid and the salt isolated by evaporating the solvent or otherwise separating the salt and solvent.
  • the compounds provided herein may form solvates with standard low molecular weight solvents using methods known to the skilled artisan.
  • Compounds provided herein can be converted to "pro-drugs.”
  • the term “pro-drugs” refers to compounds that have been modified with specific chemical groups known in the art and when administered into an individual these groups undergo biotransformation to give the parent compound. Pro-drugs can thus be viewed as compounds provided herein containing one or more specialized nontoxic protective groups used in a transient manner to alter or to eliminate a property of the compound.
  • the "pro-drug” approach is utilized to facilitate oral absorption.
  • a thorough discussion is provided in T. Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems Vol. 14 of the A.C.S. Symposium Series; and in Bioreversible Carriers in Drug Design, ed. Edward B. Roche, American Pharmaceutical Association and Pergamon Press, 1987.
  • “combination-therapy” comprising admixing at least one compound according to any of the compound embodiments disclosed herein, together with at least one known pharmaceutical agent as described herein and a pharmaceutically acceptable carrier.
  • cannabinoid CB 2 receptor modulators are utilized as active ingredients in pharmaceutical compositions, these are not intended for use in humans only, but in non-human mammals as well.
  • active agents such as cannabinoid CB 2 receptor modulators
  • livestock animals e.g., horses, cows, etc.
  • the reactor contents were cooled to 0°C and 3M HC1 (6.5 L) was added to the mixture slowly so as to maintain an internal temperature of ⁇ 10°C.
  • the phases were separated and the organic layer washed with 3M HC1 (2.6 L).
  • the combined aqueous layer was extracted twice with 5.8 kg and 2.4 kg of MTBE respectively.
  • the combined MTBE layers were concentrated to one half the original volume and was used directly in the oxidation step, see Example 2.
  • a buffer solution was made by adding in a 50 L reactor, potassium phosphate dibasic (3.5 kg), potassium phosphate monobasic (0.37 kg), potassium bromide (0.24 kg), and water (6.5 L). The mixture was stirred at ambient for 5 minutes, at which time a clear solution formed.
  • the buffer solution was charged to a 50 L reactor containing (lS,2S,5R)-bicyclo[3.1.0]hexan-2-ol (prepared according to Example 1).
  • TEMPO (0.06 kg) was charged to the reaction mixture followed by the controlled addition of sodium hypochlorite (11.4 L) via an addition funnel so as to keep the temperature below 10°C. The reaction was deemed complete when the amount of (lS,2S,5R)-bicyclo[3.1.0]hexan-2-ol was less than 2% (peak area) as determined by GC.
  • the reaction mixture was quenched with 2.4M aqueous sodium sulfite solution (2.6 L) until the reaction was negative for oxidant by starch iodide indicator paper.
  • the mixture was filtered and the filtrate set aside.
  • the filter cake was washed with MTBE (2.93 kg).
  • the filtrate and the wash were combined and transferred to the reactor.
  • the phases were separated and the aqueous phase was extracted twice with 4.8 kg and 2.4 kg of MTBE respectively.
  • Example 3 Preparation of (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/ - cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He)).
  • the internal temperature of the reaction mixture was kept between 35-40°C and the mixture monitored by HPLC for the disappearance of starting material (i.e., (lS,5R)-bicyclo[3.1.0]hexan-2-one).
  • the reactor contents were then cooled to 30°C before adding 2-hydrazinylpyrazine (2.43 kg, 22.07 mol, 1.06 eq.) to the reactor.
  • 2-hydrazinylpyrazine (2.43 kg, 22.07 mol, 1.06 eq.
  • 6N HC1 (6.76 L) was charged slowly to the reactor while maintaining an internal temperature of 35-40°C.
  • the mixture was then stirred at 35-40°C and monitored for reaction completion by HPLC.
  • the reaction was considered complete when the diketo methyl ester intermediate of Formula (Ilc-i) was less than 3 area % as determined by HPLC
  • the mixture was concentrated by distillation under vacuum at a jacket temperature of 25-50°C until no appreciable distillate was observed.
  • Water (19.3 kg) was charged to the reactor and the mixture concentrated by distillation under maximum vacuum at a jacket temperature of 25-50°C until no appreciable distillate was observed.
  • Water (9.65kg) was added to the reaction mixture and the mixture stirred at 20-25°C for at least 1 hour.
  • the mixture was filtered and the filter cake washed with water (2 x 9.65 kg).
  • the solids were placed in a vacuum oven set at 45-55°C and dried to a constant weight.
  • a powder X-ray diffraction (PXRD) pattern for a sample containing (4aS',5a l S')-methyl 1- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is shown in Figure 1.
  • Certain PXRD peaks for (4a l S',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate are shown in Table 3.
  • Example 4 Preparation of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l//- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (IIf».
  • reaction mixture was stirred under nitrogen at 45°C for over 3 hours until the amount of starting material (i.e., (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate) was less than 0.5% (area) as determined by HPLC.
  • starting material i.e., (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate
  • the mixture was concentrated by distillation under vacuum at a jacket temperature of 25-50°C until no appreciable distillate was observed. Thereafter, ethyl acetate (41.3 kg) was charged to the reactor and the mixture was concentrated by distillation under maximum vacuum at a jacket temperature of 25-50°C until 2 to 5 volumes remained in the reactor.
  • Water (2.5 kg) was charged to the reactor followed by acetone (12.2 kg) and the mixture was heated to reflux for at least 1 hour.
  • the mixture was cooled to 50°C and acetone (23.9 kg) was added to the reaction mixture to form a slurry comprising crystalline product.
  • the mixture was stirred at 20-25°C for 1 hour.
  • the mixture was filtered and the filter cake washed with acetone (24.4 kg).
  • Example 5 Preparation of 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l//- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Formula (Ilg)).
  • Hydrogen peroxide solution (30 wt.% aqueous; 5.54 kg, 48.9 moles, 4.03 eq.) was charged in four portions over a 4-hour period while maintaining the stirred reactor contents at a temperature not to exceed 60°C with reactor jacket cooling.
  • the reaction mixture was stirred under nitrogen at 55°C for over 2 hours until the amount of free acid (i.e., (laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3- diaza-cyclopropa[a]pentalene-4-carboxylic acid of Formula (Ilf-i)) was less than 0.6% (area) as determined by HPLC.
  • a powder X-ray diffraction (PXRD) pattern for a sample containing 3-((4a l S',5a l S r )-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is shown in Figure 3.
  • Certain powder X-ray diffraction peaks for 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide are shown in Table 5.
  • Example 6 Preparation of (la5,5a5)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/ -2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2-dimethyl-propyl)-amide (Compound 1, Formula (I)).
  • the progress of the reaction is monitored by HPLC and the reaction was deemed complete when the amount of starting material (i.e., 3-((4a l S',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide) was less than 0.5% (area) as determined by HPLC.
  • starting material i.e., 3-((4a l S',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide
  • the filter cake was transferred to the reactor and dissolved in acetonitrile (3.84 kg)/water (1.62 kg) at 55-65°C. The solution was cooled to 35-40°C and water (8.10 kg) was added slowly. The mixture was stirred at 35-40°C vigorously for a minimum of 2 hours when solids began to form.
  • Example 7 Evaluation of the oxidation step in the preparation of 3-((4a5,5aS)-3-carboxy- 4,4a,5,5a-tetrahydro-l/ -cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Formula dig)).
  • volume used in this example is merely a scaling factor and refers to the number of milliliters used for a respective solvent per gram of Compound of Formula (Ilf), for example formic acid (10 volume) refers to 5 mL/0.5 gram, 10 mL/1 gram, 50 mL/5 grams, 100 mL/10 gram, and the like of formic acid/Compound of
  • the oxidation step was modified reducing the amount of formic acid from 10 to 6 volumes with hydrogen peroxide (30 wt.%).
  • the progress of the reaction was followed by HPLC for the conversion of the Compound of Formula (Ilf) to the Compound of Formula (Ilg) and is shown below in Table 8.
  • Table 11 shows the progress of the oxidation step using formic acid (3 volumes) and trifluoroacetic acid (3 vol) with 30 wt.% hydrogen peroxide at 50-55°C.
  • Example 8 Representative gas chromatography (GC) methods for in-process control (IPC) and/or assessing chiral purity for (S)-l,2-epoxy-5-hexene (Formula (Ila)), (1S,2S,5R)- bicyclo[3.1.0]hexan-2-ol (Formula (lib)), and (15,5tf )-bicyclo[3.1.0]hexan-2-one (Formula (lie)).
  • GC gas chromatography
  • the sample was prepared as a solution in acetonitrile. The sample was then injected onto a
  • the chiral purity was determined quantitatively by comparing the peak area ratios for the sample chromatograms to the corresponding peak area ratios for the standard chromatograms.
  • the method conditions and a representative chromatogram for the sample solution are presented below:
  • the sample was prepared as a solution in acetonitrile.
  • the sample was then injected onto a Restek Rtx-5 capillary GC column and monitored using a flame ionization detector (FID).
  • FID flame ionization detector
  • the chiral purity was determined quantitatively by comparing the peak area ratios for the sample chromatograms to the corresponding peak area ratios for the standard chromatograms.
  • the method conditions and a representative chromatogram for the sample solution are presented below:
  • the sample was prepared as a solution in acetonitrile.
  • the sample was then injected onto a Restek RtyDEXsa GC column and monitored using a flame ionization detector (FID).
  • FID flame ionization detector
  • the chiral purity was determined quantitatively by comparing the peak area ratios for the sample chromatograms to the corresponding peak area ratios for the standard chromatograms.
  • the method conditions and a representative chromatogram for the sample solution are presented below:
  • Example 9 Representative high-performance liquid chromatography/high-pressure liquid chromatography (HPLC) methods for in-process control (IPC) and/or assessing purity for:
  • the sample solution in acetonitrile was injected onto an Agilent Poroshell 120 EC-C18 column and monitored by UV detection at 268 nm.
  • the working concentration was about 0.18 mg/mL for assay, related substance, and identification.
  • the percent assay of (4a l S',5aS')-methyl l-(pyrazin-2-yl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He)) in the starting material is determined against an external reference standard(s).
  • the positive identification of (He) is established by comparing the retention time of the (He) peak against that of the reference standard.
  • the method conditions are presented below:
  • Example 10 PathHunter ⁇ -arrestin assay; (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l//-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimethyl- propyl)-amide (Compound 1, Formula (I)).
  • Compound 1 was screened for agonist activity against the human CB2 (hCB2) receptor using the DiscoveRx PathHunter ⁇ -arrestin assay which measures the ⁇ -arrestin binding to the CB2 receptor upon its activation.
  • CB2 was cloned into the pCMV-PK vector (DiscoveRx, Fremont, CA; catalog # 93- 0167) and transfected into the CHO-K1 EA-Arrestin parental cell line (DiscoveRx, Fremont, CA; catalog # 93-0164).
  • CHO-K1 positive clones stably expressing the CB2-ProLink fusion protein were identified by their responses to the CB2 agonist CP55,940.
  • Clone # 61 was chosen for its big agonist window and homogenous expression as detected by anti-HA flow cytometry.
  • the PathHunter ⁇ -arrestin assay measures the interaction of ⁇ -arrestin with activated GPCRs using Enzyme Fragment Complementation (Yan et al., J. Biomol. Screen. 7: 451- 459, 2002).
  • a small, 42 amino acid ⁇ -galactosidase fragment, Prolink is fused to the c-terminus of a GPCR, and ⁇ -arrestin is fused to the larger ⁇ -galactosidase fragment, EA (Enzyme Acceptor).
  • Binding of ⁇ -arrestin to the activated GPCR causes the complementation of the two enzyme fragments, forming an active ⁇ -galactosidase enzyme which can be measured using the chemiluminiescent PathHunter Flash Detection Kit (DiscoveRx, Fremont, CA: catalog # 93-0001).
  • the assay The stable CHO-K1 cells expressing CB2-Prolink fusion protein were plated overnight in 384-well plates (Optiplate 384-Plus, PerkinElmer, Fremont CA; catalog # 6007299) at 5000 cells/5 ⁇ in the Opti-MEM medium (Invitrogen, Carlsbad, CA; catalog # 31985088) with 1 % FBS. 5uL of test compound diluted in Opti-MEM supplemented with 1 % BSA was transferred to each well of the Optiplate. The plates were then incubated at 37°C/5 C0 2 for two hours.
  • Assay readout ⁇ -Arrestin assay readout was accomplished using a PHERAstar (BMG Labtech Inc., Durham, NC) or an En VisionTM (PerkinElmer, Fremont CA) microplate reader. (1 ⁇ 5,5 ⁇ 3 ⁇ 4-2-(4- ⁇ ⁇ - ⁇ -2- ⁇ 1)-1 ⁇ ,2,5,5 ⁇ - ⁇ - 1/ ⁇ -2,3-(1 ⁇ - ⁇ 1 ⁇ [ ⁇ ] ⁇ 1 ⁇ -4- carboxylic acid ((3 ⁇ 4-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide (Compound 1) was observed to be a selective agonist for CB2 with an EC5 0 value of 5.4 nM and substantially inactive against hCBl .
  • Example 11 Effect of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/ -2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2-dimethyl-propyl)-amide (Compound 1) on osteoarthritis pain.
  • MIA monosodium iodoacetate
  • the osteoarthritis was induced in 200 g male Sprague Dawley rats. After brief anesthesia by isoflurane, rats received a single intra-articular injection of MIA (2 mg) (Sigma Aldrich, Saint Louis, MO, USA; Cat # 19148) dissolved in 0.9% sterile saline in a 50 volume administered through the patella ligament into the joint space of the left knee with a 30G needle. Following the injection, animals were allowed to recover from anesthesia before being returned to the main housing vivarium.
  • MIA Sigma Aldrich, Saint Louis, MO, USA
  • Hind paw weight distribution was determined by placing rats in a chamber so that each hind paw rests on a separate force plate of the incapacitance tester. The force exerted by each hind limb (measured in grams) is averaged over a 3 second period. Three measurements were taken for each rat, and the change in hind paw weight distribution calculated. Peak hind limb grip force was conducted by recording the maximum compressive force exerted on the hind limb mesh gauge set on the grip strength meter. During the testing, each rat was restrained and the paw of the injected knee was allowed to grip the mesh. The animal was then pulled in an upward motion until their grip was broken. Each rat is tested 3 times, with the contralateral paw used as a control.
  • Example 12 Powder X-ray Diffraction.
  • Powder X-ray Diffraction (PXRD) data were collected on an X'Pert PRO MPD powder diffractometer (PANalytical, Inc.) with a Cu source set at 45 kV and 40 mA, Cu(Ka) radiation and an X'Celerator detector. Samples were added to the sample holder and smoothed flat with a spatula and weigh paper. With the samples spinning, X-ray diffractograms were obtained by a 12-min scan over the 2-theta range 5-40 °2 ⁇ . Diffraction data were viewed and analyzed with the X'Pert Data Viewer Software, version 1.0a and X'Pert HighScore Software, version 1.0b.
  • DSC Differential scanning calorimetry
  • Thermogravimetric analysis was conducted using a TA Instruments TGA Q500 or Q5000 at a heating rate 10°C/min. The instrument was calibrated using a standard weight for the balance, and Alumel and Nickel standards for the furnace (Curie point measurements). Thermal events such as weight-loss are calculated using the Universal Analysis 2000 software, version 4. ID, Build 4.1.0.16.
  • thermogravimetric profile for the anhydrous crystalline form of Compound 1 is shown in Figure 5.
  • Example 15 Dynamic Moisture-Sorption Analysis.
  • DMS dynamic moisture-sorption
  • VTI dynamic moisture-sorption analyzer
  • SGA-100 static moisture-sorption analyzer
  • the instrument was calibrated using polyvinyl pyrrolidone (PVP) and NaCl.
  • Samples were prepared for DMS analysis by placing 5 mg to 20 mg of a sample in a tared sample holder. The sample was placed on the hang-down wire of the VTI balance.
  • a drying step was run, typically at 40°C and 0.5-1% RH for 1 h.
  • the isotherm temperature is 25°C.
  • % RH holds typically ranged from 10% RH to 90% RH, with intervals of 10 to 20% RH.
  • the water content of the sample equilibrated as described above was determined at each % RH hold.
  • Example 16 Analytical characterization of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro-l//-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2- dimethyl-propyl)-amide (Compound 1, Formula (I)).
  • Example 17 Headspace gas chromatography method for residual solvents presence in intermediates and Compound 1.
  • HGC Headspace gas chromatography
  • the GC cycle time in the table is a recommendation. This can vary from instrument to instrument.
  • the GC cycle time should be equal to the GC run time for the GC to equilibrate to initial conditions.
  • the method described herein has been validated, such as, specificity, linearity, accuracy, quantitation limit, detection limit, and solution stability, for methanol, ethanol, acetone, acetonitrile, methyl ieri-butyl ether, and ethyl acetate.
  • the method has certain limitations for the isomers of hexanes, for example, the separation of MTBE could not be achieved from one of the isomers of hexanes.
  • Hexanes contained 5 different isomers labeled Hexane A to Hexane E.
  • Hexane C is the second most abundant isomer of hexanes and a similar retention time as MTBE.
  • Samples were prepared using 100 mg in 1.00 mL DMSO and were used as the basis for all ppm concentrations using 20 mL borosilicate glass headspace vials with silicone/PTFE septa. The method was based on the published headspace GC method (e.g., Dai L, et. al. (2010) LCGC North America, 28 (1), 73-84).

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Abstract

The present invention relates to processes, and process intermediates useful in the preparation of (1aS,5aS)-2-(4-oxy-pyrazin-2-yl)-1a,2,5,5a-tetrahydro-1H-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-1-hydroxymethyl-2,2-dimethyl-propyl)-amide (Compound 1) and crystalline forms thereof. The compound (1aS,5aS)-2-(4-oxy-pyrazin-2-yl)-1a,2,5,5a-tetrahydro-1H-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-1-hydroxymethyl-2,2-dimethyl-propyl)-amide has been identified as a modulator of the cannabinoid 2 (CB2) receptor and useful in the treatment of CB2 receptor-mediated disorders, for example, pain, disorders of the immune system (such as, autoimmune disorders, type 1 hypersensitivity and allergic response, conditions associated with CNS inflammation, conditions associated with vascular inflammation, and other disorders associated with aberrant or unwanted immune response), bone and joint diseases (such as, osteoporosis and arthritis), eye disease, cough, cancer, and regenerative medicine. The present invention further provides pharmaceutical compositions comprising Compound 1 and the use of the pharmaceutical compositions for the treatment of CB2 receptor-mediated disorders.

Description

PROCESSES FOR THE PREPARATION OF CANNABINOID RECEPTOR MODULATORS
FIELD OF THE INVENTION
The present invention relates to processes, and process intermediates useful in the preparation of (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide (Compound 1) and crystalline forms thereof. The compound (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide has been identified as a modulator of the cannabinoid 2 (CB2) receptor and useful in the treatment of CB2 receptor-mediated disorders, for example, osteoarthritis; pain, for example bone and joint pain, muscle pain, dental pain, migraine and other headache pain, inflammatory pain, neuropathic pain, pain that occurs as an adverse effect of therapeutics, and pain associated with a disorder selected from:
osteoarthritis, cancer, multiple sclerosis, allergic reactions, nephritic syndrome, scleroderma, thyroiditis, diabetic neuropathy, fibromyalgia, HIV related-neuropathy, sciatica, and autoimmune conditions; hyperalgesia; allodynia; inflammatory hyperalgesia; neuropathic hyperalgesia; acute nociception; osteoporosis; multiple sclerosis-associated spasticity; autoimmune disorders, for example an autoimmune disorder selected from the group consisting of: multiple sclerosis, Guillan-Barre syndrome, polyradiculoneuropathy, chronic inflammatory demyelination, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylarthritis, and reactive arthritis; allergic reactions, for example, an allergic reaction associated with a disorder selected from: atopic dermatitis, pruritus, urticaria, asthma, conjunctivitis, allergic rhinitis, and anaphylaxis; CNS inflammation for example, CNS inflammation associated with a disorder selected from: Alzheimer's disease, stroke, dementia, amyotrophic lateral sclerosis, and human immunodeficiency virus; atherosclerosis; undesired immune cell activity, and inflammation associated with a disorder selected from: osteoarthritis, anaphylaxis, Behcet's disease, graft rejection, vasculitis, gout, spondylitis, viral disease, bacterial disease, lupus, inflammatory bowel disease, autoimmune hepatitis, and type 1 diabetes mellitus; age-related macular degeneration; cough; leukemia; lymphoma; CNS tumors; prostate cancer; Alzheimer's disease; stroke-induced damage; dementia; amyotrophic lateral sclerosis; and Parkinson's disease. The present invention further provides pharmaceutical compositions comprising Compound 1 and the use of the pharmaceutical compositions for the treatment of CB2 receptor-mediated disorders.
BACKGROUND OF THE INVENTION
Cannabinoids are a group of extracellular signaling molecules that are found in both plants and animals. Signals from these molecules are mediated in animals by two G-protein coupled receptors, CB l and CB2. CBl is expressed most abundantly in the neurons of the CNS but is also present at lower concentrations in a variety of peripheral tissues and cells (Matsuda, L. A. et al. (1990) Nature 346:561- 564). In contrast, CB2 is expressed predominantly, although not exclusively, in non-neural tissues, e.g. in hematopoietic cells, endothelial cells, osteoblasts, osteoclasts, the endocrine pancreas, and cancerous cell lines (Munro, S. et al. (1993) Nature 365:61-65; and as reviewed in Pacher, P. et al. (2006) Pharmacol. Rev. 58(3): 389-462). As such, CB 1 is believed to be primarily responsible for mediating the psychotropic effects of cannabinoids on the body, whereas CB2 is believed to be primarily responsible for most of their non-neural effects.
The compound (laS,5a¾-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide
(Compound 1) was shown to be a potent and selective CB2 agonist and demonstrated a robust and sustained analgesic efficacy over 6 hours in the streptozotocin-induced Painful Diabetic Peripheral Neuropathy (PDPN) model and also showed therapeutic efficacy for osteoarthritis pain in the monosodium iodoacetate-induced (MIA) model of osteoarthritis; see PCT application
PCT/US2010/002360, filed 27 August 2010 (International Publication Number WO2011/025541), and PCT application PCT/US2012/026506, filed 24 February 2012 (International Publication Number WO2012/116276), both are incorporated by reference in their entirety.
In view of the growing demand for compounds useful in the treatment of CB2 receptor- mediated disorders, the compound (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide
(Compound 1) of Formula (I) has emerge new compound:
Figure imgf000003_0001
Accordingly, new and efficient routes are needed for the preparation of (lalS',5alS')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide of Formula (I) (Compound 1), intermediates, and crystalline forms related thereto. Several improvements have now been discovered. These
improvements are described herein.
SUMMARY OF THE INVENTION
The present invention provides, inter alia, processes for the preparation (lalS',5alSr)-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide of Formula (I) (Compound 1) and intermediates thereof.
The processes and intermediates of the present invention are useful in preparing (laS,5aS)-2-(4 oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((5)- l- hydroxymethyl-2,2-dimethyl-propyl)-amide, crystalline forms, and pharmaceutical compositions that are useful in the treatment of CB2 receptor-mediated disorders.
One aspect of the present invention, inter alia, pertains to processes for preparing
Figure imgf000004_0001
(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-a (I) :
Figure imgf000004_0002
comprising the steps of:
a) reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (lie):
Figure imgf000004_0003
(He) .
with dimethyl oxalate in the presence of sodium methoxide and methanol to form one or more intermediates;
b) condensing one or more intermediates with 2-hydrazinyl pyrazine (lid)
,NHNH2
N
(lid)
or an acid salt thereof; in the presence of a condensing-step solvent and optionally a condensing- step acid, provided that at least one of the acid salt or the condensing-step acid is present, to form (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000004_0004
c) hydrolyzing the compound of Formula (He) in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form sodium (4alS',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000005_0001
d) oxidizing the compound of Formula (Ilf) in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form 3-((4alS',5alSr)-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000005_0002
e) coupling the compound of Formula (Ilg) with (S)-feri-leucinol of Formula (Ilh) or a salt thereof:
Figure imgf000005_0003
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
One aspect of the present invention pertains to one or more of the processes as described above in Steps a) through e), either provided separately or together, that are useful in the preparation of an intermediate for use directly or indirectly in the preparation of (lalS',5aS')-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl-2,2- dimethyl-propyl) -amide of Formula (I), salts, and/or crystalline forms thereof.
One aspect of the present invention pertains to processes for preparing (lalS',5alSr)-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide of Formula (I) further comprising the step of formulating (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition.
One aspect of the present invention pertains to one or more of the intermediates described herein. In some embodiments, the intermediate is (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He) and crystalline forms thereof. In some embodiments, the intermediate is sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf) and crystalline forms thereof. Each of the intermediates is useful in the preparation of Compound 1.
These and other aspects of the invention disclosed herein will be set forth in greater detail as the patent disclosure proceeds.
BRIEF DESCRIPTION OF THE DRAWINGS
Figure 1 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the crystalline form of the intermediate (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He).
Figure 2 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the crystalline form of the intermediate sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf)- Figure 3 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the crystalline form of 3-((4alS',5alSr)-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg).
Figure 4 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the anhydrous crystalline form of Compound 1 as described in Example 4, PCT WO2012/ 116276.
Figure 5 shows a differential scanning calorimetry (DSC) thermogram and a
thermogravimetric analysis (TGA) thermogram for a sample containing the anhydrous crystalline form of Compound 1 as described in Example 4, PCT WO2012/116276.
Figure 6 shows an adsorption and desorption isotherm, Dynamic Moisture Sorption (DMS), for a sample containing the anhydrous crystalline form of Compound 1 as described in Example 4, PCT WO2012/116276.
Figure 7 shows a powder X-ray diffraction (PXRD) pattern for a sample containing the anhydrous crystalline form of Compound 1 obtained from the process described in Example 6 herein.
Figure 8 shows the effect of Compound 1 in the monosodium iodoacetate (MIA) model of osteoarthritis in rats, see Example 11.
DETAILED DESCRIPTION OF THE INVENTION
DEFINITIONS
For clarity and consistency, the following definitions will be used throughout this patent document.
The term "agonist" refers to a moiety that interacts with and activates a G-protein-coupled receptor, for instance a cannabinoid receptor, and can thereby initiate a physiological or
pharmacological response characteristic of that receptor. For example, an agonist may activate an intracellular response upon binding to a receptor, or enhance GTP binding to a membrane. The term "Ci-Ce alkyl" refers to a straight or branched carbon radical containing 1 to 6 carbons. Some embodiments are 1 to 5 carbons, some embodiments are 1 to 4 carbons, some embodiments are 1 to 3 carbons and some embodiments are 1 or 2 carbons. Examples of an alkyl include, but are not limited to, methyl, ethyl, w-propyl, isopropyl, w-butyl, sec-butyl, isobutyl, ieri-butyl, pentyl, isopentyl, ieri-pentyl, weo-pentyl, 1-methylbutyl [i.e., -CH(CH3)CH2CH2CH3], 2-methylbutyl [i.e., -CH2CH(CH3)CH2CH3], w-hexyl and the like.
The term "in need of treatment" and the term "in need thereof" when referring to treatment are used interchangeably to mean a judgment made by a caregiver (e.g. physician, nurse, nurse practitioner, etc. in the case of humans; veterinarian in the case of animals, including non-human mammals) that an individual or animal requires or will benefit from treatment. This judgment is made based on a variety of factors that are in the realm of a caregiver's expertise, but that includes the knowledge that the individual or animal is ill, or will become ill, as the result of a disease, condition or disorder that is treatable by the compounds of the invention. Accordingly, the compounds of the invention can be used in a protective or preventive manner; or compounds of the invention can be used to alleviate, inhibit or ameliorate the disease, condition or disorder.
The term "individual" refers to any animal, including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, swine, cattle, sheep, horses, or primates, and most preferably humans.
The term "modulate or modulating" refers to an increase or decrease in the amount, quality, response or effect of a particular activity, function or molecule.
The term "composition" refers to a compound or crystalline form thereof, including but not limited to, salts, solvates, and hydrates of a compound of the present invention, in combination with at least one additional component, such as, a composition obtained/prepared during synthesis, preformulation, in-process testing (i.e., TLC, HPLC, NMR samples), and the like
The term "pharmaceutical composition" refers to a specific composition comprising at least one active ingredient; including but not limited to, salts, solvates, and hydrates of compounds of the present invention, whereby the composition is amenable to investigation for a specified, efficacious outcome in a mammal (for example, without limitation, a human). Those of ordinary skill in the art will understand and appreciate the techniques appropriate for determining whether an active ingredient has a desired efficacious outcome based upon the needs of the artisan.
The phrase "pharmaceutically acceptable salts, solvates, and hydrates" when referring to a compound/compounds as described herein embraces pharmaceutically acceptable solvates and/or hydrates of the compound/compounds, pharmaceutically acceptable salts of the compound/compounds, as well as pharmaceutically acceptable solvates and/or hydrates of pharmaceutically acceptable salts of the compound/compounds. It is also understood that when the phrase "pharmaceutically acceptable solvates and hydrates" or the phrase "pharmaceutically acceptable solvate or hydrate" is used when referring to a compound/compounds as described herein that are salts, it embraces
pharmaceutically acceptable solvates and/or hydrates of such salts. It is also understood by a person of ordinary skill in the art that hydrates are a subgenus of solvates. The term "therapeutically effective amount" refers to the amount of active compound or pharmaceutical agent that elicits the biological or medicinal response in a tissue, system, animal, individual or human that is being sought by a researcher, veterinarian, medical doctor or other clinician or caregiver or by an individual, which includes one or more of the following:
(1) preventing the disease, for example, preventing a disease, condition or disorder in an individual that may be predisposed to the disease, condition or disorder but does not yet experience or display the pathology or symptomatology of the disease;
(2) inhibiting the disease, for example, inhibiting a disease, condition or disorder in an individual that is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., arresting further development of the pathology and/or symptomatology); and
(3) Ameliorating the disease, for example, ameliorating a disease, condition or disorder in an individual that is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., reversing the pathology and/or symptomatology).
The term "as determined by GC" refers to the use of gas chromatography (i.e., GC), including headspace GC, to characterize a sample using a method as described herein, such as, any one of the methods described in Example 8 and Example 17 or an equivalent method thereof. For example, the phrase "a compound is present in the composition of less than or equal to 494 ppm as determined by GC" refers to using GC, such as headspace GC, and a method as described herein to determine that acetone is present in a quantity of less than or equal to 494 ppm.
The term "as determined by HPLC" refers to the use of high-performance liquid
chromatography (HPLC) (i.e., also referred to as high-pressure liquid chromatography), to characterize a sample using a method described herein, such as, the method described in Example 9 or an equivalent method thereof. For example, the phrase "a compound is present in the composition of less than or equal to 1.0% as determined by HPLC" refers to the use of HPLC and a method as described herein to determine that the compound is present in a quantity of less than or equal to 1.0%.
The term "Ci-Ce alkyl phosphonic anhydride" refers to the following formula wherein each C1-C6 alkyl group can be the same or different and has the same definition as described herein:
Figure imgf000008_0001
C C6 alkyl O
In some embodiments, each Ci-C6 alkyl group is the same. In some embodiments, each Ci-C6 alkyl group is w-propyl (i.e., 1-propanephosphonic anhydride).
It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment.
Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination. In addition, subcombinations of uses and medical indications listed in the embodiments describing such uses and medical indications described herein, are also specifically embraced by the present invention just as if each and every subcombination of uses and medical indications was individually and explicitly recited herein. CRYSTALLINE FORMS
The crystalline forms of (laS',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide and intermediates related thereto can be identified by their unique solid state signatures with respect to, for example, differential scanning calorimetry (DSC), powder X-ray diffraction (PXRD), and other solid state methods. Further characterization with respect to water or solvent content of the crystalline forms can be gauged by any of the following methods for example, thermogravimetric analysis (TGA), DSC and the like. For DSC, it is known that the temperatures observed for thermal events will depend upon sample purity, and may also depend on the rate of temperature change, as well as sample preparation technique and the particular instrument employed. Thus, the values reported herein relating to DSC thermograms can vary by plus or minus about 5°C (i.e., ± about 5°C). The values reported herein relating to DSC thermograms can also vary by plus or minus about 20 joules per gram (i.e., ± about 20 joules per gram). For PXRD, the relative intensities of the peaks can vary, depending upon the sample preparation technique, the sample mounting procedure and the particular instrument employed.
Moreover, instrument variation and other factors can often affect the 2Θ values. Therefore, the peak assignments of diffraction patterns can vary by plus or minus about 0.2° (i.e., ± about 0.2°). For TGA, the temperature features reported herein can vary by plus or minus about 5°C (i.e., ± about 5°C). The TGA % weight changes reported herein over a specified temperature range can vary by plus or minus about 2% weight change (i.e., ± about 2% weight change) due to, for example, variations in sample quality and sample size. Further characterization with respect to hygroscopicity of the crystalline form can be gauged by, for example, dynamic moisture sorption (DMS). The DMS features reported herein can vary by plus or minus about 5% relative humidity (i.e., ± about 5% relative humidity). The DMS features reported herein can also vary by plus or minus about 2% weight change (i.e., ± about 2% weight change).
A. Compound 1 (Crystalline Anhydrous Form).
One aspect of the present invention relates to novel processes for the preparation of an anhydrous form of (lalS',5aSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide
(Compound 1). The anhydrous form was previously described in International Publication Number WO2012/1 16276 and the representative physical properties are shown below in Table 1 and Table 2.
The physical properties of the crystalline form of Compound 1 (anhydrous form) are summarized in Table 1 below. Table 1
Figure imgf000010_0001
Certain other PXRD peaks for the anhydrous form of (lalS',5aS')-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl-2,2- dimethyl-propyl) -amide (Compound 1) are shown in Table 2 below.
Table 2
Figure imgf000010_0002
One aspect of the present invention relates to an anhydrous crystalline form of (laS,5a5)-2-(4- oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l- hydroxymethyl-2,2-dimethyl-propyl)-amide.
One aspect of the present invention relates to an anhydrous crystalline form of (laS,5a5)-2-(4- oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l- hydroxymethyl-2,2-dimethyl-propyl)-amide prepared according to any one of the processes described herein.
One aspect of the present invention relates to an anhydrous crystalline form of (laS,5a5)-2-(4- oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide, wherein the anhydrous crystalline form has a powder X- ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, and 10.7° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, and 16.9° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, and 1 1.1° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2(9, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, and 17.4° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, and 16.5° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, 16.5° + 0.2°, 14.5° + 0.2°, 11.8° + 0.2°, and 18.9° + 0.2°. In some embodiments, the anhydrous crystalline form has a powder X-ray diffraction pattern substantially as shown in Figure 4, wherein by "substantially" is meant that the reported peaks can vary by about + 0.2 °2 Θ.
In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 159.6°C and about 169.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 160.6°C and about 168.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 162.6°C and about 166.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 163.6°C and about 165.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature at about 164.6°C. In some embodiments, the anhydrous crystalline form has a differential scanning calorimetry thermogram substantially as shown in Figure 5, wherein by "substantially" is meant that the reported DSC features can vary by about + 5°C and that the reported DSC features can vary by about + 20 joules per gram.
In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile showing about 0.5% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile showing about 0.25% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has a thermogravimetric analysis profile substantially as shown in Figure 5, wherein by "substantially" is meant that the reported TGA features can vary by about + 5°C, and that that the reported TGA features can vary by about + 2% weight change.
In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, and 10.7° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 159.6°C and about 169.6°C; and 3) a thermogravimetric analysis profile showing about 0.5% weight loss below about 135°C. In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, and 16.9° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 160.6°C and about 168.6°C; and 3) a thermogravimetric analysis profile showing about 0.25% weight loss below about 135°C.
In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, and 11.1° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 162.6°C and about 166.6°C; and 3) a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, and 17.4° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature between about 163.6°C and about 165.6°C; and 3) a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° +
0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, and 16.5° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature at about 164.6°C; and 3) a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.5° + 0.2°, 10.7° + 0.2°, 16.9° + 0.2°, 25.4° + 0.2°, 11.1° + 0.2°, 9.8° + 0.2°, 17.4° + 0.2°, 22.1° + 0.2°, 16.5° + 0.2°, 14.5° + 0.2°, 11.8°
+ 0.2°, and 18.9° + 0.2°; 2) a differential scanning calorimetry thermogram comprising an endotherm with an extrapolated onset temperature at about 164.6°C; and 3) a thermogravimetric analysis profile showing about 0.05% weight loss below about 135°C.
In some embodiments, the anhydrous crystalline form has one or more of the following: 1) a powder X-ray diffraction pattern substantially as shown in Figure 4; 2) a differential scanning calorimetry thermogram substantially as shown in Figure 5; and 3) a thermogravimetric analysis profile substantially as shown in Figure 5.
B. (4aS,5aS)-Methyl l-(Pyrazin-2-yl)-4,4a,5,5a-tetrahydro-lf7- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He), Crystalline Form).
One aspect of the present invention relates to a novel crystalline form of (4alS',5aS')-methyl 1-
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of
(Formula (He)). One aspect of the present invention relates to a novel crystalline form of (4aS,5aS)-mefhyl 1- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of (Formula (He)) prepared according to any one of the processes described herein.
Certain PXRD peaks for the crystalline form of (4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate are shown in Table 3, Example 3.
One aspect of the present invention relates to a crystalline form of (4aS,5aS)-mefhyl l-(pyrazin- 2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He)), wherein the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2° and 11.0° + 0.2°. In some embodiments, the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, and 17.9° + 0.2°. In some embodiments, the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, and 17.9° + 0.2°. In some embodiments, the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.9° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form of Formula (He) has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.3° + 0.2°, 17.9° + 0.2°, 21.0° + 0.2°, 24.3° + 0.2°, 25.3° + 0.2°, and 25.9° + 0.2°. In some embodiments, the crystalline form of Formula (He) has a powder X-ray diffraction pattern substantially as shown in Figure 1, wherein by "substantially" is meant that the reported peaks can vary by about + 0.2 °2Θ.
C. Sodium (4a5,5aS)-l-(Pyrazin-2-yl)-4,4a,5,5a-tetrahydro-li7- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (Ilf), Crystalline Form).
One aspect of the present invention relates to a novel crystalline form of sodium (4aS,5a5)-l- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (Ilf)).
One aspect of the present invention relates to a novel crystalline form of sodium
Figure imgf000013_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (Ilf)) prepared according to any one of the processes described herein.
Certain PXRD peaks for the crystalline form of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate are shown in Table 4, Example 4.
One aspect of the present invention relates to a crystalline form of sodium
Figure imgf000013_0002
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (Ilf)), wherein the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2° and 25.3° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 2 Θ, at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 1Θ, at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 2(9, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern comprising peaks, in terms of 2(9, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 11.5° + 0.2°, 12.3° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°. In some embodiments, the crystalline form of Formula (Ilf) has a powder X-ray diffraction pattern substantially as shown in Figure 2, wherein by "substantially" is meant that the reported peaks can vary by about + 0.2 °2Θ.
D. 3-((4a5,5a5)-3-carboxy-4,4a,5,5a-tetrahydro-l/ -cyclopropa[4,5]cyclopenta[l,2- c]pyrazol-l-yl)pyrazine 1-oxide (Formula (Ilg), Crystalline Form).
3-((4alS',5alSr)-3-Carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l- yl)pyrazine 1-oxide of Formula (Ilg) has also been referred to as (laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a- tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid N-oxide in WO2011/025541 (e.g., Example 1.108) and WO2012/116276 (e.g., Example 1, METHOD 2, Step E), it is understood that both chemical names refer to the same compound with the following chemical structure:
Figure imgf000014_0001
One aspect of the present invention relates to a novel crystalline form of
Figure imgf000014_0002
carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l-yl)pyrazine 1-oxide (Formula (Ilg)).
One aspect of the present invention relates to a novel crystalline form of
Figure imgf000014_0003
carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l-yl)pyrazine 1-oxide (Formula (Ilg)) prepared according to any one of the processes described herein.
Certain PXRD peaks for the crystalline form of 3-((4alS',5aS')-3-carboxy-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide are shown in Table 5, Example 5.
One aspect of the present invention relates to a crystalline form of 3-((4alS',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Formula (Ilg)), wherein the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 1Θ, at 12.6° + 0.2° and 16.8° + 0.2°. In some embodiments, the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 1Θ, at 12.6° + 0.2°, 16.8° + 0.2°, and 27.1° + 0.2°. In some embodiments, the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 12.6° + 0.2°, 16.8° + 0.2°, 22.0° + 0.2°, 26.5° + 0.2°, and 27.1° + 0.2°. In some embodiments, the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 12.6° + 0.2°, 16.8° + 0.2°, 21.6° + 0.2°, 22.0° + 0.2°, 22.6° + 0.2°, 26.5° + 0.2°, and 27.1° + 0.2°. In some embodiments, the crystalline form of
Formula (Ilg) has a powder X-ray diffraction pattern comprising peaks, in terms of 1Θ, at 12.6° + 0.2°, 12.9° + 0.2°, 16.8° + 0.2°, 21.0° + 0.2°, 21.6° + 0.2°, 22.0° + 0.2°, 22.2° + 0.2°, 22.6° + 0.2°, 26.5° + 0.2°, 27.1° + 0.2°, and 27.6° + 0.2°. In some embodiments, the crystalline form of Formula (Ilg) has a powder X-ray diffraction pattern substantially as shown in Figure 3, wherein by "substantially" is meant that the reported peaks can vary by about + 0.2 °2Θ.
The crystalline forms described herein can be prepared by any of the suitable procedures known in the art for preparing crystalline polymorphs. In some embodiments the crystalline forms described herein are prepared according to the Examples. In some embodiments, the crystalline forms described herein can be prepared by heating crystalline forms other than the crystalline forms described herein. In some embodiments, the crystalline forms described herein can be prepared by recrystahizing crystalline forms other than the crystalline forms described herein.
CERTAIN PROCESSES OF THE INVENTION
The present invention is directed, inter alia, to processes useful in the preparation of (laS^aS")- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimethyl-propyl)-amide and crystalline forms thereof.
The processes described herein can be monitored according to any suitable method known in the art. For example, product formation can be monitored by spectroscopic means, such as nuclear magnetic resonance spectroscopy {e.g., ¾ or 13C), infrared spectroscopy, spectrophotometry {e.g., UV- visible), mass spectrometry, gas chromatography (GC), or by chromatography, such as, high performance liquid chromatography (HPLC) or thin layer chromatography.
The processes described herein can be carried out in air or under an inert atmosphere unless specifically stated otherwise. Typically, reactions containing reagents or products that are substantially reactive with air can be carried out using air-sensitive synthetic techniques that are well known to one skilled in the art.
Upon carrying out preparation of compounds according to the processes described herein, the usual isolation and purification operations such as concentration, filtration, extraction, solid-phase extraction, recrystallization, enantiomeric-enrichment via recrystallization, chromatography, and the like may be used, to isolate the desired product.
Example processes and certain intermediates of the present invention are shown in Scheme I to
Scheme VIII below.
Representative cyclization step between the intermediate of Formula (Ha) with a cyclization- step base {e.g., w-butyl lithium) and optionally a 2° amine {e.g., tetramethylpiperidine (TMP)) in the presence of a cyclization-step solvent (e.g., methyl ieri-butyl ether (MTBE)) is provided below in
Scheme I.
Scheme I
cyclization-step solvent
cyclization-step base ^
optionally a 2° amine
Figure imgf000016_0001
H
(lla) (lib)
Representative ketone forming step utilizing the intermediate of Formula (lib) in the presence of an oxidizing agent (e.g., TEMPO/NaOCl), ketone-forming solvent (e.g., MTBE), and a ketone- forming aqueous buffer (e.g., potassium phosphate buffer) is provided below in Scheme II.
Scheme II
oxidizing agent
ketone-forming solvent
Figure imgf000016_0002
H ketone-forming
(lib) aqueous buffer (lie)
Representative reacting step between the intermediate of Formula (lie) and dimethyl oxalate in the presence of sodium methoxide and methanol is provided below in Scheme III.
Figure imgf000016_0003
It is appreciated that the sodium salt intermediates (i.e., sodium (£')-2-methoxy-2-oxo-l- ((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-ylidene)efhanolate of Formula (IIc-i) and sodium (Z)-2-methoxy- 2-oxo-l-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of Formula (IIc-ii)) depicted inside the brackets above can be used directly in the next step (i.e., condensing-step), or can be isolated, or the sodium salt intermediates could be subsequently neutralized to provide the diketo-ester intermediate (i.e., methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii)) as represented below:
Figure imgf000016_0004
Representative condensing step between one or more of the intermediates of Formulae (IIc-i), (IIc-ii), and (IIc-iii) with the intermediate of Formula (lid) in the presence of a condensing-step solvent is shown below in Scheme IV. It is appreciated that the condensing-step is conducted in the presence of an acid, such as, an acid salt of 2-hydrazinyl pyrazine (lid), or a condensing-step acid, or mixture thereof. Scheme IV
(IIc-i) and/or (Ilc-ii) and/or (Ilc-iii) reof
condensing-step solvent
optionally a
Figure imgf000017_0001
condensing-step acid
Alternatively, the representative reacting step and condensing step can be successively employed in a two-step process using the intermediate of Formula (lie) and dimethyl oxalate in the presence of sodium methoxide and methanol without isolation of the intermediates of Formulae (IIc-i), (Ilc-ii), and/or (Ilc-iii) followed by using the intermediate of Formula (lid) in the presence of a condensing-step solvent as provided below in Scheme V.
Scheme V
Figure imgf000017_0002
optionally a condensing-step acid
Representative hydrolyzing step and the intermediates of Formulae (He) and (Ilf) of the present invention are provided below in Scheme VI.
Figure imgf000017_0003
(He) (Ilf)
Representative oxidizing step between the intermediate of Formulae (Ilf) with hydrogen peroxide in the presence of an oxidizing-step solvent is provided below in Scheme VII.
Figure imgf000017_0004
Representative coupling step between the intermediate of Formulae (Ilg) (or a salt thereof) and the compound of Formula (Ilh) (or a salt thereof) with a coupling-step agent, coupling-step base in the presence of a coupling-step solvent is provided below in Scheme VIII.
Scheme VIII
Figure imgf000018_0001
One aspect of the present invention includes every combination of one or more process step and intermediates related thereto used in the preparation of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a- tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl- propyl)-amide of Formula (I) and crystalline forms thereof, such as those processes exemplified by Schemes I, II, III, IV, V, VI, VII, and VIII (supra) and compounds of Formulae (I), (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Ilf), (Hf-i), (Ilg), (Ilh), (Ilia), (Illb), and (IIIc).
One aspect of the present invention pertains to one or more of the intermediates, such as Compounds of Formulae (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Ilf), (Ilf-i), (Ilg), and (Ilh), as exemplified in Schemes I, II, III, IV, V, VI, VII, and VIII (supra), useful in the preparation of the Compound of Formula (I) and crystalline forms thereof.
Certain synthetic processes for the preparation of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl- propyl)-amide of Formula (I) (Compound 1) have been described in PCT application
PCT/US2010/002360, filed 27 August 2010 (International Publication Number WO2011/025541) and PCT application PCT/US2012/026506, filed 24 February 2012 (International Publication Number WO2012/116276); both are incorporated by reference in their entirety. Several improvements have been discovered and are described herein.
One improvement is the use of dimethyl oxalate in the presence of sodium methoxide and methanol as represented in Scheme III, Scheme IV, and Scheme V which was found to unexpectedly improve volume efficiency compared to the previously reported use of diethyl oxalate in absolute ethanol, THF, and potassium ieri-butoxide (see, Example 1.107, Step C, WO2011/025541 and
Example 1, Step C, WO2012/116276). Replacing absolute ethanol in the previously reported method with methanol improved the volume efficiency but provided an undesirable mixture of esters. Using dimethyl oxalate in methanol and replacing THF/potassium ieri-butoxide with sodium methoxide provided unexpected improvements over the previously reported methods, such as those cited above. Further, the Compound of Formulae (He) and the crystalline form of the Compound of Formulae (He) were not previously disclosed. Another improvement is the isolation of the sodium salt of Formula (Ilf) instead of the free acid of Formula (Ilf-i) as previously disclosed (see, Example 1.107, Step D, WO2011/025541 and Example 1, Step D, WO2012/116276).
Figure imgf000019_0001
The previous procedures referred to the free acid as a "fine precipitate" that was isolated by filtration. It was found that the previous procedures gave the free acid of Formula (Ilf-i) as a gelatinous precipitate that was extremely difficult to filter. This was observed even on smaller scaled reactions. The resulting free acid mixture clogged the filtration system resulting in lengthy filtration times. This problem was exacerbated as the scale of the reaction increased. Surprising, it was discovered that the sodium salt of Formula (Ilf) could be isolated and was found to be crystalline and stable leading to improved filtration and isolation thus improving processability of the reaction. Advantageously, the crystal form of the sodium salt of Formula (Ilf) showed improved manufacturing properties compared to the free acid, such as, processing (e.g., crystal size), processability (e.g. improved filterability on scale), and ease of drying while retaining purity and yield. In the previously reported processes (i.e., WO2011/025541 and WO2012/116276) the crystalline form of the sodium salt of Formula (Ilf) was not isolated and was not previously disclosed.
Another improvement relates to the oxidation processes for preparing the Compound of Formula (Ilg). It was previously reported that two separate oxidation procedures were required when using formic acid and hydrogen peroxide to provide the Compound of Formula (Ilg) (see, Example 1.108, WO2011/025541 and Example 1, Method 2, Step E, WO2012/116276). However, it was discovered that when the oxidation was conducted in the presence of trifluoroacetic acid (i.e., TFA) the second separate oxidation step was not necessary and the present oxidation proceeds to completion (i.e., less than 0.6% of non-oxidized free acid, see Example 5).
Another improvement relates to the coupling processes for preparing the Compound of Formula (I) from Compounds of Formula (Ilg) and (Ilh) in the presence of acetonitrile as the coupling- step solvent. It was previously reported that coupling Compounds of Formula (Ilg) and (Ilh) in DMF followed by recrystallization with CH2Cl2/hexanes and a lengthy drying step (i.e., ~65°C for 10 days) provided the Compound of Formula (I) in a 67.2% yield (see, Example 1, Method 2, Step F,
WO2012/116276). However, it has now been discovered that using acetonitrile as the coupling-step solvent and the recrystallization solvent with water gave an average yield of 70.6% for the preparation of the Compound of Formula (I) (the yields for three separate batches were 74.9%, 72.3%, and 70.7%; see Example 6). As disclosed herein, several improvements have been discovered and are useful in the preparation of Compound 1.
Compounds of the invention can also include tautomeric forms, such as keto-enol tautomers and the like. Tautomeric forms can be in equilibrium or sterically locked into one form by appropriate substitution. It is understood that the various tautomeric forms are within the scope of the compounds of the present invention.
Compounds of the invention also include all isotopes of atoms occurring in the intermediates and/or final compounds. Isotopes include those atoms having the same atomic number but different mass numbers. For example, isotopes of hydrogen include deuterium and tritium.
I. Processes useful in the preparation of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro-l//-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2- dimethyl-propyl)-amide of Formula (I).
Ia. Processes for preparing the Compound 1 of Formula (I)
One aspect of the present invention relates to processes comprising the following steps, (i.e. Steps a) to Steps e)), for preparing (laS,5a¾-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000020_0001
comprising the steps of:
a) reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (lie):
Figure imgf000020_0002
(He) .
with dimethyl oxalate in the presence of sodium methoxide and methanol to form one or more intermediates;
b) condensing one or more intermediates with 2-hydrazinyl pyrazine (lid):
NHNH2
N
(lid)
or an acid salt thereof; in the presence of a condensing-step solvent and optionally a condensing-step acid, provided that at least one of the acid salt or the condensing-step acid is present, to form (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate of Formula (He):
Figure imgf000021_0001
c) hydrolyzing the compound of Formula (He) in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000021_0002
d) oxidizing the compound of Formula (Ilf) in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form 3-((4alS',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000021_0003
e) coupling the compound of Formula (Ilg) with (S)-ieri-leucinol of Formula (Ilh) or a salt thereof:
Figure imgf000021_0004
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I). Reacting Step Using (lS,5/f)-Bicyclo[3.1.0]hexan-2-one
In some embodiments, reacting in Step a) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, reacting in Step a) is carried out in the presence of a nitrogen atmosphere.
In some embodiments, reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first- reacting mixture. In some embodiments, the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the first-reacting mixture is at a temperature of about 20°C to about 25°C.
In some embodiments, reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture.
In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 25°C to about 50°C. In some embodiments, wherein after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 30°C to about 45°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 35°C to about 40°C.
In some embodiments, sodium methoxide is in a solution comprising methanol. In some embodiments, sodium methoxide is about 5 weight percent to about 30 weight percent in methanol. In some embodiments, sodium methoxide is about 25 weight percent in methanol solution.
In some embodiments, dimethyl oxalate and sodium methoxide are present in molar excess compared to (lS,5R)-bicyclo[3.1.0]hexan-2-one.
In some embodiments, sodium methoxide is present in molar excess compared to dimethyl oxalate.
In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1 : 1.05: 1.05 to about 1 :2.4:2.8. In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1 : 1.2: 1.4.
In some embodiments, the intermediate is sodium (£)-2-methoxy-2-oxo-l-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of the formula:
Figure imgf000023_0001
or sodium (Z)-2-methoxy-2-oxo-l-((llS',5lS')-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of the formula:
Figure imgf000023_0002
or a mixture thereof.
In some embodiments, the intermediate is not isolated.
In some embodiments, the intermediate is isolated.
In some embodiments, the intermediate is isolated and is methyl 2-oxo-2-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (Ilc-i):
Figure imgf000023_0003
or an enol form thereof, or a mixture of methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3- yl)acetate and the enol form thereof. It is understood that at least one enol form may exist.
Condensing Step
In some embodiments, condensing in Step b) further comprises the step of mixing the intermediate and the condensing-step solvent together to form a first-condensing mixture.
In some embodiments, the condensing-step solvent comprises methanol.
In some embodiments, the condensing is carried out in the presence of the condensing-step acid.
In some embodiments, the condensing-step acid is HCl. In some embodiments, the condensing- step acid is aqueous HCl. In some embodiments, the condensing-step acid is about IN aqueous HCl to about 12N aqueous HCl. In some embodiments, the condensing-step acid is about 3N aqueous HCl to about ION aqueous HCl. In some embodiments, the condensing-step acid is about 5N aqueous HCl to about 7N aqueous HCl. In some embodiments, the condensing-step acid is about 6N aqueous HCl.
In some embodiments, condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture.
In some embodiments, the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing- step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about
20°C to about 50°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
In some embodiments, condensing in Step b) further comprises the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-condensing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, ^aS^aS^-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 90% or greater as determined by HPLC. In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
Hydrolyzing Step
In some embodiments, hydrolyzing in Step c) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, hydrolyzing in Step c) is carried out in the presence of a nitrogen atmosphere.
In some embodiments, hydrolyzing in Step c) further comprising the step of mixing (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the hydrolyzing- step solvent together to form a first-hydrolyzing mixture.
In some embodiments, the hydrolyzing-step solvent comprises methanol.
In some embodiments, the first-hydrolyzing mixture is at a temperature of about 20°C to about
45°C.
In some embodiments, hydrolyzing in Step c) further comprises the step of adding the sodium hydroxide to the first-hydrolyzing mixture to form a second-hydrolyzing mixture.
In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 25°C to about 60°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 35°C to about 55°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 40°C to about 50°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 45°C.
In some embodiments, hydrolyzing in Step c) further comprises the step of isolating sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-hydrolyzing mixture. In some embodiments, isolating is by filtration.
In some embodiments, sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline. In some embodiments, crystalline sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
Oxidizing Step
In some embodiments, oxidizing in Step d) further comprises the step of mixing sodium
(4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the oxidizing-step solvent together to form a first-oxidizing mixture.
In some embodiments, the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to
1.00:0.71 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2. In some embodiments, the molar ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5. In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000026_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000026_0002
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.94:3.06 to about 1.00:3.06:2.94 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
In some embodiments, the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C.
In some embodiments, oxidizing in Step d) further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
In some embodiments, oxidizing in Step d) further comprises the step of isolating 3-((4aS,5aS)- 3-carboxy-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5] cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide from the second-oxidizing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline. Coupling Step
In some embodiments, the coupling in Step e) further comprises the step of mixing 3- ((4alS',5alS')-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, the coupling-step base, and the coupling-step solvent together to form a first-coupling mixture.
In some embodiments, the first-coupling mixture is at a temperature of about -15°C to about
25°C. In some embodiments, the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
In some embodiments, the coupling in Step e) further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
In some embodiments, after addition of the coupling-step agent the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about
-5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C.
In some embodiments, the coupling in Step e) further comprises the step of adding (S)-tert- leucinol to the second-coupling mixture.
In some embodiments, (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
In some embodiments, (S)-ieri-leucinol is added substantially all at once. It was found that adding (S)-ieri-leucinol all at once instead of over a longer period of time minimized the amount of a side product.
In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent.
In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro-l/i-benzotriazole-l-yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and A^A^A^-tetramefhyl-CKS^-dihydro- 4-oxo-l, 2, 3-benzotriazin-3-yl)uranium tetrafluoroborate (TDBTU). In some embodiments, the coupling-step agent is 2-(7-aza-l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium
hexafluorophosphate (HATU).
In some embodiments, the coupling-step agent is a phosphonic acid anhydride. In some embodiments, the phosphonic acid anhydride is a Ci-C6 alkyl phosphonic anhydride. In some embodiments, the phosphonic acid anhydride is 1-propanephosphonic anhydride.
In some embodiments, the coupling-step base comprises a tertiary amine. In some
embodiments, the coupling-step base is selected from: A^N-diisopropylefhylamine (DIEA), triethylamine (TEA), N-mefhylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimethylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4- (dimethylamino)pyridine (DBDMAP). In some embodiments, the coupling-step base is triethylamine.
In some embodiments, the coupling-step solvent is acetonitrile.
In some embodiments, the coupling in Step e) further comprises the step of isolating (la^SaS")- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture. In some embodiments, the isolating is by filtration.
In some embodiments, (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
In some embodiments, the coupling in Step e) further comprises the step of formulating (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition. In some embodiments, the formulating comprises admixing (lalS',5alS')-2-(4-oxy-pyrazin-2- yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl- 2,2-dimethyl-propyl)-amide with a pharmaceutical excipient. In some embodiments, the formulating is admixing (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene- 4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
Ib. Processes for preparing the Compound 1 of Formula (I)
One aspect of the present invention relates to processes comprising the following steps, i.e. Steps a) to Steps c), for preparing (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000028_0001
comprising the steps of:
a) hydrolyzing (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydi
cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000029_0001
(He)
in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form sodium
Figure imgf000029_0002
l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000029_0003
(Ilf)
b) oxidizing the compound of Formula (Ilf) in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form 3-((4alS',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000029_0004
c) coupling the compound of Formula (Ilg) with (S)-ieri-leucinol of Formula (Ilh):
Figure imgf000029_0005
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
Hydrolyzing Step
In some embodiments, hydrolyzing in Step a) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, hydrolyzing in Step a) is carried out in the presence of a nitrogen atmosphere.
In some embodiments, hydrolyzing in Step a) further comprises the step of mixing (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the hydrolyzing- step solvent together to form a first-hydrolyzing mixture.
In some embodiments, the hydrolyzing-step solvent comprises methanol.
In some embodiments, the first-hydrolyzing mixture is at a temperature of about 20°C to about
45°C.
In some embodiments, hydrolyzing in Step a) further comprises the step of adding the sodium hydroxide to the first-hydrolyzing mixture to form a second-hydrolyzing mixture.
In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 25°C to about 60°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 35°C to about 55°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 40°C to about 50°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 45°C.
In some embodiments, hydrolyzing in Step a) further comprises the step of isolating sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-hydrolyzing mixture. In some embodiments, isolating is by filtration.
In some embodiments, sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
In some embodiments, crystalline sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
Oxidizing Step
In some embodiments, oxidizing in Step b) further comprises the step of mixing sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the oxidizing-step solvent together to form a first-oxidizing mixture.
In some embodiments, the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to 1.00:0.71 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2. In some embodiments, the molar ratio of sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5. In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
In some embodiments, the ratio of sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000031_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000031_0002
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.94:3.06 to about 1.00:3.06:2.94 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
In some embodiments, the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C.
In some embodiments, oxidizing in Step b) further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions.
In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
In some embodiments, oxidizing in Step b) further comprises the step of isolating 3-((4aS,5aS)- 3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide from the second-oxidizing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, 3-((4alS',5alSr)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
Coupling Step
In some embodiments, the coupling in Step c) further comprises the step of mixing 3- ((4alS',5alS')-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, the coupling-step base, and the coupling-step solvent together to form a first-coupling mixture.
In some embodiments, the first-coupling mixture is at a temperature of about -15°C to about 25°C. In some embodiments, the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
In some embodiments, the coupling in Step c) further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
In some embodiments, after addition of the coupling-step agent the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about -5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C. In some embodiments, the coupling in Step c) further comprises the step of adding (S)-tert- leucinol to the second-coupling mixture.
In some embodiments, (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
In some embodiments, (S)-ieri-leucinol is added substantially all at once.
In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent. In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro-l/i-benzotriazole-l-yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and A^A^A^-tetramefhyl-CKS^-dihydro- 4-oxo-l, 2,3-benzotriazin-3-yl)uranium tetrafluoroborate (TDBTU). In some embodiments, the coupling-step agent is 2-(7-aza-l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium
hexafluorophosphate (HATU).
In some embodiments, the coupling-step agent is a phosphonic acid anhydride. In some embodiments, the phosphonic acid anhydride is a Ci-C6 alkyl phosphonic anhydride. In some embodiments, the phosphonic acid anhydride is 1-propanephosphonic anhydride.
In some embodiments, the coupling-step base comprises a tertiary amine. In some
embodiments, the coupling-step base is selected from: A^N-diisopropylefhylamine (DIEA), triefhylamine (TEA), N-mefhylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimefhylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4- (dimethylamino)pyridine (DBDMAP). In some embodiments, the coupling-step base is triethylamine.
In some embodiments, the coupling-step solvent is acetonitrile.
In some embodiments, the coupling in Step c) further comprises the step of isolating (laS,5aS)- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture. In some embodiments, the isolating is by filtration.
In some embodiments, (lalS',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
In some embodiments, the coupling in Step c) further comprises the step of formulating (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition. In some embodiments, the formulating comprises admixing (laS,5aS)-2-(4-oxy-pyrazin-2- yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl- 2,2-dimethyl-propyl)-amide with a pharmaceutical excipient. In some embodiments, the formulating is admixing (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene- 4-carboxylic acid ((S)-l -hydro xymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient. Ic. Processes for preparing the Compound 1 of Formula (I)
One aspect of the present invention relates to processes comprising the following Steps a) and Steps b) for preparing (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000034_0001
comprising the steps of:
a) oxidizing sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydi
cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000034_0002
(Ilf)
in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form 3-((4alS',5alS')-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000034_0003
b) coupling the compound of Formula (Ilg) with (S)-ieri-leucinol of Formula (Ilh):
Figure imgf000034_0004
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
Oxidizing Step
In some embodiments, oxidizing in Step a) further comprises the step of mixing sodium
(4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate and the oxidizing-step solvent together to form a first-oxidizing mixture.
In some embodiments, the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to
1.00:0.71 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2. In some embodiments, the molar ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5. In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000036_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000036_0002
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.94:3.06 to about 1.00:3.06:2.94 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
In some embodiments, the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C.
In some embodiments, oxidizing in Step a) further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
In some embodiments, oxidizing in Step a) further comprises the step of isolating 3-((4aS,5aS)- 3-carboxy-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5] cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide from the second-oxidizing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
Coupling Step
In some embodiments, the coupling in Step b) further comprises the step of mixing 3- ((4alS',5alS')-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, the coupling-step base, and the coupling-step solvent together to form a first-coupling mixture. In some embodiments, the first-coupling mixture is at a temperature of about -15°C to about 25°C. In some embodiments, the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
In some embodiments, the coupling in Step b) further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
In some embodiments, after addition of the coupling-step agent the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about -5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C.
In some embodiments, the coupling in Step b) further comprises the step of adding (S)-tert- leucinol to the second-coupling mixture.
In some embodiments, (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
In some embodiments, (S)-ieri-leucinol is added substantially all at once.
In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent.
In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- 1-yl)- 1 , 1 ,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro- lH-benzotriazole- 1 -yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and N,N,N,N etramethyl-0-(3,4-dihydro- 4-oxo-l, 2,3-benzotriazin-3-yl)uranium tetrafluoroborate (TDBTU). In some embodiments, the coupling-step agent is 2-(7-aza-l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium
hexafluorophosphate (HATU).
In some embodiments, the coupling-step agent is a phosphonic acid anhydride. In some embodiments, the phosphonic acid anhydride is a Ci-C6 alkyl phosphonic anhydride. In some embodiments, the phosphonic acid anhydride is 1-propanephosphonic anhydride.
In some embodiments, the coupling-step base comprises a tertiary amine. In some
embodiments, the coupling-step base is selected from: A^N-diisopropylefhylamine (DIEA), triethylamine (TEA), N-methylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimethylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4-
(dimethylamino)pyridine (DBDMAP). In some embodiments, the coupling-step base is triethylamine.
In some embodiments, the coupling-step solvent is acetonitrile. In some embodiments, the coupling in Step b) further comprises the step of isolating (la^^a^- 2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture. In some embodiments, the isolating is by filtration.
In some embodiments, (lalS',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
In some embodiments, the coupling in Step b) further comprises the step of formulating (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition. In some embodiments, the formulating comprises admixing (lalS',5alS')-2-(4-oxy-pyrazin-2- yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymefhyl- 2,2-dimethyl-propyl)-amide with a pharmaceutical excipient. In some embodiments, the formulating is admixing (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene- 4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
Id. Processes for preparing the Compound 1 of Formula (I)
One aspect of the present invention relates to processes for preparing (lalS',5alS')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)- (I):
Figure imgf000038_0001
comprising the step:
coupling 3-((4alS',5aS')-3-carboxy-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilg):
Figure imgf000038_0002
with (S)-ieri-leucinol of Formula (Ilh):
Figure imgf000039_0001
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
In some embodiments, coupling further comprises the step of mixing 3-((4alS',5alSr)-3-carboxy-
4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide, the coupling- step base, and the coupling-step solvent together to form a first-coupling mixture.
In some embodiments, the first-coupling mixture is at a temperature of about -15°C to about 25°C. In some embodiments, the first-coupling mixture is at a temperature of about -10°C to about 15°C. In some embodiments, the first-coupling mixture is at a temperature of about -5°C to about 10°C. In some embodiments, the first-coupling mixture is at a temperature of about 0°C to about 5°C.
In some embodiments, coupling further comprises the step of adding the coupling-step agent to the first-coupling mixture to form a second-coupling mixture.
In some embodiments, after addition of the coupling-step agent the temperature of the second- coupling mixture is maintained at a temperature of about -15°C to about 25°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about -10°C to about 15°C. In some embodiments, after addition of the coupling- step agent the temperature of the second-coupling mixture is maintained at a temperature of about -5°C to about 10°C. In some embodiments, after addition of the coupling-step agent the temperature of the second-coupling mixture is maintained at a temperature of about 0°C to about 5°C.
In some embodiments, coupling further comprises the step of adding (S)-ieri-leucinol to the second-coupling mixture.
In some embodiments, (S)-ieri-leucinol is added to the second-coupling mixture as a solution comprising acetonitrile.
In some embodiments, (S)-ieri-leucinol is added substantially all at once.
In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent. In some embodiments, the coupling-step agent is a tetramethyluronium-based coupling agent selected from: 2-(l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HBTU), 2-(lH- benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium tetrafluoroborate (TBTU), 2-(7-aza- 1/i-benzotriazole- l-yl)-l, 1,3,3-tetramethyluronium hexafluorophosphate (HATU), 2-(6-chloro-l/i-benzotriazole-l-yl)- 1, 1,3,3-tetramethylaminium hexafluorophosphate (HCTU), and A^A^A^-tetramefhyl-CKS^-dihydro- 4-oxo-l, 2,3-benzotriazin-3-yl)uranium tetrafluoroborate (TDBTU). In some embodiments, the coupling-step agent is 2-(7-aza-l/i-benzotriazole-l-yl)-l, 1,3,3-tetramethyluronium
hexafluorophosphate (HATU). In some embodiments, the coupling-step agent is a phosphonic acid anhydride. In some embodiments, the phosphonic acid anhydride is a C1-C6 alkyl phosphonic anhydride. In some embodiments, the phosphonic acid anhydride is 1-propanephosphonic anhydride.
In some embodiments, the coupling-step base comprises a tertiary amine. In some
embodiments, the coupling-step base is selected from: A^N-diisopropylefhylamine (DIEA), triethylamine (TEA), N-mefhylmorpholine (NMM), 4-dimethylaminopyridine (DMAP), 2,4,6- trimethylpyridine (collidine), 2,3,5,6-tetramethylpyridine (TEMP), and 2,6-di-ieri-butyl-4- (dimethylamino)pyridine (DBDMAP). In some embodiments, the coupling-step base is triethylamine.
In some embodiments, the coupling-step solvent is acetonitrile.
In some embodiments, coupling further comprises the step of isolating (lalS',5alS')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide from the second-coupling mixture. In some embodiments, the isolating is by filtration.
In some embodiments, (lalS',5aS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide is crystalline.
In some embodiments, coupling further comprises the step of formulating (lalS',5alS')-2-(4-oxy- pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide to form a pharmaceutical composition. In some embodiments, the formulating comprises admixing (laS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- 1 /i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydro xymethyl-2,2-dimethyl- propyl)-amide with a pharmaceutical excipient. In some embodiments, the formulating is admixing (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide with a pharmaceutical excipient. II. Processes used for the preparation of (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l//-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He) and intermediates related thereto.
Ila. Processes for preparing the Compound of Formula (He)
In one embodiment, (4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He) is prepared from methyl 2- oxo-2-((lS,5¾-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) that was previously prepared according to any one of the processes described herein or by an alternative process.
Accordingly, one aspect of the present invention relates to the use of methyl 2-oxo-2-((lS,5S)- 2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) in processes for preparing (4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000041_0001
comprising the step:
condensing an intermediate of Formula (Ilc-iii):
Figure imgf000041_0002
with 2-hydrazinyl pyrazine (lid):
Figure imgf000041_0003
or an acid salt thereof; in the presence of a condensing-step solvent and optionally a condensing-step acid, provided that at least one of the acid salt or the condensing-step acid is present, to form (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate.
In some embodiments, the condensing step further comprises the step of mixing the intermediate of Formula (Ilc-iii) and the condensing-step solvent together to form a first-condensing mixture.
In some embodiments, the condensing-step solvent comprises methanol.
In some embodiments, the condensing is carried out in the presence of the condensing-step acid. In some embodiments, the condensing-step acid is HCl. In some embodiments, the condensing- step acid is aqueous HCl. In some embodiments, the condensing-step acid is about IN aqueous HCl to about 12N aqueous HCl. In some embodiments, the condensing-step acid is about 3N aqueous HCl to about ION aqueous HCl. In some embodiments, the condensing-step acid is about 5N aqueous HCl to about 7N aqueous HCl. In some embodiments, the condensing-step acid is about 6N aqueous HCl.
In some embodiments, the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture.
In some embodiments, the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing- step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 20°C to about 50°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
In some embodiments, the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 20°C to about 50°C. In some embodiments, the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing step further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
In some embodiments, the condensing step further comprises the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-condensing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline and has a powder X-ray diffraction pattern comprising one or more peaks as described herein, in terms of 1Θ.
In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 90% or greater as determined by HPLC. In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
lib. Processes for preparing the Compound of Formula (He)
The Compound of Formula (He) can be prepared using either: 1) the Compound of Formula
(IIc-iii) that is isolated from the reaction between the Compound of Formula (lie) and dimethyl oxalate or 2) the intermediate(s) that are generated in situ (as described herein) and used directly without isolation in the condensing step with 2-hydrazinyl pyrazine (lid). One aspect of the present invention relates to the two-step processes wherein the intermediate is generated in situ and used in the preparation of (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He).
Accordingly, some embodiments relate to the preparation of (4aS,5aS)-methyl l-(pyrazin-2-yl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000043_0001
comprising the steps of:
a) reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (lie):
Figure imgf000043_0002
(lie)
with dimethyl oxalate in the presence of sodium methoxide and methanol to form one or more intermediates; and
b) condensing one or more intermediates with 2-hydrazinyl pyrazine (Hd):
Figure imgf000043_0003
or an acid salt thereof; in the presence of a condensing-step solvent and optionally a condensing- step acid, provided that at least one of the acid salt or the condensing-step acid is present, to form (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate.
Reacting Step Using (15,5/f)-Bicyclo[3.1.0]hexan-2-one
In some embodiments, reacting in Step a) is carried out in the presence of an inert anhydrous atmosphere. In some embodiments, reacting in Step a) is carried out in the presence of a nitrogen atmosphere.
In some embodiments, reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture.
In some embodiments, the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the first-reacting mixture is at a temperature of about 20°C to about 25°C.
In some embodiments, reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, reacting in Step a) further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first- reacting mixture, wherein the first-reacting mixture is at a temperature of about 20°C to about 25°C.
In some embodiments, reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture.
In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
In some embodiments, reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, reacting in Step a) further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 25°C to about 50°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 30°C to about 45°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 35°C to about 40°C.
In some embodiments, the sodium methoxide is in a solution comprising methanol. In some embodiments, the sodium methoxide is about 5 weight percent to about 30 weight percent in methanol. In some embodiments, the sodium methoxide is about 25 weight percent in methanol solution. In some embodiments, the dimethyl oxalate and the sodium methoxide are present in molar excess compared to (lS,5R)-bicyclo[3.1.0]hexan-2-one.
In some embodiments, the sodium methoxide is present in molar excess compared to dimethyl oxalate.
In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.05: 1.05 to about 1 :2.4:2.8. In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.2: 1.4.
In some embodiments, the intermediate is sodium (£)-2-methoxy-2-oxo-l-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of the formula:
Figure imgf000045_0001
or sodium (Z)-2-methoxy-2-oxo-l-((llS',5lSr)-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate of the formula:
Figure imgf000045_0002
or a mixture thereof.
In some embodiments, the intermediate is not isolated.
In some embodiments, reacting in Step a) further comprises neutralizing the intermediate. In some embodiments, reacting in Step a) further comprises neutralizing the intermediate with HC1.
In some embodiments, reacting in Step a) further comprises neutralizing the intermediate comprising sodium (£)-2-methoxy-2-oxo- 1 -(( 1 S,5S)-2-oxobicyclo [3.1.0]hexan-3-ylidene)ethanolate and/or sodium (Z)-2-methoxy-2-oxo-l-((llS',5lS')-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate. In some embodiments, reacting in Step a) further comprises neutralizing the intermediate comprising sodium (£)-2-methoxy-2-oxo-l-((llS',5lSr)-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate and/or sodium (Z)-2-methoxy-2-oxo-l-((llS',5lSr)-2-oxobicyclo[3.1.0]hexan-3-ylidene)ethanolate with HC1.
In some embodiments, the intermediate is isolated.
In some embodiments, the intermediate is isolated and is methyl 2-oxo-2-((lS,5S)-2- oxobicyclo[3.1.0]hexan-3-yl)acetate of the formula:
Figure imgf000045_0003
or an enol form thereof, or a mixture of methyl 2-oxo-2-((lS,5¾-2-oxobicyclo[3.1.0]hexan-3-yl)acetate and the enol form thereof. Condensing Step
In some embodiments, condensing in Step b) further comprises the step of mixing the intermediate and the condensing-step solvent together to form a first-condensing mixture.
In some embodiments, the condensing-step solvent comprises methanol.
In some embodiments, the condensing is carried out in the presence of the condensing-step acid. In some embodiments, the condensing-step acid is HC1. In some embodiments, the condensing- step acid is aqueous HC1. In some embodiments, the condensing-step acid is about IN aqueous HC1 to about 12N aqueous HC1. In some embodiments, the condensing-step acid is about 3N aqueous HC1 to about ION aqueous HC1. In some embodiments, the condensing-step acid is about 5N aqueous HC1 to about 7N aqueous HC1. In some embodiments, the condensing-step acid is about 6N aqueous HC1.
In some embodiments, condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture.
In some embodiments, the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 55°C or below. In some embodiments, the condensing- step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about
20°C to about 50°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
In some embodiments, condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 55°C or below. In some embodiments, condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second- condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second- condensing mixture at a temperature of about 20°C to about 50°C. In some embodiments, condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 30°C to about 45°C. In some embodiments, condensing in Step b) further comprises the step of adding the condensing-step acid to the first-condensing mixture to form a second-condensing mixture, wherein the condensing-step acid is added at a rate so as to maintain the second-condensing mixture at a temperature of about 35°C to about 40°C.
In some embodiments, condensing in Step b) further comprises the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from the second-condensing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline. In some embodiments, (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline and has a powder X-ray diffraction pattern comprising one or more peaks as described herein, in terms of 1Θ.
In some embodiments, (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 90% or greater as determined by HPLC. In some embodiments, (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
In one embodiment, the condensing-step solvent is substantially methanol, the same solvent used in the processes used to prepare the Compound of Formula (IIc-iii). However, it is understood that an additional solvent could be added thereby providing a condensing-step solvent comprising methanol (i.e., methanol in the presence of at least one other added solvent). Alternatively, prior to the start of the condensing-step, methanol can be substantially removed completely and an alternative condensing-step solvent used, commonly referred in the art as a "solvent exchange" step.
III. Processes used for the preparation of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/ -cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf).
One aspect of the present invention relates to processes for preparing sodium
Figure imgf000047_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000047_0002
comprising the step of: hydrolyzing (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000047_0003
(He)
in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form the sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate. In some embodiments, hydrolyzing is carried out in the presence of an inert anhydrous atmosphere.
In some embodiments, hydrolyzing is carried out in the presence of a nitrogen atmosphere. In some embodiments, hydrolyzing further comprising the step of mixing (4aS,5aS)-methyl 1- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate and the hydrolyzing- step solvent together to form a first-hydrolyzing mixture.
In some embodiments, the hydrolyzing-step solvent comprises methanol.
In some embodiments, the first-hydrolyzing mixture is at a temperature of about 20°C to about
45°C.
In some embodiments, hydrolyzing further comprises the step of adding the sodium hydroxide to the first-hydrolyzing mixture to form a second-hydrolyzing mixture.
In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 25°C to about 60°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 35°C to about 55°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 40°C to about 50°C. In some embodiments, after the addition of the sodium hydroxide to the second-hydrolyzing mixture, the temperature of the second-hydrolyzing mixture is maintained at about 45°C.
In some embodiments, hydrolyzing further comprises the step of isolating sodium
Figure imgf000048_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate from the second-hydrolyzing mixture. In some embodiments, isolating is by filtration.
In some embodiments, sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
In some embodiments, crystalline sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
IV. Processes used for the preparation of 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l//- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilg).
One aspect of the present invention relates to processes for preparing 3-((4alS',5alS')-3-carboxy-
4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilg):
Figure imgf000049_0001
comprising the step:
oxidizing sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000049_0002
(Ilf)
in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form the 3-((4alS',5aS')-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5] cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide.
In some embodiments, oxidizing further comprises the step of mixing sodium
Figure imgf000049_0003
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate and the oxidizing-step solvent together to form a first-oxidizing mixture.
In some embodiments, the oxidizing-step solvent comprises a mixture of trifluoroacetic acid and formic acid.
In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.40 to 1.00:0.71 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about
1.00: 1.35 to 1.00:0.74 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.22 to 1.00:0.82 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.17 to 1.00:0.85 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.13 to about 1.00:0.88 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.08 to about 1.00:0.93 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.04 to about 1.00:0.96 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.02 to about 1.00:0.98 by volume. In some embodiments, the ratio of formic acid to trifluoroacetic acid is about 1.00: 1.00 by volume.
In some embodiments, the molar ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 12.5:6 to about 1 :8.5:2. In some embodiments, the molar ratio of sodium (4aS,5aS)-l -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5. In some embodiments, the molar ratio of sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 10.5:4.0.
In some embodiments, the ratio of sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.50:3.50 to about 1.00:3.50:2.50 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.55:3.45 to about 1.00:3.45:2.55 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.70:3.20 to about 1.00:3.20:2.70 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.76:3.24 to about 1.00:3.24:2.76 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000050_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.82:3.18 to about 1.00:3.18:2.82 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5aS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.88:3.12 to about 1.00:3.12:2.88 in terms of gram:mL:mL. In some embodiments, the ratio of sodium
Figure imgf000050_0002
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.94:3.06 to about 1.00:3.06:2.94 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:2.97:3.03 to about 1.00:3.03:2.97 in terms of gram:mL:mL. In some embodiments, the ratio of sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate, formic acid, and trifluoroacetic acid is about 1.00:3.00:3.00 in terms of gram:mL:mL.
In some embodiments, the first-oxidizing mixture is at a temperature of about 25°C to about 60°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 35°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 45°C to about 55°C. In some embodiments, the first-oxidizing mixture is at a temperature of about 50°C. In some embodiments, oxidizing further comprises the step of adding hydrogen peroxide to the first-oxidizing mixture to form a second-oxidizing mixture.
In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 50°C to about 60°C. In some embodiments, hydrogen peroxide is added to the first-oxidizing mixture in multiple portions at a rate so as to maintain the second-oxidizing mixture at a temperature of about 55°C.
In some embodiments, oxidizing further comprises the step of isolating 3-((4aS,5aS)-3- carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide from the second-oxidizing mixture. In some embodiments, the isolating is by filtration.
In some embodiments, 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is crystalline.
V. Processes used for the preparation of methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3- yl)acetate of Formula (IIc-iii).
The intermediate methyl 2-oxo-2-((lS,5¾-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) can be either isolated or used directly in the condensing step as described herein.
In one embodiment, the intermediate methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3- yl)acetate of Formula (IIc-iii) is isolated. It is understood that one or more tautomers are possible for the Compound of Formula (IIc-iii), for example:
Figure imgf000051_0001
The concept of tautomerization is well understood and appreciated in the art. Therefore, although only one chemical structure is shown for the Compound of Formula (IIc-iii) it is understood that all tautomeric forms are embraced by the present invention just as if each and every tautomer was specifically disclosed.
Accordingly, one aspect of the present invention relates to processes for preparing methyl 2- oxo-2-((lS,5¾-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii):
Figure imgf000052_0001
comprising the step:
reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (lie):
Figure imgf000052_0002
(He) .
with dimethyl oxalate in the presence of sodium methoxide and methanol to form methyl 2- oxo-2-((lS,5¾-2-oxobicyclo[3.1.0]hexan-3-yl)acetate.
In some embodiments, reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (He) with dimethyl oxalate, is carried out in the presence of an inert anhydrous atmosphere. In some
embodiments, the reacting step is carried out in the presence of a nitrogen atmosphere.
In some embodiments, the reacting step further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture.
In some embodiments, the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the first-reacting mixture is at a temperature of about 20°C to about 25°C.
In some embodiments, the reacting step further comprises the step of mixing (IS,5R)- bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first- reacting mixture wherein the first-reacting mixture is at a temperature of about 0°C to about 35°C. In some embodiments, the reacting step further comprises the step of mixing (lS,5R)-bicyclo[3.1.0]hexan- 2-one of Formula (He), dimethyl oxalate, and methanol together to form a first-reacting mixture wherein the first-reacting mixture is at a temperature of about 10°C to about 35°C. In some embodiments, the reacting step further comprises the step of mixing (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first-reacting mixture wherein the first-reacting mixture is at a temperature of about 15°C to about 30°C. In some embodiments, the reacting step further comprises the step of mixing (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (He), dimethyl oxalate, and methanol together to form a first-reacting mixture wherein the first-reacting mixture is at a temperature of about 20°C to about 25°C.
In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 0°C to about 50°C. In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 10°C to about 40°C. In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 15°C to about 30°C. In some embodiments, the reacting step further comprises adding sodium methoxide to the first-reacting mixture to form a second-reacting mixture, wherein the sodium methoxide is added to the first-reacting mixture at a rate so as to maintain the temperature of the second-reacting mixture at about 20°C to about 25°C.
In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 25°C to about 50°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 30°C to about 45°C. In some embodiments, after complete addition of sodium methoxide, the second-reacting mixture is maintained at a temperature of about 35°C to about 40°C.
In some embodiments, the sodium methoxide is in a solution comprising methanol. In some embodiments, the sodium methoxide is about 5 weight percent to about 30 weight percent in methanol. In some embodiments, the sodium methoxide is about 25 weight percent in methanol solution.
In some embodiments, the dimethyl oxalate and the sodium methoxide are present in molar excess compared to (lS,5R)-bicyclo[3.1.0]hexan-2-one.
In some embodiments, the sodium methoxide is present in molar excess compared to dimethyl oxalate.
In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.05: 1.05 to about 1 :2.4:2.8. In some embodiments, the molar ratio of (lS,5R)-bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1: 1.2: 1.4.
In some embodiments, the reacting step further comprises the step of neutralizing with a reacting-step acid to form methyl 2-oxo-2-((lS,5¾-2-oxobicyclo[3.1.0]hexan-3-yl)acetate. In some embodiments, the reacting-step acid is HCl. In some embodiments, the reacting-step acid is aqueous HCl. In some embodiments, the reacting-step acid is 6N aqueous HCl.
In some embodiments, the reacting step further comprises the step of isolating methyl 2-oxo-2- ((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-yl)acetate of Formula (IIc-iii) from the second-condensing mixture.
Compounds
One aspect of the present invention includes every combination of one or more compounds selected from the compound of Formulae (I), (Ila), (lib), (He), (IIc-i), (IIc-ii), (IIc-iii), (lid), (He), (Hf)> (Ilf-i), (Ilg), and (Hh), and salts, solvates, and hydrates thereof.
One aspect of the present invention relates to a compound selected from the compound of Formula (He), and salts, solvates, and hydrates thereof:
Figure imgf000054_0001
In some embodiments, the compound is (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate.
In some embodiments, (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
In some embodiments, the crystalline form of (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2° and 11.0° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, and 17.9° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, and 17.9° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.9° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.3° + 0.2°, 17.9° + 0.2°, 21.0° + 0.2°, 24.3° + 0.2°, 25.3° + 0.2°, and 25.9° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern substantially as shown in Figure 1.
One aspect of the present invention relates to isolated sodium (4aS,5aS)-l-(pyrazin-2-yl)-
4,4a,5,5a-tetrahydro- lH-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000055_0001
One aspect of the present invention relates to a crystalline form of sodium (4aS,5aS)-l- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-lH-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf).
In some embodiments, the crystalline form of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-lH-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2° and 25.3° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, and 25.3° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 2Θ, at 8.3° + 0.2°, 10.9° + 0.2°, 11.1° + 0.2°, 11.5° + 0.2°, 12.3° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°. In some embodiments, the crystalline form has a powder X-ray diffraction pattern substantially as shown in Figure 2.
One aspect of the present invention relates to the Compound of Formula (Ilia), 3-((4aS,5aS)-3- (((lS,)-3,3-dimethyl-l-(((4a,S,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide.
Indications and Methods of Treatment
In addition to the diseases and disorders described previously, Compound 1 and pharmaceutical compositions comprising Compound 1 are useful in the treatment of several other diseases and disorders, and/or in the amelioration of symptoms thereof.
Accordingly, one aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of the human or animal body by therapy.
Another aspect of the present invention relates to methods for the treatment of a CB2 receptor- mediated disorder in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein. In some embodiments, the CB2 receptor-mediated disorder is any one or more of the disorders as described herein.
Another aspect of the present invention relates to the use of a pharmaceutical composition as described herein, in the treatment of a CB2 receptor-mediated disorder. In some embodiments, the CB2 receptor-mediated disorder is any one or more of the disorders as described herein.
Another aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of a CB2 receptor-mediated disorder. In some embodiments, the CB2 receptor-mediated disorder is any one or more of the disorders as described herein.
Without limitation, additional diseases and disorders include the following.
I. Pain
The analgesic properties of cannabinoids have been recognized for many years. For example, animal studies have demonstrated that the CBi/CB2 agonists anandamide, THC, CP55,940 and WIN 55212-2 are effective against acute and chronic pain from chemical, mechanical, and thermal pain stimuli (reviewed in Walker and Huang (2002) Pharmacol. Ther. 95: 127-135; reviewed in Pacher, P et al. (2006) Pharmacol. Rev. 58(3): 389-462). In humans, topical administration of the CBi/CB2 agonist HU-210 attenuates capsaicin-induced hyperalgesia and allodynia (Rukwied, R. et al. (2003) Pain 102:283-288), and co-administration of the CBi/CB2 agonist THC and cannabidiol (nabiximols, trademark Sativex®) provides relief from cancer-associated pain (GW Pharmaceuticals press release Jan 19, 2005, Jun 19, 2007) and multiple-sclerosis-associated pain and spasticity (GW Pharmaceuticals press release Sept 27, 2005, Mar 11, 2009).
The role of CBi in mediating these analgesic effects is well-documented (reviewed in
Manzanares, J. et al. (2006) Current Neuropharmacology 4:239-57; reviewed in Pacher, P. et al. (2006) Pharmacol. Rev. 58(3): 389-462). For example, blockade of peripheral or central CBi leads to hyperalgesia (Richardson, J. D. et al. (1997) Eur. J. Pharmacol. 345: 145-153; Calignano, A. et al. (1998) Nature 394:277-281), whereas CBi activation by exogenous administration of a CBi agonist arachidonyl-2-chloroethylamide reduces pain (Furuse, S. et al. (2009) Anesthesiology 111(1): 173-86).
Although less well-documented, CB2 also plays a role in mediating analgesic effects of cannabinoids (reviewed in Guindon and Hohmann (2008) Br. J. Pharmacol. 153:319-334). For example, systemic delivery of the CB2-selective agonist AM1241 suppresses hyperalgesia induced in the carrageenan, capsaicin, and formalin models of inflammatory pain in rodents (reviewed in Guindon and Hohmann (2008) Br. J. Pharmacol. 153:319-334). Local (subcutaneous) or systemic administration of AM 1241 also reverses tactile and thermal hypersensitivity in rats following ligation of spinal nerves in the chronic constriction injury model of neuropathic pain (Malan, T. P. et al. (2001) Pain 93:239- 245; Ibrahim, M. M. et al. (2003) Proc. Natl. Acad. Sci. 100(18): 10529-10533), an effect which is inhibited by treatment with the CB2-selective antagonist AM630 (Ibrahim, M. M. et al. (2005) Proc. Natl. Acad. Sci. 102(8):3093-8). The CB2-selective agonist GW405833 administered systemically significantly reverses hypersensitivity to mechanical stimuli in rats following ligation of spinal nerves (Hu, B. et al. (2009) Pain 143:206-212). Thus, CB2-selective agonists have also been demonstrated to attenuate pain in experimental models of acute, inflammatory, and neuropathic pain, and hyperalgesia.
Accordingly, CB2-specific agonists and/or CBi/CB2 agonists find use in the treatment and/or prophylaxis of acute nociception and inflammatory hyperalgesia, as well as the allodynia and hyperalgesia produced by neuropathic pain. For example, these agonists are useful as an analgesic to treat pain arising from autoimmune conditions; allergic reactions; bone and joint pain; muscle pain; dental pain; nephritic syndrome; scleroderma; thyroiditis; migraine and other headache pain; pain associated with diabetic neuropathy; fibromyalgia, HIV-related neuropathy, sciatica, and neuralgias; pain arising from cancer; and pain that occurs as an adverse effect of therapeutics for the treatment of disease.
Furthermore, although cannabinoids exert their antinociceptive effects by complex mechanisms involving effects on the central nervous system, spinal cord, and peripheral sensory nerves (reviewed in Pacher, P. et al. (2006) Pharmacol. Rev. 58(3): 389-462), an analysis of models of inflammatory and neuropathic pain in mice that are deficient for CBi only in nociceptive neurons localized in the peripheral nervous system demonstrates that the contribution of CB type receptors expressed on the peripheral terminals of nociceptors to cannabinoid-induced analgesia is paramount (Agarwal, N. et al. (2007) Nat. Neurosci. 10(7): 870-879). Accordingly, CBi agonists that are unable to cross the blood brain barrier still find use in the treatment and/or prophylaxis of acute pain, inflammatory pain, neuropathic pain, and hyperalgesia.
Another aspect of the present invention relates to methods for the treatment of pain in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein. Some embodiments relate to methods for the treatment of pain associated with osteoarthritis in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein. Some embodiments relate to methods for the treatment of neuropathic pain in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein. Some embodiments relate to methods for the treatment of acute post-operative pain in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
Another aspect of the present invention relates to the use of a pharmaceutical composition as described herein, in the treatment of pain. Some embodiments relate to the use of a pharmaceutical composition as described herein, in the treatment of pain associated with osteoarthritis. Some embodiments relate to the use of a pharmaceutical composition as described herein, in the treatment of neuropathic pain. Some embodiments relate to the use of a pharmaceutical composition as described herein, in the treatment of acute post-operative pain.
Another aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of pain. Some embodiments relate to pharmaceutical compositions as described herein, for use in a method of treatment of pain associated with osteoarthritis. Some embodiments relate to pharmaceutical compositions as described herein, for use in a method of treatment of neuropathic pain. Some embodiments relate to pharmaceutical compositions as described herein, for use in a method of treatment of acute post-operative pain.
II. Disorders of the Immune System
Ila. Autoimmune disorders. Cannabinoid receptor agonists have been demonstrated to attenuate aberrant immune responses in autoimmune disorders, and in some cases, to provide protection to the tissue that is being inappropriately targeted by the immune system.
For example, Multiple Sclerosis (MS) is an autoimmune disorder that results in the demyelination of neurons in the CNS. The CBi/CB2 agonist THC significantly inhibits the severity of clinical disease in the Experimental Autoimmune Encephalomyelitis (EAE) mouse model of MS, an effect that is believed to be mediated by CBi on neurons and CB2 on immune cell (Maresz, K. et al. (2007) Nat. Med. 13(4):492-497). Consistent with these results, CB selective agonist WIN 55212-2 provides significant neuroprotection in the experimental allergic uveitis (EAU) model in mice (Pryce, G. et al. (2003) Brain 126:2191-2202), whereas CB2-selective agonist HU-308 markedly reduces the recruitment of immature myeloid cells and T cells, microglial and infiltrating myeloid cell proliferation, and axonal loss in the EAE model (Palazuelos, J. et al. (2008). J. Biol. Chem. 283(19): 13320-9).
Likewise, the CBi/CB2 agonist WIN 55212-2 significantly inhibits leukocyte rolling and adhesion in the brain in the EAE mouse model, an effect that is blocked by the CB2-selective antagonist SR144528 but not the CB selective antagonist SR141716A (Ni, X. et al. Mult. Sclerosis 10(2): 158-64).
Accordingly, CB2-selective agonists and/or CBi/CB2 agonists find use in the treatment and/or prophylaxis of Multiple Sclerosis and related autoimmune demyelinating diseases, e.g. Guillan-Barre syndrome, polyradiculoneuropathy and chronic inflammatory demyelination.
As another example, the autoimmune disease Rheumatoid Arthritis (RA) is a chronic, systemic inflammatory disorder of the skeletal system that principally attacks the joints to produce an inflammatory synovitis and that often progresses to destruction of the articular cartilage and ankylosis of the joints. The CBi/CBz agonists WIN 55212-2 and HU-210 significantly inhibit IL-1 alpha- stimulated proteoglycan and collagen degradation in bovine nasal cartilage explants in vitro
(Mbvundula, E. et al. (2006) J. Pharm. and Pharmacol. 58:351-358). Accordingly, CB2-selective agonists and/or CBi/CB2 agonists find use in the treatment and/or prophylaxis of autoimmune arthritic diseases, for example, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylarthritis, and reactive arthritis.
lib. Type 1 Hypersensitivity and Allergic response. Cannabinoid receptor agonists have been demonstrated to attenuate aberrant immune responses in allergic reactions as well. In type- 1, or immediate, hypersensitivity, plasma cells that have been activated by an allergen secrete IgE antibodies, which bind to Fc receptors on the surface of tissue mast cells and blood basophils and eosinophils. Repeated exposure to the same allergen results in cross-linking of the bound IgE on sensitized cells, resulting in secretion of pharmacologically active mediators such as histamine, leukotriene and prostaglandin. These mediators are responsible for the symptoms associated with allergies, including vasodilation and increased permeability, smooth muscle spasms, and leukocyte extravasation. Topical administration of the CBi/CB2 agonist HU-210 reduces these histamine-induced responses in human skin (Dvorak, M. et al. (2003) Inflamm. Res. 52:238-245). Similarly, subcutaneous injection of CBi/CB2 agonist THC or increased levels of endogenous cannabinoids reduces cutaneous inflammation and the pruritis (itch) associated with it in a mouse model for allergic contact dermatitis. (Karsak et al. (2007) Science, 316(5830), 1494-1497). In contrast, injection of the CBi receptor antagonist S141716A or the CB2 receptor antagonist SR144528 exacerbates this inflammation and pruritis. (Karsak et al. (2007) Science, 316(5830), 1494-1497). Accordingly, CB2-selective agonists and/or CBi/CBz agonists find use in the treatment of allergic reactions including atopic dermatitis (pruritis/itch), urticaria (hives), asthma, conjunctivitis, allergic rhinitis (hay fever), and anaphylaxis.
lie. Conditions Associated with CNS Inflammation. CB2 agonists have been demonstrated to attenuate inflammation in the CNS. For example, administration of CB2 agonists prevents the activation of microglia in rodent models of Alzheimer's Disease (Ashton J. C, et al. (2007) Curr. Neuropharmacol. 5(2):73-80). Likewise, administration of CB2 agonists reduces the volume of infarcts by 30% in a rodent occlusion model of stroke (Zhang, M. et al. (2007) J. Cereb. Blood Flow Metab. 27: 1387-96). Thus, CB2 agonists find use in the treatment and/or prophylaxis of neuropathologies associated with CNS inflammation, e.g. Alzheimer's, stroke-induced damage, dementia, ALS, and HIV.
lid. Conditions Associated with Vascular Inflammation. CB2 is expressed in macrophages and T cells in atherosclerotic plaques, and the CBi/CB2 agonist THC reduces the progression of atherosclerosis in ApoE knockout mice, a well-studied mouse model of atherosclerosis. The CB2-specific antagonist SRI 44528 completely blocks this effect in vitro and in vivo (Steffens, S. et al. (2005) Nature 434:782-786). Thus, CB2 agonists find use in treating atherosclerosis.
He. Other Disorders Associated with Aberrant or Unwanted Immune Response. Given the expression of CB2 on a number of different types of immune cells and the attenuating effects that CB2 agonists have been observed to have on the activities of these cells, CB2 agonists are useful for the treatment and/or prophylaxis of other disorders wherein undesired immune cell activity and/or inflammation is observed. Such exemplary disorders include osteoarthritis, anaphylaxis, Behcet' s disease, graft rejection, vasculitis, gout, spondylitis, viral and bacterial diseases, e.g. AIDS, and meningitis; and other autoimmune disorders such as lupus, e.g. systemic lupus erythematosus;
inflammatory bowel disease, e.g. Crohn' s disease, ulcerative colitis; psoriasis; autoimmune hepatitis; and type 1 diabetes mellitus.
Another aspect of the present invention relates to methods for the treatment of osteoarthritis in an individual, comprising administering to the individual in need thereof, a therapeutically effective amount of a pharmaceutical composition as described herein.
Another aspect of the present invention relates to the use of a pharmaceutical composition as described herein, in the treatment of osteoarthritis. Another aspect of the present invention relates to pharmaceutical compositions as described herein, for use in a method of treatment of osteoarthritis.
III. Bone and Joint Diseases
Ilia. Osteoporosis. CB2 is expressed in osteoblasts, osteocytes, and osteoclasts. Osteoblasts make new bone, whereas osteoclasts degrade it. The CB2-specific agonist HU-308 enhances endocortical osteoblast numbers and activity while simultaneously inhibiting proliferation of osteoclast precursors in bone marrow-derived osteoblasts/stromal cells in vitro, and attenuates ovariectomy- induced bone loss and stimulates cortical thickness by stimulating endocortical bone formation and suppressing osteoclast number in vivo (Ofek, O. et al. (2006) Proc. Natl. Acad. Sci. 103(3):696-701). Thus, CB2 agonists are useful for the treatment and/or prophylaxis of disease wherein bone density is decreased, such as osteoporosis.
Illb. Arthritis. As discussed above, CB2-selective agonists and CBi/CB2 agonists are useful for the treatment and/or prophylaxis of autoimmune arthritic diseases, for example, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylarthritis, and reactive arthritis, and for the treatment and/or prophylaxis of inflammation associated with osteoarthritis. In addition, as discussed above, CB selective agonists and CBi/CB2 agonists are useful for the treatment of pain associated with these arthritic disorders.
IV. Eye Disease
Retinal pigment epithelial (RPE) cells provide trophic support to photoreceptor cells in the eye, and RPE cell death has been demonstrated to be a major contributor to Age-related Macular
Degeneration (AMD). The CBi/CB2 agonist CP55,940 significantly protects RPE cells from oxidative damage; the CB2 receptor agonist, JWH015 provides comparable protection (Wei, Y. et al. (2009) Mol.
Vis. 15: 1243-51). Accordingly, CB2-selective agonists find use in preventing the onset or progression of vision loss associated with AMD.
V. Cough
The cough reflex is predominantly under the control of two classes of sensory afferent nerve fibers, the myelinated A-delta fibers and the non-myelinated C-fibers, the activation of which (i.e. depolarization) elicits cough via the vagus nerve afferent pathway. The CBi/CB2 agonist CP55,940 reduces capsaicin-, PGE2- and hypertonic saline-induced depolarization of guinea pig and human vagus nerve preparations in vitro (Patel, H. J. et al. (2003) British J. Pharma. 140:261-8). The CBi/CB2 agonists WIN 55212-2 produced a dose-dependent inhibition of the number of capsaicin-induced coughs in mice (Morita, K. et al. (2003) Eur. J. Pharmacol. 474:269-272). The CBi/CBz agonist anandamide produced a dose-dependent inhibition of the number of capsaicin-induced coughs in guinea pigs (Calignano, A. et al. (2000) Nature 408:96-101). CB specific antagonist SR141716A attenuates the antitussive effects of WN 55212-2 and anandamide (Morita, K. et al. (2003) Eur. J. Pharmacol. 474:269-272; Calignano, A. et al. (2000) Nature 408:96- 101). The CB2-selective agonist JWH133 reduces capsaicin-, PGE2- and hypertonic saline-induced depolarization of guinea pig and human vagus nerve preparations in vitro, and administration of CB2-selective agonist JWH133 prior to exposure to the tussive agent citric acid significantly reduces cough in conscious guinea-pigs (Patel, H. J. et al. (2003) British J. Pharma. 140:261-8). Thus, both CBi and CB2 play an important role in mediating the antitussive effect of cannabinoids, and CB selective agonists and CBi/CB2 agonists are useful in the treatment and/or prophylaxis of cough.
VI. Cancer
A number of human leukemia and lymphoma cell lines, including Jurkat, Molt-4 and Sup-Tl, express CB2 and not CBi, and agonists of CB2 induce apoptosis in these and primary acute lymphoblastic leukemia (ALL) cells (Nagarkatti, L. C. et al. US2004/0259936). Similarly, CB2 is expressed on glioblastoma cell lines and treatment with agonists of CB2 induces apoptosis of these cells in vitro (Widmer, M. (2008) /. Neurosci. Res. 86(14):3212-20). Accordingly, CB2-selective agonists are useful in attenuating the growth of a malignancy of the immune system, for example, leukemias, lymphomas, and solid tumors of the glial lineage.
In addition, as discussed above, CBi-selective agonists and CBi/CB2 agonists are useful in providing relief from pain associated with cancer (GW Pharmaceuticals press release Jan 19, 2005, Jun 19, 2007).
CB2-mediated signaling is involved in the in vivo and in vitro growth inhibition of prostate cancer cells, which suggests that CB2 agonists have potential therapeutic interest in the management of prostate cancer. (Inhibition of human tumour prostate PC-3 cell growth by cannabinoids R(+)- Methanandamide and JWH-015: Involvement of CB2; Olea-Herrero, et al. British Journal of Cancer advance online publication 18 August 2009; doi: 10.1038/sj.bjc. 6605248).
VII. Regenerative Medicine
Agonists of CB2 modulate the expansion of the progenitor pool of neurons in the CNS. CB2 antagonists inhibit the proliferation of cultured neural stem cells and the proliferation of progenitor cells in the SVZ of young animals, whereas CB2-selective agonists stimulate progenitor cell proliferation in vivo, with this effect being more pronounced in older animals (Goncalves, M. B. et al. (2008) Mol. Cell Neurosci. 38(4):526-36). Thus, agonists of CB2 are useful in regenerative medicine, for example to promote the expansion of progenitor cells for the replacement of neurons lost during injury or disease, such as Alzheimer's Disease, stroke-induced damage, dementia, amyotrophic lateral sclerosis (ALS) and Parkinson's Disease.
Polymorphs and Pseudopolymorphs
Polymorphism is the ability of a substance to exist as two or more crystalline phases that have different arrangements and/or conformations of the molecules in the crystal lattice. Polymorphs show the same properties in the liquid or gaseous state but they behave differently in the solid state.
Besides single-component polymorphs, drugs can also exist as salts and other multicomponent crystalline phases. For example, solvates and hydrates may contain an API host and either solvent or water molecules, respectively, as guests. Analogously, when the guest compound is a solid at room temperature, the resulting form is often called a cocrystal. Salts, solvates, hydrates, and cocrystals may show polymorphism as well. Crystalline phases that share the same API host, but differ with respect to their guests, may be referred to as pseudopolymorphs of one another.
Solvates contain molecules of the solvent of crystallization in a definite crystal lattice. Solvates, in which the solvent of crystallization is water, are termed hydrates. Because water is a constituent of the atmosphere, hydrates of drugs may be formed rather easily.
By way of example, Stahly recently published a polymorph screens of 245 compounds consisting of a "wide variety of structural types" revealed that about 90% of them exhibited multiple solid forms. Overall, approximately half the compounds were polymorphic, often having one to three forms. About one-third of the compounds formed hydrates, and about one-third formed solvates. Data from cocrystal screens of 64 compounds showed that 60% formed cocrystals other than hydrates or solvates. (G. P. Stahly, Crystal Growth & Design (2007), 7(6), 1007-1026).
Isotopes
The present disclosure includes all isotopes of atoms occurring in the present compounds, salts, and crystalline forms thereof. Isotopes include those atoms having the same atomic number but different mass numbers. One aspect of the present invention includes every combination of one or more atoms in the present compounds, salts, and crystalline forms thereof that is replaced with an atom having the same atomic number but a different mass number. One such example is the replacement of an atom that is the most naturally abundant isotope, such as ¾ or 12C, found in one of the present compounds, salts, and crystalline forms thereof, with a different atom that is not the most naturally
2 3 1 11 13 14 12
abundant isotope, such as H or H (replacing H), or C, C, or C (replacing C). A compound or salt thereof wherein such a replacement has taken place is commonly referred to as being isotopically- labeled. Isotopic-labeling of the present compounds, salts, and crystalline forms thereof can be accomplished using any one of a variety of different synthetic methods know to those of ordinary skill in the art and they are readily credited with understanding the synthetic methods and available reagents needed to conduct such isotopic-labeling. By way of general example, and without limitation, isotopes of hydrogen include 2H (deuterium) and 3H (tritium). Isotopes of carbon include nC, 13C, and 14C.
13 15 15 17 18
Isotopes of nitrogen include N and N. Isotopes of oxygen include O, O, and C. An isotope of fluorine includes 18F. An isotope of sulfur includes 35S. An isotope of chlorine includes 36C1. Isotopes of bromine include 75Br, 76Br, 77Br, and 82Br. Isotopes of iodine include 123I, 124I, 125I, and 131I. Another aspect of the present invention includes compositions, such as, those prepared during synthesis, preformulation, and the like, and pharmaceutical compositions, such as, those prepared with the intent of using in a mammal for the treatment of one or more of the disorders described herein, comprising one or more of the present compounds, salts, and crystalline forms thereof, wherein the naturally occurring distribution of the isotopes in the composition is perturbed. Another aspect of the present invention includes compositions and pharmaceutical compositions comprising a compound, a salt, or a crystalline form thereof as described herein wherein the compound, salt, or crystalline form is enriched at one or more positions with an isotope other than the most naturally abundant isotope. Methods are readily available to measure such isotope perturbations or enrichments, such as, mass spectrometry, and for isotopes that are radio-isotopes additional methods are available, such as, radio-detectors used in connection with HPLC or GC. Compositions and Formulations
One aspect of the present invention, inter alia, pertains to compositions and/or pharmaceutical compositions comprising: the anhydrous crystalline form of (lalS',5aS')-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2- dimethyl-propyl) -amide (Compound 1); and at least one compound selected from: a compound of Formulae (Ila), (lib), (lie), (Ilc-i), (Ilc-ii), (Ilc-iii), (lid), (He), (Iff), (Hf-i), (Hg), (Hh), (Ilia),
(Hlb), (IIIc); a solvent; and a reagent; wherein the solvent and the reagent is as described in Scheme I to Scheme VIII and the embodiments related thereto, and Example 1 to Example 6. In some embodiments, 3-((4aR,5aR)-3-(((lSr)- l-hydroxy-3,3-dimethylbutan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (IIIc) is optional.
One aspect of the present invention relates to compositions comprising:
a. the anhydrous crystalline form of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- 1 /i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydro xymethyl-2,2-dimethyl- propyl)-amide (Compound 1); and
b. at least one compound selected from:
3-((4a.S,,5alS,)-3-(((lS,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia);
(4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb);
acetonitrile; acetone; and ethyl acetate;
and optionally 3-((4aR,5aR)-3-(((lS')-l-hydroxy-3,3-dimethylbutan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (IIIc).
One aspect of the present invention relates to pharmaceutical compositions comprising:
a. the anhydrous crystalline form of (laS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl- propyl)-amide (Compound 1);
b. at least one compound selected from:
3-((4aS,,5aS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia);
(4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb); acetonitrile; acetone; and ethyl acetate; and
c. a pharmaceutically acceptable carrier;
and optionally 3-((4aR,5aR)-3-(((lS')-l-hydroxy-3,3-dimethylbutan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (IIIc).
One aspect of the present invention relates to compositions comprising:
a. the anhydrous crystalline form of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymefhyl-2,2-dimefhyl- propyl)-amide (Compound 1); and
b. a compound selected from:
3-((4a^,5a5)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia); and
(4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb).
Another aspect of the present invention relates to pharmaceutical compositions comprising: a. the anhydrous crystalline form of (laS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymefhyl-2,2-dimefhyl- propyl)-amide (Compound 1);
b. a compound selected from:
3-((4aS,,5aS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia); and
(4aS,5aS)-7V-((.S,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb);
and
c. a pharmaceutically acceptable carrier.
In some embodiments, the composition is substantially free of methylene chloride.
As used herein, "substantially free of methylene chloride" refers to the amount of methylene chloride present in a composition or pharmaceutical composition as being 600 ppm or less as determined by GC, such as head space GC.
In some embodiments, the composition is free of methylene chloride.
In some embodiments, the composition further comprises a pharmaceutically acceptable carrier.
In some embodiments, the pharmaceutical composition is substantially free of methylene chloride.
In some embodiments, the pharmaceutical composition is free of methylene chloride.
In some embodiments for the composition and/or pharmaceutical composition, the compound is 3-((4a^,5a5)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia). In some embodiments, 3-((4alS,,5alS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia) is present in the composition of less than or equal to 1.0% as determined by HPLC. In some embodiments, 3-((4alS,,5alS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (IHa) is present in the composition of less than or equal to 0.5% as determined by HPLC. In some embodiments, 3-((4alS,,5alS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia) is present in the composition of less than or equal to 0.2% as determined by HPLC.
In some embodiments for the composition and/or pharmaceutical composition, the compound is
(4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb). In some embodiments, (4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb) is present in the composition of less than or equal to 1.0% as determined by HPLC. In some embodiments,
Figure imgf000065_0001
((lSr)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb) is present in the composition of less than or equal to 0.5% as determined by HPLC. In some embodiments,
Figure imgf000065_0002
((lSr)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb) is present in the composition of less than or equal to 0.05% as determined by HPLC.
In some embodiments for the composition and/or pharmaceutical composition, the compound is acetone. In some embodiments, acetone is present in the composition of less than or equal to 5000 ppm as determined by GC. In some embodiments, acetone is present in the composition of less than or equal to 2500 ppm as determined by GC. In some embodiments, acetone is present in the composition of less than or equal to 494 ppm as determined by GC.
In some embodiments for the composition and/or pharmaceutical composition, the compound is ethyl acetate. In some embodiments, ethyl acetate is present in the composition of less than or equal to 5000 ppm as determined by GC. In some embodiments, ethyl acetate is present in the composition of less than or equal to 2500 ppm as determined by GC. In some embodiments, ethyl acetate is present in the composition of less than or equal to 5 ppm as determined by GC.
In some embodiments for the composition and/or pharmaceutical composition, the pharmaceutical composition is suitable for oral, rectal, nasal, topical, buccal, sub-lingual, vaginal, parenteral, intramuscular, sub-cutaneous, or intravenous administration, or in a form suitable for administration by inhalation, insufflation, or by a transdermal patch. In some embodiments, the pharmaceutical composition is suitable for oral administration.
Formulations may be prepared by any suitable method, typically by uniformly mixing the active compound(s) with liquids or finely divided solid carriers, or both, in the required proportions and then, if necessary, forming the resulting mixture into a desired shape.
Conventional excipients, such as binding agents, fillers, acceptable wetting agents, tabletting lubricants and disintegrants can be used in tablets and capsules for oral administration. Liquid preparations for oral administration can be in the form of solutions, emulsions, aqueous or oily suspensions and syrups. Alternatively, the oral preparations can be in the form of dry powder that can be reconstituted with water or another suitable liquid vehicle before use. Additional additives such as suspending or emulsifying agents, non-aqueous vehicles (including edible oils), preservatives and flavorings and colorants can be added to the liquid preparations. Parenteral dosage forms can be prepared by dissolving the compound provided herein in a suitable liquid vehicle and filter sterilizing the solution before filling and sealing an appropriate vial or ampule. These are just a few examples of the many appropriate methods well known in the art for preparing dosage forms.
A compound provided herein can be formulated into pharmaceutical compositions using techniques well known to those in the art. Suitable pharmaceutically-acceptable carriers, outside those mentioned herein, are known in the art; for example, see Remington, The Science and Practice of Pharmacy, 20th Edition, 2000, Lippincott Williams & Wilkins, (Editors: Gennaro et al).
While it is possible that, for use in the prophylaxis or treatment, a compound provided herein can, in an alternative use, be administered as a raw or pure chemical, it is preferable however to present the compound or active ingredient as a pharmaceutical formulation or composition further comprising a pharmaceutically acceptable carrier.
Pharmaceutical formulations include those suitable for oral, rectal, nasal, topical (including buccal and sub-lingual), vaginal or parenteral (including intramuscular, sub-cutaneous and intravenous) administration or in a form suitable for administration by inhalation, insufflation or by a transdermal patch. Transdermal patches dispense a drug at a controlled rate by presenting the drug for absorption in an efficient manner with minimal degradation of the drug. Typically, transdermal patches comprise an impermeable backing layer, a single pressure sensitive adhesive and a removable protective layer with a release liner. One of ordinary skill in the art will understand and appreciate the techniques appropriate for manufacturing a desired efficacious transdermal patch based upon the needs of the artisan.
The compounds provided herein, together with a conventional adjuvant, carrier, or diluent, can thus be placed into the form of pharmaceutical formulations and unit dosages thereof and in such form may be employed as solids, such as tablets or filled capsules, or liquids such as solutions, suspensions, emulsions, elixirs, gels or capsules filled with the same, all for oral use, in the form of suppositories for rectal administration; or in the form of sterile injectable solutions for parenteral (including
subcutaneous) use. Such pharmaceutical compositions and unit dosage forms thereof can comprise conventional ingredients in conventional proportions, with or without additional active compounds or principles and such unit dosage forms may contain any suitable effective amount of the active ingredient commensurate with the intended daily dosage range to be employed.
For oral administration, the pharmaceutical composition may be in the form of, for example, a tablet, capsule, suspension or liquid. The pharmaceutical composition is preferably made in the form of a dosage unit containing a particular amount of the active ingredient. Examples of such dosage units are capsules, tablets, powders, granules or a suspension, with conventional additives such as lactose, mannitol, corn starch or potato starch; with binders such as crystalline cellulose, cellulose derivatives, acacia, corn starch or gelatins; with disintegrators such as corn starch, potato starch or sodium carboxymethyl-cellulose; and with lubricants such as talc or magnesium stearate. The active ingredient may also be administered by injection as a composition wherein, for example, saline, dextrose or water may be used as a suitable pharmaceutically acceptable carrier.
Compounds provided herein or a solvate, hydrate or physiologically functional derivative thereof can be used as active ingredients in pharmaceutical compositions, specifically as cannabinoid CB2 receptor modulators. The term "active ingredient", defined in the context of a "pharmaceutical composition"," refers to a component of a pharmaceutical composition that provides the primary pharmacological effect, as opposed to an "inactive ingredient" which would generally be recognized as providing no pharmaceutical benefit.
The dose when using the compounds provided herein can vary within wide limits and as is customary and is known to the physician, it is to be tailored to the individual conditions in each individual case. It depends, for example, on the nature and severity of the illness to be treated, on the condition of the individual, such as a patient, on the compound employed, on whether an acute or chronic disease state is treated, or prophylaxis conducted, or on whether further active compounds are administered in addition to the compounds provided herein. Representative doses include, but are not limited to, about 0.001 mg to about 5000 mg, about 0.001 mg to about 2500 mg, about 0.001 mg to about 1000 mg, about 0.001 mg to about 500 mg, about 0.001 mg to about 250 mg, about 0.001 mg to 100 mg, about 0.001 mg to about 50 mg and about 0.001 mg to about 25 mg. Multiple doses may be administered during the day, especially when relatively large amounts are deemed to be needed, for example 2, 3 or 4 doses. Depending on the individual and as deemed appropriate from the healthcare provider it may be necessary to deviate upward or downward from the doses described herein.
All dosage amounts disclosed herein are calculated with respect to the active moiety, i.e., the molecule or ion that gives the intended pharmacologic or physiologic action.
The amount of active ingredient, or an active salt or derivative thereof, required for use in treatment will vary not only with the particular salt selected but also with the route of administration, the nature of the condition being treated and the age and condition of the individual and will ultimately be at the discretion of the attendant physician or clinician. In general, one skilled in the art understands how to extrapolate in vivo data obtained in a model system, typically an animal model, to another, such as a human. In some circumstances, these extrapolations may merely be based on the weight of the animal model in comparison to another, such as a mammal, preferably a human, however, more often, these extrapolations are not simply based on weights, but rather incorporate a variety of factors.
Representative factors include the type, age, weight, sex, diet and medical condition of the individual, the severity of the disease, the route of administration, pharmacological considerations such as the activity, efficacy, pharmacokinetic and toxicology profiles of the particular compound employed, whether a drug delivery system is utilized, whether an acute or chronic disease state is being treated or prophylaxis conducted or whether further active compounds are administered in addition to the compounds provided herein such as part of a drug combination. The dosage regimen for treating a disease condition with the compounds and/or compositions provided herein is selected in accordance with a variety factors as cited above. Thus, the actual dosage regimen employed may vary widely and therefore may deviate from a preferred dosage regimen and one skilled in the art will recognize that dosage and dosage regimen outside these typical ranges can be tested and, where appropriate, may be used in the methods disclosed herein.
The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four, or more sub-doses per day. The sub-dose itself may be further divided, e.g., into a number of discrete loosely spaced administrations. The daily dose can be divided, especially when relatively large amounts are administered as deemed appropriate, into several, for example 2, 3, or 4 part administrations. If appropriate, depending on individual behavior, it may be necessary to deviate upward or downward from the daily dose indicated.
The compounds provided herein can be administered in a wide variety of oral and parenteral dosage forms. It will be obvious to those skilled in the art that the dosage forms may comprise, as the active component, either a compound provided herein or a pharmaceutically acceptable salt, hydrate, or solvate of a compound provided herein.
For preparing pharmaceutical compositions from the compounds provided herein, the selection of a suitable pharmaceutically acceptable carrier can be either solid, liquid or a mixture of both. Solid form preparations include powders, tablets, pills, capsules, cachets, suppositories and dispersible granules. A solid carrier can be one or more substances which may also act as diluents, flavoring agents, solubilizers, lubricants, suspending agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material.
In powders, the carrier is a finely divided solid which is in a mixture with the finely divided active component.
In tablets, the active component is mixed with the carrier having the necessary binding capacity in suitable proportions and compacted to the desired shape and size.
The powders and tablets may contain varying percentage amounts of the active compound. A representative amount in a powder or tablet may contain from 0.5 to about 90 percent of the active compound; however, an artisan would know when amounts outside of this range are necessary. Suitable carriers for powders and tablets are magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter and the like. The term "preparation" refers to the formulation of the active compound with encapsulating material as carrier providing a capsule in which the active component, with or without carriers, is surrounded by a carrier, which is thus in association with it. Similarly, cachets and lozenges are included. Tablets, powders, capsules, pills, cachets and lozenges can be used as solid forms suitable for oral administration.
For preparing suppositories, a low melting wax, such as an admixture of fatty acid glycerides or cocoa butter, is first melted and the active component is dispersed homogeneously therein, as by stirring. The molten homogenous mixture is then poured into convenient sized molds, allowed to cool and thereby to solidify.
Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or sprays containing in addition to the active ingredient such carriers as are known in the art to be appropriate.
Liquid form preparations include solutions, suspensions and emulsions, for example, water or water-propylene glycol solutions. For example, parenteral injection liquid preparations can be formulated as solutions in aqueous polyethylene glycol solution. Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.
The compounds provided herein may thus be formulated for parenteral administration (e.g. by injection, for example bolus injection or continuous infusion) and may be presented in unit dose form in ampoules, pre-filled syringes, small volume infusion or in multi-dose containers with an added preservative. The pharmaceutical compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Alternatively, the active ingredient may be in powder form, obtained by aseptic isolation of sterile solid or by lyophilization from solution, for constitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use.
Aqueous formulations suitable for oral use can be prepared by dissolving or suspending the active component in water and adding suitable colorants, flavors, stabilizing and thickening agents, as desired.
Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, or other well-known suspending agents. Also included are solid form preparations which are intended to be converted, shortly before use, to liquid form preparations for oral administration. Such liquid forms include solutions, suspensions and emulsions. These preparations may contain, in addition to the active component, colorants, flavors, stabilizers, buffers, artificial and natural sweeteners, dispersants, thickeners, solubilizing agents and the like.
For topical administration to the epidermis the compounds provided herein may be formulated as ointments, creams or lotions, or as a transdermal patch.
Ointments and creams may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening and/or gelling agents. Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilizing agents, dispersing agents, suspending agents, thickening agents, or coloring agents.
Formulations suitable for topical administration in the mouth include lozenges comprising active agent in a flavored base, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert base such as gelatin and glycerin or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier.
Solutions or suspensions are applied directly to the nasal cavity by conventional means, for example with a dropper, pipette or spray. The formulations may be provided in single or multi-dose form. In the latter case of a dropper or pipette, this may be achieved by the patient administering an appropriate, predetermined volume of the solution or suspension. In the case of a spray, this may be achieved for example by means of a metering atomizing spray pump.
Administration to the respiratory tract may also be achieved by means of an aerosol formulation in which the active ingredient is provided in a pressurized pack with a suitable propellant. If the compounds provided herein or pharmaceutical compositions comprising them are administered as aerosols, for example as nasal aerosols or by inhalation, this can be carried out, for example, using a spray, a nebulizer, a pump nebulizer, an inhalation apparatus, a metered inhaler or a dry powder inhaler. Pharmaceutical forms for administration of the compounds provided herein as an aerosol can be prepared by processes well known to the person skilled in the art. For their preparation, for example, solutions or dispersions of the compounds provided herein in water, water/alcohol mixtures or suitable saline solutions can be employed using customary additives, for example benzyl alcohol or other suitable preservatives, absorption enhancers for increasing the bioavailability, solubilizers, dispersants and others and, if appropriate, customary propellants, for example include carbon dioxide, CFCs, such as, dichlorodifluoromethane, trichlorofluoromethane, or dichlorotetrafluoroethane; and the like. The aerosol may conveniently also contain a surfactant such as lecithin. The dose of drug may be controlled by provision of a metered valve.
In formulations intended for administration to the respiratory tract, including intranasal formulations, the compound will generally have a small particle size for example of the order of 10 microns or less. Such a particle size may be obtained by means known in the art, for example by micronization. When desired, formulations adapted to give sustained release of the active ingredient may be employed.
Alternatively the active ingredients may be provided in the form of a dry powder, for example, a powder mix of the compound in a suitable powder base such as lactose, starch, starch derivatives such as hydroxypropylmethyl cellulose and polyvinylpyrrolidone (PVP). Conveniently the powder carrier will form a gel in the nasal cavity. The powder composition may be presented in unit dose form for example in capsules or cartridges of, e.g., gelatin, or blister packs from which the powder may be administered by means of an inhaler.
The pharmaceutical preparations are preferably in unit dosage forms. In such form, the preparation is subdivided into unit doses containing appropriate quantities of the active component. The unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packeted tablets, capsules and powders in vials or ampoules. Also, the unit dosage form can be a capsule, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form.
Tablets or capsules for oral administration and liquids for intravenous administration are preferred compositions.
The compounds provided herein may optionally exist as pharmaceutically acceptable salts including pharmaceutically acceptable acid addition salts prepared from pharmaceutically acceptable non-toxic acids including inorganic and organic acids. Representative acids include, but are not limited to, acetic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethenesulfonic, dichloroacetic, formic, fumaric, gluconic, glutamic, hippuric, hydrobromic, hydrochloric, isethionic, lactic, maleic, malic, mandelic, methanesulfonic, mucic, nitric, oxalic, pamoic, pantothenic, phosphoric, succinic, sulfiric, tartaric, oxalic, j-toluenesulfonic and the like. Certain compounds provided herein which contain a carboxylic acid functional group may optionally exist as pharmaceutically acceptable salts containing non-toxic, pharmaceutically acceptable metal cations and cations derived from organic bases.
Representative metals include, but are not limited to, aluminium, calcium, lithium, magnesium, potassium, sodium, zinc and the like. In some embodiments the pharmaceutically acceptable metal is sodium. Representative organic bases include, but are not limited to, benzathine (TV^A^-dibenzylethane- 1,2-diamine), chloroprocaine (2-(diethylamino)ethyl 4-(chloroamino)benzoate), choline,
diethanolamine, ethylenediamine, meglumine ((2R,3R,4R,5S)-6-(methylamino)hexane-l,2,3,4,5- pentaol), procaine (2-(diethylamino)ethyl 4-aminobenzoate), and the like. Certain pharmaceutically acceptable salts are listed in Berge, et al., Journal of Pharmaceutical Sciences, 66: 1-19 (1977).
The acid addition salts may be obtained as the direct products of compound synthesis. In the alternative, the free base may be dissolved in a suitable solvent containing the appropriate acid and the salt isolated by evaporating the solvent or otherwise separating the salt and solvent. The compounds provided herein may form solvates with standard low molecular weight solvents using methods known to the skilled artisan. Compounds provided herein can be converted to "pro-drugs." The term "pro-drugs" refers to compounds that have been modified with specific chemical groups known in the art and when administered into an individual these groups undergo biotransformation to give the parent compound. Pro-drugs can thus be viewed as compounds provided herein containing one or more specialized nontoxic protective groups used in a transient manner to alter or to eliminate a property of the compound. In one general aspect, the "pro-drug" approach is utilized to facilitate oral absorption. A thorough discussion is provided in T. Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems Vol. 14 of the A.C.S. Symposium Series; and in Bioreversible Carriers in Drug Design, ed. Edward B. Roche, American Pharmaceutical Association and Pergamon Press, 1987.
Some embodiments include a method of producing a pharmaceutical composition for
"combination-therapy" comprising admixing at least one compound according to any of the compound embodiments disclosed herein, together with at least one known pharmaceutical agent as described herein and a pharmaceutically acceptable carrier.
It is noted that when the cannabinoid CB2 receptor modulators are utilized as active ingredients in pharmaceutical compositions, these are not intended for use in humans only, but in non-human mammals as well. Recent advances in the area of animal health-care mandate that consideration be given for the use of active agents, such as cannabinoid CB2 receptor modulators, for the treatment of a CB2 receptor-associated disease or disorder in companionship animals {e.g., cats, dogs, etc.) and in livestock animals {e.g., horses, cows, etc.) Those of ordinary skill in the art are readily credited with understanding the utility of such compounds in such settings.
As will be recognized, the steps of the methods provided herein need not be performed any particular number of times or in any particular sequence. Additional objects, advantages and novel features of the invention(s) will become apparent to those skilled in the art upon examination of the following examples thereof, which are intended to be illustrative and not intended to be limiting.
EXAMPLES
The compounds disclosed herein and their syntheses are further illustrated by the following examples. The following examples are provided to further define the invention without, however, limiting the invention to the particulars of these examples. The compounds described herein, supra and infra, are named according to the AutoNom version 2.2, CS ChemDraw Ultra Version 9.0.7, or ChemBioDraw Ultra 12.0.2.1076. In certain instances common names are used and it is understood that these common names would be recognized by those skilled in the art.
Chemistry: Proton nuclear magnetic resonance (¾ NMR) spectra were recorded on a Bruker Avance III-400 equipped with a 5 mm BBFO probe. Chemical shifts are given in parts per million (ppm) with the residual solvent signal used as reference. NMR abbreviations are used as follows: s = singlet, d = doublet, dd = doublet of doublets, t = triplet, q = quartet, m = multiplet, bs = broad singlet, sxt = sextet. Microwave irradiations were carried out using a Smith Synthesizer™ or an Emrys
Optimizer™ (Biotage). Thin-layer chromatography (TLC) was performed on silica gel 60 F254 (Merck), preparatory thin-layer chromatography (prep TLC) was performed on PK6F silica gel 60 A 1 mm plates (Whatman) and column chromatography was carried out on a silica gel column using Kieselgel 60, 0.063-0.200 mm (Merck). Evaporation was done under reduced pressure on a Biichi rotary evaporator. Celite® 545 was used for filtration of palladium.
LCMS spec: HPLC- Agilent 1200; pumps: G1312A; DAD:G1315B; Autosampler: G1367B;
Mass spectrometer- Agilent G1956A; ionization source: ESI; Drying Gas Flow: 10 L/min; Nebulizer Pressure: 40 psig; Drying Gas Temperature: 350°C; Capillary Voltage: 2500 V) Software: Agilent Chemstation Rev.B.04.03. Example 1: Preparation of (lS,2S,5tf)-bicyclo[3.1.0]hexan-2-ol (Formula (lib)).
A solution of (S)-l,2-epoxy-5-hexene (1.3 kg, 13.25 mol), methyl ieri-butyl ether (MTBE; 9.62 kg), and tetramethylpiperidine (TMP; 0.94 kg, 6.65 mol) was charged to a 50 L glass lined reactor equipped with overhead agitation, jacket temperature control, and a nitrogen inlet. The reactor contents were cooled to -10°C to 0°C before adding w-butyllifhium (9.94 L). The addition rate was such so as to keep the temperature of the reactor below 0°C. The reaction to form (lS,2S,5R)-bicyclo[3.1.0]hexan-2- ol was deemed complete when the amount of (S)-l,2-epoxy-5-hexene was less than 2% (peak area) as determined by GC.
The reactor contents were cooled to 0°C and 3M HC1 (6.5 L) was added to the mixture slowly so as to maintain an internal temperature of < 10°C. The phases were separated and the organic layer washed with 3M HC1 (2.6 L). The combined aqueous layer was extracted twice with 5.8 kg and 2.4 kg of MTBE respectively. The combined MTBE layers were concentrated to one half the original volume and was used directly in the oxidation step, see Example 2. ¾ NMR (CDC13, 400 MHz) δ -0.02-0.01 (m, 1H), 0.38-0.43 (m, 1H), 1.26-1.36 (m, 2H), 1.37-1.42 (m, 1H), 1.53 (dd, J = 8.4, 14.4 Hz, 1H), 1.64 (dd, J = 8.0, 12.4 Hz, 1H), 1.88-1.97 (m, 1H), 2.42 (s, 1H), 4.20 (d, 7 = 4.8 Hz, 1H); 13C NMR (CDC13, 100 MHz) δ 6.8, 16.2, 24.2, 24.6, 30.3, 74.3.
Example 2: Preparation of (lS,5/f)-bicyclo[3.1.0]hexan-2-one (Formula (lie)).
A buffer solution was made by adding in a 50 L reactor, potassium phosphate dibasic (3.5 kg), potassium phosphate monobasic (0.37 kg), potassium bromide (0.24 kg), and water (6.5 L). The mixture was stirred at ambient for 5 minutes, at which time a clear solution formed. The buffer solution was charged to a 50 L reactor containing (lS,2S,5R)-bicyclo[3.1.0]hexan-2-ol (prepared according to Example 1). TEMPO (0.06 kg) was charged to the reaction mixture followed by the controlled addition of sodium hypochlorite (11.4 L) via an addition funnel so as to keep the temperature below 10°C. The reaction was deemed complete when the amount of (lS,2S,5R)-bicyclo[3.1.0]hexan-2-ol was less than 2% (peak area) as determined by GC.
The reaction mixture was quenched with 2.4M aqueous sodium sulfite solution (2.6 L) until the reaction was negative for oxidant by starch iodide indicator paper. The mixture was filtered and the filtrate set aside. The filter cake was washed with MTBE (2.93 kg). The filtrate and the wash were combined and transferred to the reactor. The phases were separated and the aqueous phase was extracted twice with 4.8 kg and 2.4 kg of MTBE respectively. The combined organic phase was concentrated under reduced pressure and assayed for 0.99 kg, 77.9% yield; ¾ NMR (CDC13, 400 MHz) δ 0.92-0.95 (m, 1H), 1.17-1.23 (m, 1H), 1.74-1.78 (m, 1H), 1.97-2.18 (m, 5H); 13C NMR (CDC13, 100 MHz) δ 13.5, 21.6, 22.6, 27.4, 31.4, 215.2.
Example 3: Preparation of (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/ - cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He)).
To a nitrogen-purged 50-L reactor was charged (lS,5R)-bicyclo[3.1.0]hexan-2-one (2.0 kg, 20.81 mol), dimethyl oxalate (2.84 kg, 24.05 mol, 1.2 eq.), and methanol (1.53 kg). The mixture was stirred at 20-25°C and sodium methoxide solution (25 wt% in methanol, 6.49 kg, 30.04 mol, 1.44 eq.) added slowly so as to keep the temperature of the reaction mixture below 40°C. The internal temperature of the reaction mixture was kept between 35-40°C and the mixture monitored by HPLC for the disappearance of starting material (i.e., (lS,5R)-bicyclo[3.1.0]hexan-2-one). The reactor contents were then cooled to 30°C before adding 2-hydrazinylpyrazine (2.43 kg, 22.07 mol, 1.06 eq.) to the reactor. Thereafter, 6N HC1 (6.76 L) was charged slowly to the reactor while maintaining an internal temperature of 35-40°C. The mixture was then stirred at 35-40°C and monitored for reaction completion by HPLC. The reaction was considered complete when the diketo methyl ester intermediate of Formula (Ilc-i) was less than 3 area % as determined by HPLC
The mixture was concentrated by distillation under vacuum at a jacket temperature of 25-50°C until no appreciable distillate was observed. Water (19.3 kg) was charged to the reactor and the mixture concentrated by distillation under maximum vacuum at a jacket temperature of 25-50°C until no appreciable distillate was observed. Water (9.65kg) was added to the reaction mixture and the mixture stirred at 20-25°C for at least 1 hour. The mixture was filtered and the filter cake washed with water (2 x 9.65 kg). The solids were placed in a vacuum oven set at 45-55°C and dried to a constant weight. The yield of (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate was 3.09 kg (58%); ¾ NMR (DMSO- , 400 MHz): δ 0.42-0.45 (td, J = 4.5, 7.8 Hz, 1H), 1.25- 1.30 (m, 1H), 2.28-2.34 (m, 1H), 2.71 -2.74 (m, 1H), 2.77 (d, / = 16.0 Hz, 1H), 2.90 (dd, / = 6.5, 16.4 Hz, 1H), 3.83 (s, 3H), 8.61 (dd, / = 1.5, 2.6 Hz, 1H), 8.68 (d, J = 2.5 Hz, 1H), 9.16 (d, / = 1.40 Hz, 1H); 13C NMR (DMSO- , 100 MHz) δ 16.80, 17.06, 23.50, 26.18, 51.83, 129.28, 135.15, 139.57, 142.47, 142.89, 147.07, 154.47, 161.62.
A powder X-ray diffraction (PXRD) pattern for a sample containing (4aS',5alS')-methyl 1- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is shown in Figure 1. Certain PXRD peaks for (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate are shown in Table 3. Table 3
Figure imgf000075_0002
Example 4: Preparation of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l//- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (IIf».
In a 50-L reactor was charged (4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (3.05 kg, 11.90 mol 1.00 eq.) followed by methanol (29.46 kg). To the stirred reaction mixture was added sodium hydroxide (50 wt.% aqueous; 1.45 kg, 18.1 moles, 1.52 eq.). The reaction mixture was stirred under nitrogen at 45°C for over 3 hours until the amount of starting material (i.e., (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate) was less than 0.5% (area) as determined by HPLC.
The mixture was concentrated by distillation under vacuum at a jacket temperature of 25-50°C until no appreciable distillate was observed. Thereafter, ethyl acetate (41.3 kg) was charged to the reactor and the mixture was concentrated by distillation under maximum vacuum at a jacket temperature of 25-50°C until 2 to 5 volumes remained in the reactor. Water (2.5 kg) was charged to the reactor followed by acetone (12.2 kg) and the mixture was heated to reflux for at least 1 hour. The mixture was cooled to 50°C and acetone (23.9 kg) was added to the reaction mixture to form a slurry comprising crystalline product. The mixture was stirred at 20-25°C for 1 hour. The mixture was filtered and the filter cake washed with acetone (24.4 kg). The solids were placed in a vacuum oven set at 50- 60°C and dried to a constant weight. The yield of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate was 3.28 kg (quantitative); ¾ NMR (DMSO- , 400 MHz): δ 0.31-0.32 (m, IH), 1.18-1.23 (m, IH), 2.17-2.24 (m, IH), 2.66 (bs, IH), 2.71 (d, 7 = 17.4 Hz, IH), 2.81 (dd, 7 = 6.1, 16.2 Hz, IH), 8.53 (bs, IH), 8.55 (bs, IH), 9.31 (s, IH); 13C NMR (DMSO- , 100 MHz) δ 16.96, 17.00, 23.15, 26.64, 128.13, 135.44, 141.24, 142.09, 147.67, 150.14, 153.02, 164.92.
A powder X-ray diffraction (PXRD) pattern for a sample containing sodium
Figure imgf000075_0001
(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is shown in Figure 2. Certain PXRD peaks for sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate are shown in Table 4. Table 4
Figure imgf000076_0002
Example 5: Preparation of 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l//- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Formula (Ilg)).
To a 50-L reactor was charged sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (3.20 kg, 12.11 mol 1.00 eq.), trifluoroacetic acid (14.50 kg, 127.2 mol) and formic acid (11.78 kg). The reaction mixture was stirred and heated at 50°C. Hydrogen peroxide solution (30 wt.% aqueous; 5.54 kg, 48.9 moles, 4.03 eq.) was charged in four portions over a 4-hour period while maintaining the stirred reactor contents at a temperature not to exceed 60°C with reactor jacket cooling. The reaction mixture was stirred under nitrogen at 55°C for over 2 hours until the amount of free acid (i.e., (laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3- diaza-cyclopropa[a]pentalene-4-carboxylic acid of Formula (Ilf-i)) was less than 0.6% (area) as determined by HPLC.
Figure imgf000076_0001
(Ilf-i)
Water (32.2 kg) was charged to the reactor and the mixture stirred at 0-5°C for 1 hour. The mixture was filtered and the filter cake washed with water (64.4 kg). The solids were placed in a vacuum oven set at 60°C and dried to a constant weight. The yield of 3-((4aS,5aS)-3-carboxy-4,4a,5,5a- tetrahydro- l/i-cyclopropa[4,5]cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide was 1.65 kg (52.8%); ¾ NMR (DMSO- , 400 MHz): δ 0.42 (td, J = 4.5, 7.8 Hz, 1H), 1.23-1.28 (m, 1H), 2.25-2.31 (m, 1H), 2.66-2.69 (m, 1H), 2.73 (d, J = 16.6 Hz, 1H), 2.86 (dd, / = 6.4, 16.5 Hz, 1H), 8.31 (dd, / = 1.5, 4.1 Hz, 1H), 8.48 (d, / = 4.0 Hz, 1H), 8.52 (d, / = 1 Hz, 1H), 13.06 (bs, 1H); 13C NMR (DMSO- , 100 MHz) δ 16.87, 17.18, 23.53, 26.17, 123.94, 129.51, 132.45, 141.11, 145.97, 150.96, 154.71, 162.43.
A powder X-ray diffraction (PXRD) pattern for a sample containing 3-((4alS',5alSr)-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide is shown in Figure 3. Certain powder X-ray diffraction peaks for 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide are shown in Table 5. Table 5
Figure imgf000077_0001
Example 6: Preparation of (la5,5a5)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/ -2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2-dimethyl-propyl)-amide (Compound 1, Formula (I)).
To 3-((4alS',5alSr)-3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l-yl)pyrazine 1 -oxide (1.62 kg, 6.27 mol) was added acetonitrile (12.75 kg) and the slurry cooled to 0- 5°C. To the mixture was added triethylamine (1.26 kg; 12.5 mol; 2.0 eq.) and the mixture stirred at 0- 5°C for 0.5 hour. To the resulting solution at 0-5°C was added HATU (2.85 kg, 7.50 mol, 1.2 eq.) and the mixture stirred at 0-5°C for 0.25 hour. Thereafter, a solution of (lSr)-2-amino-3,3-dimethylbutan-l-ol (i.e., (S)-ieri-leucinol) (0.87 kg, 7.42 mol, 1.18 eq.) in acetonitrile (7.38 kg) was added all at once. The resulting mixture was stirred at 0-5°C. The progress of the reaction is monitored by HPLC and the reaction was deemed complete when the amount of starting material (i.e., 3-((4alS',5aS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide) was less than 0.5% (area) as determined by HPLC.
To the mixture was added concentrated aqueous hydrochloric acid (0.08 kg) in purified water
(0.75 kg) and the stirred reactor contents evaporated under reduced pressure at 20-45°C until no appreciable distillate was observed. To the reactor contents was added water (16.23 kg) and the mixture evaporated under reduced pressure at 20-45°C until no appreciable distillate was observed. Thereafter, ethyl acetate (14.61 kg), and potassium carbonate (0.81 kg) dissolved in water (1.62 kg) was added to the reactor contents. The mixture was stirred at room temperature for at least 0.5 hour. The biphasic solution was allowed to settle and the phases separated. The aqueous layer was extracted twice with ethyl acetate (2 x 7.32 kg).
The combined ethyl acetate layer was evaporated under reduced pressure at 25-45°C. The mixture was then chased with ethanol (12.9 kg) under reduced pressure at 20-45°C. Thereafter, aqueous potassium carbonate solution (0.32 kg dissolved in 1.62 kg water) was added and the mixture was heated to reflux for 2 hours. The resulting solution was cooled to 35-40°C and water (19.44 kg) was added slowly. The mixture was stirred at 35-40°C for a minimum of 2 hours when solids began to form. Additional water (8.14 kg) was added to the mixture slowly at 35-40°C. Thereafter, the mixture was cooled to 0-5°C and stirred for a minimum of 2 hours and filtered. The filter cake was washed with water (2 x 8.1 kg).
The filter cake was transferred to the reactor and dissolved in acetonitrile (3.84 kg)/water (1.62 kg) at 55-65°C. The solution was cooled to 35-40°C and water (8.10 kg) was added slowly. The mixture was stirred at 35-40°C vigorously for a minimum of 2 hours when solids began to form.
Additional water (14.58 kg) was slowly added to the mixture at 35-40°C. Thereafter, the mixture was stirred for a minimum of 3 hours at 0-5 °C and filtered. The filter cake was washed with water (2 x 8.1 kg). The resulting filter cake was dried in a vacuum oven at 65°C to a constant weight (i.e., dry time was 40.5 hrs). The yield of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimethyl-propyl)-amide
(Compound 1) was 1.68 kg (74.9%; two additional batches were prepared in a similar manner as described herein with yields of 72.3% and 70.7% respectively for an average yield for the three batches of 72.6%); HPLC purity 99.8%; enantiomeric purity > 99.9% as determined by the amount of the Compound of Formula (IIIc) present (i.e., < 0.1% (quantitation limit)); water content (Karl Fisher) 0.1 %; for more anal
Figure imgf000078_0001
¾ NMR (DMSO- , 400 MHz): δ 0.42 (td, / = 4.4, 7.8 Hz, 1H), 0.93 (s, 9H), 1.22-1.28 (m, 1H), 2.26-2.32 (m, 1H), 2.64-2.67 (m, 1H), 2.77 (d, / = 16.7 Hz, 1H), 2.85 (dd, / = 6.2, 16.6 Hz, 1H), 3.53-3.59 (m, 1H), 3.68-3.73 (m, 1H), 3.81-3.86 (m, 1H), 4.56 (t, / = 5.4 Hz, 1H, OH), 7.69 (d, / = 9.8 Hz, 1H), 8.28 (dd, / = 1.6, 4.2 Hz, 1H), 8.45 (d, / = 4.3 Hz, 1H), 9.08 (d, / = 1.1 Hz, 1H); 13C NMR (DMSO- , 100 MHz) δ 16.88, 16.95, 23.75, 26.01, 26.95, 34.01, 58.63, 60.30, 124.16, 127.84, 132.07, 143.35, 145.78, 151.10, 154.68, 160.89.
The PXRD of the isolated product is shown in Figure 7 and conforms to the PXRD for the anhydrous crystalline form of Compound 1 previously reported in International Publication Number WO2012/116276. Representative PXRD peaks for Compound 1 prepared according to Example 6 are shown below in Table 6. Table 6
Figure imgf000079_0002
Example 7: Evaluation of the oxidation step in the preparation of 3-((4a5,5aS)-3-carboxy- 4,4a,5,5a-tetrahydro-l/ -cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Formula dig)).
The intermediate, 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide, was previously prepared in
WO2011/025541(see Example 1.108) and WO2012/116276 (see Example 1, METHOD 2, Step E) wherein both procedures use substantially the same procedure of formic acid and 50 wt. % aqueous solution of hydrogen peroxide at a temperature 58 to 64°C. However, in both procedures it was noted that under these conditions the isolated product contained 2 % starting material by ¾ NMR. In addition, it was observed that using 50 wt. % aqueous solution of hydrogen peroxide with formic acid resulted in an exothermic reaction that was difficult to control on large scale. Accordingly, the reaction conditions for the oxidation step for converting (laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a-tetrahydro-lH- 2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid (Compound of Formula (Ilf)) to 3 -((4aS,5aS)-3- carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide (Compound of Formula (Ilg)) were evaluated to address these and other issues. The term "volume" used in this example is merely a scaling factor and refers to the number of milliliters used for a respective solvent per gram of Compound of Formula (Ilf), for example formic acid (10 volume) refers to 5 mL/0.5 gram, 10 mL/1 gram, 50 mL/5 grams, 100 mL/10 gram, and the like of formic acid/Compound of
Figure imgf000079_0001
(iif) (iig)
As a preliminary study, the oxidation step was evaluated using formic acid (10 volume) with hydrogen peroxide (30 wt.%, 7 eq.) was monitored by HPLC for 17 hours at 50-55°C. Progress of reaction for the conversion of (laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a-tetrahydro-lH-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid (Compound of Formula (Ilf)) to 3-((4aS,5aS)-3-carboxy- 4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide (Compound of Formula (Ilg)) is shown below in Table 7.
Table 7
Figure imgf000080_0001
After 17 hours, the reaction mixture was diluted with water (15x initial volume) and the resulting product mixture was isolated and analyzed. The oxidation step converting the Compound of Formula (Ilf) to the Compound of Formula (Ilg) did not go to completion under these conditions even in the presence of 7 eq. of hydrogen peroxide (30 wt.%) after 17 hours at 50-55°C.
The oxidation step was modified reducing the amount of formic acid from 10 to 6 volumes with hydrogen peroxide (30 wt.%). The progress of the reaction was followed by HPLC for the conversion of the Compound of Formula (Ilf) to the Compound of Formula (Ilg) and is shown below in Table 8.
Table 8
Figure imgf000080_0002
Results from the progress of the reaction in formic acid (6 volumes) with 30 wt.% hydrogen peroxide (4 eq. added incrementally) at 50 -55°C. When the conversion became slow (after 3 hours), trifluoroacetic acid was added incrementally. Although the addition of TFA was observed to increase the rate of formation to the Compound of Formula (Ilg) the reaction stalled after 6 hours.
The oxidation step was further modified replacing formic acid with trifluoroacetic acid (4 volume) and using hydrogen peroxide (30 wt.%). In a similar manner, the progress of the reaction was followed by HPLC for the conversion of the Compound of Formula (Ilf) to the Compound of Formula (Ilg) and is shown below in Table 9. Table 9
Figure imgf000081_0001
During the reaction with trifluoroacetic acid, it was observed that only 0.2 % area of the Compound of Formula (Ilf) remained after 4 hours of reaction time. Although encouraging, other impurities increased during the synthesis leading to a reduced yield and a more complicated mixture that resulted in a difficult purification requiring repeated manipulation to provide the Compound of Formula (Iig).
The oxidation step was again further modified using formic acid (2.5 volume) and trifluoroacetic acid (3.0 volume) with Hydrogen Peroxide (30 wt.%). In a similar manner, the progress of the reaction was followed by HPLC for the conversion of the Compound of Formula (Ilf) to the Compound of Formula (Iig) and is shown below in Table 10.
Table 10
Figure imgf000081_0002
The reaction was observed to form the Compound of Formula (Iig) with only 1.1 area% of starting material (i.e., Compound of Formula (Ilf)) remaining after 4 hours of reaction. The isolated purity of Compound of Formula (Iig) was 98.3 % area.
Table 11 shows the progress of the oxidation step using formic acid (3 volumes) and trifluoroacetic acid (3 vol) with 30 wt.% hydrogen peroxide at 50-55°C.
Table 11
Figure imgf000081_0003
The isolated purity of Compound of Formula (Iig) was 98.5 %area. Other impurities were at or less than 0.3 %area, including the Compound of Formula (Ilf)- These results led to the procedure described in Example 5. Example 8: Representative gas chromatography (GC) methods for in-process control (IPC) and/or assessing chiral purity for (S)-l,2-epoxy-5-hexene (Formula (Ila)), (1S,2S,5R)- bicyclo[3.1.0]hexan-2-ol (Formula (lib)), and (15,5tf )-bicyclo[3.1.0]hexan-2-one (Formula (lie)).
Example 8.1: GC Method 1.
The sample was prepared as a solution in acetonitrile. The sample was then injected onto a
Restek RtyDEXsa GC column and monitored using a flame ionization detector (FID). The chiral purity was determined quantitatively by comparing the peak area ratios for the sample chromatograms to the corresponding peak area ratios for the standard chromatograms. The method conditions and a representative chromatogram for the sample solution are presented below:
Figure imgf000082_0001
The retention times (ir, minutes) for a certain list of intermediates were determined using the method directly above and are shown in the following table.
Figure imgf000082_0002
Example 8.2: GC Method 2.
The sample was prepared as a solution in acetonitrile. The sample was then injected onto a Restek Rtx-5 capillary GC column and monitored using a flame ionization detector (FID). The chiral purity was determined quantitatively by comparing the peak area ratios for the sample chromatograms to the corresponding peak area ratios for the standard chromatograms. The method conditions and a representative chromatogram for the sample solution are presented below:
Figure imgf000082_0003
100% methanol for (lib)
Needle wash
100% acetonitrile for (Ila) and (lie)
Split ratio 1 :25
Carrier gas Helium, constant flow at ~1 mL/min
Hydrogen flow 40 mL/min
Air flow 450 mL/min
Make up gas flow 50 mL/min, constant make up
The retention times (tT, minutes) for a certain list of intermediates were determined using the method directly above and are shown in the following table.
Figure imgf000083_0001
Example 8.3: GC Method 3.
The sample was prepared as a solution in acetonitrile. The sample was then injected onto a Restek RtyDEXsa GC column and monitored using a flame ionization detector (FID). The chiral purity was determined quantitatively by comparing the peak area ratios for the sample chromatograms to the corresponding peak area ratios for the standard chromatograms. The method conditions and a representative chromatogram for the sample solution are presented below:
Figure imgf000083_0002
The retention times (tT, minutes) for a certain list of intermediates were determined using the method directly above and are shown in the following table.
Figure imgf000083_0003
Example 9: Representative high-performance liquid chromatography/high-pressure liquid chromatography (HPLC) methods for in-process control (IPC) and/or assessing purity for:
(lS,5tf)-bicyclo[3.1.0]hexan-2-one (Formula (lie)); methyl 2-oxo-2-((lS,5S)-2-oxobicyclo[3.1.0]hexan-3-yl)acetate (intermediate in the condensing step);
(4aS,5aS) -methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/ -cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate (Formula (He));
(laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/ -2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid of (Formula (Ilf-i)) or the free acid of the sodium salt of Formula (IIf»;
3-((4a5,5a5)-3-carboxy-4,4a,5,5a-tetrahydro-l/ -cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l-yl)pyrazine 1-oxide (Formula (Ilg)); and
(laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/ -2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2-dimethyl-propyl)-amide (Compound 1, Formula (I)).
The sample solution in acetonitrile was injected onto an Agilent Poroshell 120 EC-C18 column and monitored by UV detection at 268 nm. The working concentration was about 0.18 mg/mL for assay, related substance, and identification. The percent assay of (4alS',5aS')-methyl l-(pyrazin-2-yl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate (Formula (He)) in the starting material is determined against an external reference standard(s). The positive identification of (He) is established by comparing the retention time of the (He) peak against that of the reference standard. The method conditions are presented below:
Figure imgf000084_0001
The retention times (tT, minutes) for a certain list of intermediates were determined using the method directly above and are shown in the following table.
Figure imgf000084_0002
Observed
Compound
retention time
(4aS,5aS)-mefhyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate 9.1 minutes
(Formula (He))
(laS,5aS)-2-(pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid 6.1 minutes (i.e., free acid of sodium salt of Formula (Ilf))
3-((4aS,5a¾-3-carboxy-4,4a,5,5a-tetrahydro-l#-
4.4 minutes cyclopropa[4,51cyclopenta|T,2-c|pyrazol-l-yl)pyrazine 1-oxide (Formula (Ilg))
(laS,5a¾-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymefhyl-2,2-dimethyl- 7.7 minutes propyl)-amide (Compound 1, Formula (I))
Example 10: PathHunter β-arrestin assay; (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l//-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)- l-hydroxymethyl-2,2-dimethyl- propyl)-amide (Compound 1, Formula (I)).
Compound 1 was screened for agonist activity against the human CB2 (hCB2) receptor using the DiscoveRx PathHunter β-arrestin assay which measures the β-arrestin binding to the CB2 receptor upon its activation. CB2 was cloned into the pCMV-PK vector (DiscoveRx, Fremont, CA; catalog # 93- 0167) and transfected into the CHO-K1 EA-Arrestin parental cell line (DiscoveRx, Fremont, CA; catalog # 93-0164). CHO-K1 positive clones stably expressing the CB2-ProLink fusion protein were identified by their responses to the CB2 agonist CP55,940. Clone # 61 was chosen for its big agonist window and homogenous expression as detected by anti-HA flow cytometry.
Principle of the assay: The PathHunter β-arrestin assay measures the interaction of β-arrestin with activated GPCRs using Enzyme Fragment Complementation (Yan et al., J. Biomol. Screen. 7: 451- 459, 2002). A small, 42 amino acid β-galactosidase fragment, Prolink, is fused to the c-terminus of a GPCR, and β-arrestin is fused to the larger β-galactosidase fragment, EA (Enzyme Acceptor). Binding of β-arrestin to the activated GPCR causes the complementation of the two enzyme fragments, forming an active β-galactosidase enzyme which can be measured using the chemiluminiescent PathHunter Flash Detection Kit (DiscoveRx, Fremont, CA: catalog # 93-0001).
The assay: The stable CHO-K1 cells expressing CB2-Prolink fusion protein were plated overnight in 384-well plates (Optiplate 384-Plus, PerkinElmer, Fremont CA; catalog # 6007299) at 5000 cells/5 μίΛνεΙΙ in the Opti-MEM medium (Invitrogen, Carlsbad, CA; catalog # 31985088) with 1 % FBS. 5uL of test compound diluted in Opti-MEM supplemented with 1 % BSA was transferred to each well of the Optiplate. The plates were then incubated at 37°C/5 C02 for two hours. 12 μΕ of substrate prepared from the PathHunter Flash Detection Kit (DiscoveRx, Fremont, CA: catalog # 93- 0001) was transferred to each well of the Optiplate. The plate was then incubated in the dark at room temperature for 2 h, after which the assay plate was read.
Assay readout: β-Arrestin assay readout was accomplished using a PHERAstar (BMG Labtech Inc., Durham, NC) or an En Vision™ (PerkinElmer, Fremont CA) microplate reader. (1α5,5α¾-2-(4-ο γ-ργΓαζϊη-2-γ1)-1α,2,5,5α-ΙεΐΓώγάΓθ- 1/ί-2,3-(1ϊαζα-ογο1ορΓορα[α]ρεηΙα1εηε-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide (Compound 1) was observed to be a selective agonist for CB2 with an EC50 value of 5.4 nM and substantially inactive against hCBl . Example 11: Effect of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/ -2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2-dimethyl-propyl)-amide (Compound 1) on osteoarthritis pain.
Injection of monosodium iodoacetate (MIA) into a joint (Kalbhen D. A., J. Rheumatol., 1987, May; 14 Spec No: 130-1; Combe, R., et. al., Neuroscience Letters, 2004, 370, 236-240) inhibits the activity of glyceraldehyde-3-phosphate dehydrogenase in chondrocytes, resulting in disruption of glycolysis and eventually in cell death. The progressive loss of chondrocytes results in histological and morphological changes of the articular cartilage, closely resembling those seen in osteoarthritis patients.
The osteoarthritis was induced in 200 g male Sprague Dawley rats. After brief anesthesia by isoflurane, rats received a single intra-articular injection of MIA (2 mg) (Sigma Aldrich, Saint Louis, MO, USA; Cat # 19148) dissolved in 0.9% sterile saline in a 50
Figure imgf000086_0001
volume administered through the patella ligament into the joint space of the left knee with a 30G needle. Following the injection, animals were allowed to recover from anesthesia before being returned to the main housing vivarium.
Typically during disease progression, there is an inflammation period of 0-7 days post-intra- articular injection followed by progressive degeneration of the cartilage and subchondral bone from days 14-55. Efficacy studies with a compound of the present invention for pain development took place from day 14 onwards and were performed twice a week with at least 3 days' wash-out in between each assay. Three different assays were used to measure pain. Tactile allodynia was measured via von Frey assay, hind limb paw weight distribution was monitored using an incapacitance tester (Columbus Instruments, Columbus, OH, USA) and hind limb grip strength was measured using a grip strength meter (Columbus Instruments, Columbus, OH, USA). Briefly, the von Frey assay was performed using the standard up/down method with von-Frey filaments. Hind paw weight distribution was determined by placing rats in a chamber so that each hind paw rests on a separate force plate of the incapacitance tester. The force exerted by each hind limb (measured in grams) is averaged over a 3 second period. Three measurements were taken for each rat, and the change in hind paw weight distribution calculated. Peak hind limb grip force was conducted by recording the maximum compressive force exerted on the hind limb mesh gauge set on the grip strength meter. During the testing, each rat was restrained and the paw of the injected knee was allowed to grip the mesh. The animal was then pulled in an upward motion until their grip was broken. Each rat is tested 3 times, with the contralateral paw used as a control.
Animals were base-lined prior to treatment of the test compound. The MIA treated groups of rats (6 per group) were then dosed with either vehicle (0.5% methylcellulose, orally) or Compound 1 (at 0.3 mg/kg, 1 mg kg, 3 mg kg, 10 mg kg, and 30 mg kg, orally). Dosing volume was 500 μΕ. One hour after dosing, von Frey assay, hind limb weight distribution and/or hind limb grip analysis was performed to measure the efficacy of the test compound. Increase in paw withdrawal threshold (PWT) by Compound 1 in comparison with vehicle is shown in Figure 8 and is indicative of Compound 1 exhibiting therapeutic efficacy in the MIA model of osteoarthritis.
Example 12: Powder X-ray Diffraction.
Powder X-ray Diffraction (PXRD) data were collected on an X'Pert PRO MPD powder diffractometer (PANalytical, Inc.) with a Cu source set at 45 kV and 40 mA, Cu(Ka) radiation and an X'Celerator detector. Samples were added to the sample holder and smoothed flat with a spatula and weigh paper. With the samples spinning, X-ray diffractograms were obtained by a 12-min scan over the 2-theta range 5-40 °2Θ. Diffraction data were viewed and analyzed with the X'Pert Data Viewer Software, version 1.0a and X'Pert HighScore Software, version 1.0b.
The PXRD profile for:
1) the crystalline form of the Compound of Formula (He) is shown in Figure 1;
2) the crystalline form of the Compound of Formula (Ilf) is shown in Figure 2;
3) the crystalline form of the Compound of Formula (Ilg) is shown in Figure 3; and
4) the anhydrous crystalline form of Compound 1 is shown in Figure 4 and Figure 7.
Example 13: Differential Scanning Calorimetry.
Differential scanning calorimetry (DSC) study was conducted using a TA Instruments, Q2000 at a heating rate 10°C/min. The instrument was calibrated for temperature and energy using the melting point and enthalpy of fusion of an indium standard. Thermal events (desolvation, melting, etc.) were evaluated using Universal Analysis 2000 software, version 4. ID, Build 4.1.0.16.
The differential scanning calorimetry profile for the anhydrous crystalline form of Compound 1 is shown in Figure 5.
Example 14: Thermal Gravimetric Analysis.
Thermogravimetric analysis (TGA) was conducted using a TA Instruments TGA Q500 or Q5000 at a heating rate 10°C/min. The instrument was calibrated using a standard weight for the balance, and Alumel and Nickel standards for the furnace (Curie point measurements). Thermal events such as weight-loss are calculated using the Universal Analysis 2000 software, version 4. ID, Build 4.1.0.16.
The thermogravimetric profile for the anhydrous crystalline form of Compound 1 is shown in Figure 5.
Example 15: Dynamic Moisture-Sorption Analysis.
A dynamic moisture-sorption (DMS) study was conducted using a dynamic moisture-sorption analyzer, VTI Corporation, SGA-100. The instrument was calibrated using polyvinyl pyrrolidone (PVP) and NaCl. Samples were prepared for DMS analysis by placing 5 mg to 20 mg of a sample in a tared sample holder. The sample was placed on the hang-down wire of the VTI balance. A drying step was run, typically at 40°C and 0.5-1% RH for 1 h. The isotherm temperature is 25°C. Defined % RH holds typically ranged from 10% RH to 90% RH, with intervals of 10 to 20% RH. A% weight change smaller than 0.010% over 10 min, or up to 2 h, whichever occurred first, was required before continuing to the next % RH hold. The water content of the sample equilibrated as described above was determined at each % RH hold.
The DMS profile (adsorption/desorption isotherm) for the anhydrous crystalline form of
Compound 1 is shown in Figure 7. The corresponding data in tabular form is provided in Table 12.
Table 12
Figure imgf000088_0002
Example 16: Analytical characterization of (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a- tetrahydro-l//-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((5)-l-hydroxymethyl-2,2- dimethyl-propyl)-amide (Compound 1, Formula (I)).
Three separate batches of Compound 1 were manufactured in compliance with applicable cGMPs for GLP and/or R&D use consisting of 1.23 kg, 1.68 kg, and 1.85 kg quantities. Each batch was analyzed. Certain data for each batch is shown below in Table 13.
Table 13
Figure imgf000088_0001
Batch 1 Batch 2 Batch 3
TEST
(1.23 kg) (1.68 kg) (1.85 kg)
Methanol (residual solvent) < 299 ppm (QL) < 299 ppm (QL) < 299 ppm (QL)
Hexanes (residual solvent) < 29 ppm (QL) < 29 ppm (QL) < 29 ppm (QL)
MTBE (residual solvent) < 5 ppm (QL) < 5 ppm (QL) < 5 ppm (QL)
QL = Quantitation Limit
Figure imgf000089_0001
Example 17: Headspace gas chromatography method for residual solvents presence in intermediates and Compound 1.
Headspace gas chromatography (HSGC) was conducted using an Agilent 6850 or 6890 gas chromatograph (or equivalent) equipped with a flame-ionization detector (FID) using an Agilent DB- 624 column (30 m x 0.53 mm x 3 μηι, or equivalent).
Gas Chromatography Conditions
Figure imgf000089_0002
Headspace Conditions
Figure imgf000089_0003
The GC cycle time in the table is a recommendation. This can vary from instrument to instrument. The GC cycle time should be equal to the GC run time for the GC to equilibrate to initial conditions. The method described herein has been validated, such as, specificity, linearity, accuracy, quantitation limit, detection limit, and solution stability, for methanol, ethanol, acetone, acetonitrile, methyl ieri-butyl ether, and ethyl acetate. The method has certain limitations for the isomers of hexanes, for example, the separation of MTBE could not be achieved from one of the isomers of hexanes. Hexanes contained 5 different isomers labeled Hexane A to Hexane E. Hexane C is the second most abundant isomer of hexanes and a similar retention time as MTBE.
Samples were prepared using 100 mg in 1.00 mL DMSO and were used as the basis for all ppm concentrations using 20 mL borosilicate glass headspace vials with silicone/PTFE septa. The method was based on the published headspace GC method (e.g., Dai L, et. al. (2010) LCGC North America, 28 (1), 73-84).
The retention times (tn minutes) and relative retention times versus DMSO for a certain list of solvents were determined and are shown with the quantitation for each.
Figure imgf000090_0002
Figure imgf000090_0001
The above cited method could be used for the presence of residue methylene chloride.
Those skilled in the art will recognize that various modifications, additions, substitutions, and variations to the illustrative examples set forth herein can be made without departing from the spirit of the invention and are, therefore, considered within the scope of the invention.

Claims

We claim:
1. A process for preparing (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000091_0001
comprising the steps of:
a) reacting (lS,5R)-bicyclo[3.1. of Formula (lie):
Figure imgf000091_0002
(He) .
with dimethyl oxalate in the presence of sodium methoxide and methanol to form one intermediates;
b) condensing said one or more intermediates with 2-hydrazinyl pyrazine (lid):
,NHNH2
N
(lid)
or an acid salt thereof; in the presence of a condensing-step solvent and optionally a condensing-step acid, provided that at least one of said acid salt or said condensing-step acid is present, to form (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i- cyclopropa[4,5]cyclopenta[l,2-c of Formula (He):
Figure imgf000091_0003
c) hydrolyzing said compound of Formula (He) in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000092_0001
(Ilf)
d) oxidizing said compound of Formula (Ilf) in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form 3-((4aS,5aS)-3- carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l-yl)pyrazine 1- oxide of Formula (Ilg):
Figure imgf000092_0002
(Ug) ; and e) coupling said compound of Formula (Ilg) with (S)-ieri-leucinol of Formula (Ilh) or a salt thereof:
Figure imgf000092_0003
(Ilh)
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
A process for preparing (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000093_0001
comprising the steps of:
a) hydrolyzing (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro
cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000093_0002
(He)
in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form sodium (4alS',5alSr)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole- 3-carboxylate of Formula (Ilf):
Figure imgf000093_0003
b) oxidizing said compound of Formula (Ilf) in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form
Figure imgf000093_0004
carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1- oxide of Formula (Ilg):
Figure imgf000093_0005
; and c) coupling said compound of Formula (Ilg) with (S)-ieri-leucinol of Formula (Ilh) or a salt thereof:
Figure imgf000094_0001
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
3. A process for preparing (laS,5a¾-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000094_0002
comprising the steps of:
a) oxidizing sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000094_0003
(Ilf)
in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form 3-((4alS',5alS')-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000095_0001
(Ilg) ; and b) coupling said compound of Formula (Ilg) with (S)-ieri-leucinol of Formula (Ilh) or a salt thereof:
Figure imgf000095_0002
(Ilh)
in the presence of a coupling-step agent, a coupling-step base, and a coupling-step solvent to form (lalS',5alS')-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I). 4. A process for preparing (laS,5a¾-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I):
Figure imgf000095_0003
comprising the step:
coupling 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro- IH- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000096_0001
(iig)
with (S)-ieri-leucinol of Formula (Ilh):
Figure imgf000096_0002
(Ilh)
in the presence of a coupling-step agent, a coupling-step base, and acetonitrile to form
(lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾- l-hydroxymethyl-2,2-dimefhyl-propyl)-amide of Formula (I).
5. The process according to claim 4, further comprising the step of mixing 3-((4aS,5aS)-3- carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol- l-yl)pyrazine 1- oxide, said coupling-step base, and acetonitrile together to form a first-coupling mixture, wherein said first-coupling mixture is at a temperature of about -10°C to about 15°C.
6. The process according to claim 4 or 5, further comprising the step of adding said coupling-step agent to said first-coupling mixture to form a second-coupling mixture, wherein after addition of said coupling-step agent the temperature of said second-coupling mixture is maintained at a temperature of about -10°C to about 15°C.
7. The process according to any one of claims 4 to 6, wherein (S)-ieri-leucinol is added
substantially all at once.
8. The process according to any one of claims 4 to 7, wherein said coupling-step agent is a
tetramethyluronium-based coupling agent. 9. The process according to any one of claims 4 to 8, wherein said coupling-step agent is 2-(7-aza- 1/i-benzotriazole- 1 -yl)- 1 , 1,3,3-tetramethyluronium hexafluorophosphate (HATU).
10. The process according to any one of claims 4 to 9, wherein said coupling-step base comprises a tertiary amine.
11. The process according to any one of claims 4 to 10, wherein said coupling-step base is triefhylamine.
12. The process according to any one of claims 4 to 1 1, wherein said coupling further comprising the step of isolating (laS,5aS)-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro-l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide from said second-coupling mixture.
13. The process according to any one of claims 4 to 12, wherein (laS,5aS)-2-(4-oxy-pyrazin-2-yl)- la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l- hydroxymethyl-2,2-dimethyl-propyl)-amide is crystalline.
14. The process according to any one of claims 4 to 13, further comprising the step of formulating (lalS',5alSr)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza-cyclopropa[a]pentalene-4- carboxylic acid ((¾-l-hydroxymethyl-2,2-dimefhyl-propyl)-amide to form a pharmaceutical composition.
15. The process according to claim 14, wherein said formulating comprises admixing
Figure imgf000097_0001
(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro-l/i-2,3-diaza-cyclopropa[a]pentalene-4-carboxylic acid ((S)-l-hydroxymethyl-2,2-dimethyl-propyl)-amide with a pharmaceutical excipient.
16. A process for preparing (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000097_0002
(lie)
comprising the steps of:
a) reacting (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (lie):
Figure imgf000097_0003
(lie)
with dimethyl oxalate in the presence of sodium methoxide and methanol to form one or more intermediates; and b) condensing said one or more intermediates with 2-hydrazinyl pyrazine (lid):
Figure imgf000098_0001
or an acid salt thereof; in the presence of a condensing-step solvent and optionally a condensing- step acid, provided that at least one of said acid salt or said condensing-step acid is present, to form (4alS',5aS')-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate.
The process according to claim 16, wherein said reacting in Step a) further comprising the step of mixing (lS,5R)-bicyclo[3.1.0]hexan-2-one of Formula (lie), dimethyl oxalate, and methanol together to form a first-reacting mixture, wherein said first-reacting mixture is at a temperature of about 15°C to about 30°C.
The process according to claim 17, further comprising adding sodium methoxide to said first- reacting mixture to form a second-reacting mixture.
The process according to claim 18, wherein sodium methoxide is added to said first-reacting mixture at a rate so as to maintain the temperature of said second-reacting mixture at about 10°C to about 40°C.
The process according to claim 18 or 19, wherein after complete addition of sodium methoxide, said second-reacting mixture is maintained at a temperature of about 30°C to about 45°C.
The process according to any one of claims 16 to 20, wherein sodium methoxide is about 25 weight percent in methanol solution.
The process according to any one of claims 16 to 21, wherein the molar ratio of (IS,5R)- bicyclo[3.1.0]hexan-2-one, dimethyl oxalate, and sodium methoxide is about 1 : 1.05: 1.05 to about 1 :2.4:2.8.
The process according to any one of claims 16 to 22, wherein said condensing-step solvent comprises methanol.
The process according to any one of claims 16 to 23, wherein said condensing is carried out in the presence of aqueous HC1. The process according to any one of claims 16 to 24, wherein said condensing in Step b) further comprising the step of mixing said one or more intermediates and said condensing-step solvent together to form a first-condensing mixture.
The process according to claim 25, further comprising the step of adding said condensing-step acid to said first-condensing mixture to form a second-condensing mixture.
27. The process according to claim 26, wherein said condensing-step acid is added at a rate so as to maintain said second-condensing mixture at a temperature of about 20°C to about 50°C.
28. The process according to claim 26 or 27, further comprising the step of isolating (4aS,5aS)- methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from said second-condensing mixture.
29. The process according to any one of claims 16 to 28, wherein (4aS,5aS)-methyl l-(pyrazin-2- yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a purity of about 94% or greater as determined by HPLC.
A process for preparing sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000099_0001
(Ilf)
comprising the step:
hydrolyzing (4aS,5aS)-methyl 1 -(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- IH- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (He):
Figure imgf000099_0002
(He) in the presence of aqueous sodium hydroxide and a hydrolyzing-step solvent to form said sodium (4alS',5alS')-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2- c]pyrazole-3-carboxylate.
31. The process according to claim 30, wherein said hydrolyzing further comprising the step of mixing (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate and said hydrolyzing-step solvent together to form a first-hydrolyzing mixture, wherein said first-hydrolyzing mixture is at a temperature of about 20°C to about 45°C.
32. The process according to claim 30 or 31, wherein said hydrolyzing-step solvent comprises methanol.
33. The process according to claim 31 or 32, further comprising the step of adding said sodium hydroxide to said first-hydrolyzing mixture to form a second-hydrolyzing mixture, wherein after the addition of said sodium hydroxide the temperature of said second-hydrolyzing mixture is maintained at about 35°C to about 55°C.
34. The process according to claim 33, further comprising the step of isolating sodium (4aS,5aS)-l- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate from said second-hydrolyzing mixture.
35. The process according to any one of claims 30 to 34, wherein sodium (4alS',5aS')-l-(pyrazin-2- yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
36. The process according to claim 35, wherein crystalline sodium (4alS',5alSr)-l-(pyrazin-2-yl)- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate has a water content of about 5% to about 8% as determined by thermogravimetric analysis and/or Karl Fisher titration.
37. A process for preparing 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilg):
Figure imgf000101_0001
(iig)
comprising the step:
oxidizing sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000101_0002
(Ilf)
in the presence of hydrogen peroxide and an oxidizing-step solvent comprising formic acid and trifluoroacetic acid to form said 3-((4aS,5aS)-3-carboxy-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5] cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide. 38. The process according to claim 37, wherein said oxidizing further comprising the step of mixing sodium (4aS,5a¾- l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate and said oxidizing-step solvent together to form a first-oxidizing mixture, wherein said first-oxidizing mixture is at a temperature of about 45°C to about 55°C.
39. The process according to claim 37 or 38, wherein the molar ratio of sodium (4aS,5aS)-l- (pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3- carboxylate, trifluoroacetic acid, and hydrogen peroxide is about 1 : 11 :5 to about 1 : 10:3.5. 40. The process according to claim 38 or 39, further comprising the step of adding hydrogen peroxide to said first-oxidizing mixture to form a second-oxidizing mixture, wherein hydrogen peroxide is added to said first-oxidizing mixture in multiple portions.
41. The process according to claim 40, wherein hydrogen peroxide is added to said first-oxidizing mixture in multiple portions at a rate so as to maintain said second-oxidizing mixture at a temperature of about 50°C to about 60°C. The process according to claim 40 or 41, further comprising the step of isolating
Figure imgf000102_0001
3-carboxy-4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1- oxide from said second-oxidizing mixture.
43. The process according to any one of claims 37 to 42, wherein 3-((4alS',5alS')-3-carboxy- 4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide is crystalline. 44. A compound selected from the compound of Formula (He) and salts, solvates, and hydrates
Figure imgf000102_0002
The compound according to claim 44, wherein (4aS,5aS)-methyl l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate is crystalline.
46. The crystalline form according to claim 45, wherein the crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 10.5° + 0.2°, 11.0° + 0.2°, 11.9° + 0.2°, 17.1° + 0.2°, 17.9° + 0.2°, and 25.3° + 0.2°.
A crystalline form of sodium (4aS,5aS)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000102_0003
(Ilf) The crystalline form according to claim 47, wherein said crystalline form has a powder X-ray diffraction pattern comprising peaks, in terms of 26», at 8.3° + 0.2°, 10.9° + 0.2°, 1 1.1° + 0.2°, 21.6° + 0.2°, 25.3° + 0.2°, and 29.8° + 0.2°.
A compound that is isolated sodium (4aS,5a¾-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxylate of Formula (Ilf):
Figure imgf000103_0001
A composition comprising:
a. (laS,5a¾-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide (anhydrous crystalline form); and
b. a compound selected from:
3-((4a.S,,5alS,)-3-(((lS,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2- yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide of Formula (Ilia):
Figure imgf000103_0002
(Ilia) and
(4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb):
Figure imgf000104_0001
51. The composition according to claim 50, wherein the composition is substantially free of methylene chloride.
52. The composition according to claim 50 or 51, wherein the compound is 3-((435,5α?)-3-(((¾- 3,3-dimethyl-l-(((4aS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1-oxide of Formula (Ilia).
53. The composition according to claim 50 or 51, wherein the compound is (4alS',5alS')-Ai-((lS')-l- hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-l/i- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb).
54. The composition according to any one of claims 50 to 53, wherein the composition further comprises a pharmaceutically acceptable carrier.
55. A pharmaceutical composition comprising:
a. (laS,5aS)-2-(4-oxy-pyrazin-2-yl)-la,2,5,5a-tetrahydro- l/i-2,3-diaza- cyclopropa[a]pentalene-4-carboxylic acid ((S)- 1 -hydroxymethyl-2,2-dimethyl-propyl)-amide (anhydrous crystalline form);
b. a compound selected from:
3-((4aS,,5aS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a- tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2- yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1-oxide of Formula (Ilia):
Figure imgf000105_0001
(Ilia) ; and
(4aS,5aS)-N-((S)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a- tetrahydro-lH-cyclopropa[4,5 -3-carboxamide of Formula (Illb):
Figure imgf000105_0002
and
c. a pharmaceutically acceptable carrier.
56. The pharmaceutical composition according to claim 55, wherein the composition is
substantially free of methylene chloride.
57. The pharmaceutical composition according to claim 55 or 56, wherein the compound is 3- ((4a5,5a¾-3-(((¾-3,3-dimethyl-l-(((4alS',5aS)-l-(l-oxidopyrazin-3-yl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2-yl)carbamoyl)- 4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazol-l-yl)pyrazine 1 -oxide of Formula (Ilia).
The pharmaceutical composition according to claim 55 or 56, wherein the compound is (4a,S,,5alS,)-iV-((lS,)-l-hydroxy-3,3-dimethylbutan-2-yl)-l-(pyrazin-2-yl)-4,4a,5,5a-tetrahydro-m- cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carboxamide of Formula (Illb).
The pharmaceutical composition according to any one of claims 55 to 58, wherein the pharmaceutical composition is suitable for oral, rectal, nasal, topical, buccal, sub-lingual, vaginal, parenteral, intramuscular, sub-cutaneous, or intravenous administration, or in a form suitable for administration by inhalation, insufflation, or by a transdermal patch.
60. The pharmaceutical composition according to any one of claims 55 to 59, wherein the pharmaceutical composition is suitable for oral administration.
61. A method for the treatment of a CB2 receptor-mediated disorder in an individual, comprising administering to said individual in need thereof, a therapeutically effective amount of a pharmaceutical composition according to any one of claims 55 to 60.
62. A method for the treatment of pain in an individual, comprising administering to said individual in need thereof, a therapeutically effective amount of a pharmaceutical composition according to any one of claims 55 to 60.
63. A method for the treatment of pain associated with osteoarthritis, neuropathic pain, or acute post-operative pain in an individual, comprising administering to said individual in need thereof, a therapeutically effective amount of a pharmaceutical composition according to any one of claims 55 to 60.
64. A method for the treatment of osteoarthritis in an individual, comprising administering to said individual in need thereof, a therapeutically effective amount of a pharmaceutical composition according to any one of claims 55 to 60.
65. Use of a pharmaceutical composition according to any one of claims 55 to 60, in the treatment of a CB2 receptor-mediated disorder.
66. Use of a pharmaceutical composition according to any one of claims 55 to 60, in the treatment of pain.
67. Use of a pharmaceutical composition according to any one of claims 55 to 60, in the treatment of pain associated with osteoarthritis, neuropathic pain, or acute post-operative pain. 68. Use of a pharmaceutical composition according to any one of claims 55 to 60, in the treatment of osteoarthritis.
69. A pharmaceutical composition according to any one of claims 55 to 60, for use in a method of treatment of the human or animal body by therapy.
70. A pharmaceutical composition according to any one of claims 55 to 60, for use in a method of treatment of a CB2 receptor-mediated disorder.
71. A pharmaceutical composition according to any one of claims 55 to 60, for use in a method of treatment of pain.
72. A pharmaceutical composition according to any one of claims 55 to 60, for use in a method of treatment of pain associated with osteoarthritis, neuropathic pain, or acute post-operative pain.
73. A pharmaceutical composition according to any one of claims 55 to 60, for use in a method of treatment of osteoarthritis.
74. A compound that is 3-((4alS,,5alS,)-3-(((.S,)-3,3-dimethyl-l-(((4alS,,5aS)-l-(l-oxidopyrazin-3-yl)- 4,4a,5,5a-tetrahydro-l/i-cyclopropa[4,5]cyclopenta[l,2-c]pyrazole-3-carbonyl)oxy)butan-2- yl)carbamoyl)-4,4a,5,5a-tetrahydro- l/i-cyclopropa[4,5]cyclopenta[ 1 ,2-c]pyrazol- 1 -yl)pyrazine 1 -oxide.
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