AU2005272074B2 - Crystal forms of (3r)-1-(2-methylalanyl-d-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide - Google Patents

Abstract

Crystalline polymorphs of (3 R)-1-(2-methylalanyl-D-tryptophyl)-3­ (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide which are useful as pharmaceutical agents are disclosed. Methods of production and isolation of these polymorphs and pharmaceutical compositions which include these polymorphs and pharmaceutical methods of treatment are also disclosed. The crystalline polymorphs of the present invention are useful as they act directly on the pituitary gland cells to release growth hormone.

Description

WO 2006/016995 PCT/US2005/022408 CRYSTAL FORMS OF (3R)-1-(2-METHYLALANYL-D-TRYPTOPHYL)-3 (PHENYLMETHYL)-3-PIPERIDINECARBOXYLIC ACID 1,2,2 TRIMETHYLHYDRAZIDE RELATED APPLICATION 5 This application claims the benefit of U.S. Provisional Application No. 60/583,757, filed on June 29, 2004. The entire teachings of the above application are incorporated herein by reference. BACKGROUND OF THF INVENTION Growth hormone is a major participant in the control of several complex 10 physiologic processes, including growth and metabolism. Growth hormone is known to have a number of effects on metabolic processes, e.g., stimulation of protein synthesis and free fatty acid mobilization and to cause a switch in energy metabolism from carbohydrate to fatty acid metabolism. Deficiency in growth hormone can result in a number of severe medical disorders, e.g., dwarfism. 15 The release of growth hormone from the pituitary is controlled, directly or indirectly, by number of hormones and neurotransmitters. Growth hormone release can be stimulated by growth hormone releasing hormone (GHRH) and inhibited by somatostatin. In both cases the hormones are released from the hypothalamus but their action is mediated primarily via specific receptors located in the pituitary. 20 Other compounds which stimulate the release of growth hormone from the pituitary have also been described. For example, arginine, L-3,4-dihydroxyphenylalanine (1 Dopa), glucagon, vasopressin, PACAP (pituitary adenylyl cyclase activating peptide), muscarinic receptor agonists and a synthetic hexapeptide, GHRP (growth hormone releasing peptide) release endogenous growth hormone either by a direct 25 effect on the pituitary or by affecting the release of GHRH and/or somatostatin from the hypothalamus. The use of certain compounds for increasing the levels of growth hormone in mammals has previously been proposed. For example, U.S. Patents 6,303,620 and 6,576,648 (the entire contents of which are incorporated herein by reference), 30 disclose a compound: (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- -2/1 piperidinecarboxylic acid 1,2,2-trimethylhydrazide, having the following chemical structure: HN 0

CH

3

H

3 CN

H

2 N N ' N CH 3 N H CH3

CH

3 0 which acts directly on the pituitary cells under normal experimental conditions in 5 vitro to release growth hormone therefrom. This growth hormone releasing compound can be utilized in vitro as a unique research tool for understanding, inter alia, how growth hormone secretion is regulated at the pituitary level. Moreover, this growth hormone releasing compound can also be administered in vivo to a mammal to increase endogenous growth hormone release. 10 Each document, reference, patent application or patent cited in this text is expressly incorporated herein in their entirety by reference, which means that it should be read and considered by the reader as part of this text. That the document, reference, patent application or patent cited in this text is not repeated in this text is merely for reasons of conciseness. 15 Reference to cited material or information contained in the text should not be understood as a concession that the material or information was part of the common general knowledge or was known in Australia or any other country. SUMMARY OF THE INVENTION Throughout this specification, unless the context requires otherwise, the 20 word "comprise" or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers - 2/2 It has been found that (3R)-1 -(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide can be crystallized. At least four crystalline forms with purities of greater than 99 area 5 percent have been produced. These crystalline forms of (3R)-1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide are expected to be useful as pharmaceutical compositions, which can be used to increase endogenous growth hormone release. In one embodiment, the present invention is crystalline (3R)-1 -(2 10 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide.

WO 2006/016995 PCT/US2005/022408 -3 In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group 5 consisting of about: 10.1, 11.1, 17.6, 20.0 and 20.8. In yet another embodiment, the present invention is a crystalline composition, having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group listed above and at least one or at least two 20 values measured using Cu Ka radiation are selected from the group consisting of about: 9.6, 10 17.3, 18.2, 22.1 and 23.5. In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group 15 consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3. In yet another embodiment, the present invention is a crystalline composition, having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group listed above and at least one or at least two 20 values measured using Cu Ka radiation are selected from the group consisting about: 9.3, 20 23.3, 20.4, 22.9, and 23.5. In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group 25 consisting of about 19.2, 20.1, 23.0, 26.2, and 27.0. In yet another embodiment, the present invention is a crystalline composition, having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group listed above and at least one or at least two 20 values measured using Cu Ka radiation are selected from the group consisting about: 16.3, 30 21.4, 24.0, 29.8, and 31.5. In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- -4 In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu Ka radiation are selected from the group 5 consisting of about 10.2, 11.2, 18.7, 20.6, and 23.4. In yet another embodiment, the present invention is a crystalline composition, having an X-ray powder diffraction pattern wherein at least one or at least two 20 values measured using Cu K 0 radiation are selected from the group listed above and at least one or at least two 20 values measured using Cu K 0 radiation are selected from the group consisting of about 9.9, 10 13.8, 14.3, 16.7, and 19.8. In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu Ka radiation is selected from the group consisting of: 15 10.1, 17.0, 20.6, and 23.0, and optionally a second additional 20 value measured using Cu Ka, radiation is selected from the group consisting of about: 17.6, 19.2, 23.0, 23.4, 26.2, and 33.3 or optionally a second additional 20 value measured using Cu Ka radiation is selected from the group consisting of: 9.5, 16.7, 17.5, 17.9, 20.0, 21.5, 23.5, 23.9, and 27.5. 20 Further the present invention is directed to a process for preparing crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide. The method comprises combining (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide with a solvent. Precipitating the crystals from the solvent 25 wherein the solvent is selected from the group consisting of alcohols, water, and mixtures thereof and isolating the crystals. In one embodiment, the solvent is selected from the group consisting of: water, methanol, ethanol, n-propanol, isopropanol, butanol, and mixtures thereof. In another embodiment, the solvent is a mixture of methanol and water. In a further embodiment, the solvent includes 30 between 40% v/v methanol and 60 % v/v methanol. In a still further embodiment, the present invention is - 5/1 crystalline (3R)-I-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method described above. Further the present invention is a process for preparing crystalline (3R)-1-(2 5 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide. The method comprises the steps of combining {1 -[(1 R)-2 ](3R)-3-Benzyl-3-(N,N',N'-trimethylhydrazinocarbonyl)piperidin-1-yl-]-1-(1 H indol-3 -ylmethyl)-2-oxo-ethylcarbamoyl] -1 -methylethyl } carbamic acid tert-butyl ester with a solvent wherein the solvent is selected from the group consisting of 10 alcohols, water, and mixtures thereof and an acid. The method further comprises neutralizing the mixture, precipitating the crystals from the solvent, and isolating the crystals. In one embodiment the solvent is methanol, and the mixture is neutralized with a mixture of potassium hydroxide and water. In another embodiment the acid is methanesulfonic acid. In yet another embodiment the present invention is a 15 crystalline (3R)- 1 -(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method described above. Further, the present invention is directed to a pharmaceutical composition comprising crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 20 piperidinecarboxylic acid 1,2,2-trimethylhydrazide and at least one pharmaceutically acceptable carrier or diluent. Further the present invention is directed to a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 25 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide. Further, the present invention is directed to a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 30 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu Ka radiation is selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0 and 20.8.

Further, the present invention is directed to a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 5 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu Ka radiation is selected from the group consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3. Further, the present invention is directed to a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising 10 administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu Ka radiation is selected from the group consisting of about: 19.2, 20.1, 23.0, 26.2, and 27.0. 15 Further, the present invention is directed to a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least 20 one 20 value measured using Cu Ka radiation is selected from the group consisting of about: 10.2, 11.2, 18.7, 20.6, and 23.4. Further, the present invention is directed to a method of A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount 25 of crystalline (3R)-I-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu Ka radiation is selected from the group consisting of about: 10.1, 17.0, 20.6, and 23.0. The present invention provides crystalline polymorphic forms of (3R)-1-(2 30 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide, each of which are more stable, and have advantageous manufacturing properties relative to amorphous forms of this compound. Due to the high purity of these crystalline forms, they can fulfill more exacting pharmaceutical regulations and specifications often required for pharmaceutical formulations.

WO 2006/016995 PCT/US2005/022408 -6 BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is a characteristic X-Ray Powder Diffraction (XRPD) pattern for form A (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method 5 described in Example 1 (Vertical axis: Intensity (CPS); Horizontal axis: Diffraction Angle, in Two Theta (degrees). Figure 2 is a characteristic X-Ray Powder Diffraction (XRPD) pattern for form B (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method 10 described in Example 2 (Vertical axis: Intensity (CPS); Horizontal axis: Diffraction Angle, in Two Theta (degrees). Figure 3 is a characteristic X-Ray Powder Diffraction (XRPD) pattern for form D (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method 15 described in Example 5 (Vertical axis: Intensity (CPS); Horizontal axis: Diffraction Angle, in Two Theta (degrees). Figure 4 is a characteristic X-Ray Powder Diffraction (XRPD) pattern for form C (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method 20 described in Example 8 (Vertical axis: Intensity (CPS); Horizontal axis: Diffraction Angle, in Two Theta (degrees). Figure 5 is a characteristic Differential Scanning Calorimetry (DSC) thennogram for form C (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide as described in Example 8. 25 DETAILED DESCRIPTION OF THE INVENTION The foregoing and other objects, features and advantages of the invention will be apparent from the following more particular description of preferred embodiments of the invention, as illustrated in the accompanying drawings in which like reference characters refer to the same parts throughout the different views. The 30 drawings are not necessarily to scale, emphasis instead being placed upon illustrating the principles of the invention.

WO 2006/016995 PCT/US2005/022408 -7 The present invention is directed to crystalline compounds of the following Structural Formula: HN 0 CH 3 0

H

3 C O

H

2 N N N CH 3 HI

CH

3

CH

3 0/ 5 The chemical name of Formula I is 1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trinethylhydrazide which is the equivalent of 2-amino-N-[(1R)-2-[-3-benzyl-3-(NN',N' trimethylhydrazinocarbonyl)piperidin-1-yl]-l-((1H-indol-3-yl)methyl)-2-oxoethyl] 10 2-methylpropionamide. In a preferred embodiment, the compound of Formula I has the (R) configuration at the chiral carbon designated by the asterisk (*) in Formula I. The chemical name of the compound of Formula I having the (R) configuration at the designated chiral carbon is: (3R)-1 -(2-methylalanyl-D-tryptophyl)-3 15 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide: WO 2006/016995 PCT/US2005/022408 -8 HN 0

CH

3

H

3 C N

H

2 N N N CH 3 N H~

OH

3

CH

3 0 II It is to be understood that crystalline (3R)-1 -(2-methylalanyl-D-tryptophyl) 5 3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide includes hydrates thereof, unless specifically excluded. In a preferred embodiment crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide is in the form of a dihydrate. As used herein a "dihydrate" is when each molecule of (3R)-1 -(2-methylalanyl-D 10 tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide is associated with about two molecules of water. As used herein a "monohydrate" is when each molecule of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide is associated with about one molecule of water. 15 As used herein a "crystalline form" is a solid substance having a highly regular chemical structure. When a compound recrystallizes from a solution or slurry, it may crystallize with different spatial lattice arrangements, a property referred to as "polymorphism," with the different crystal forms individually being referred to as a "polymorph." While polymorphs of a given substance have the same 20 chemical composition, they may differ from each other with respect to one or more physical properties, such as solubility and dissociation, true density, melting point, crystal shape, compaction behavior, flow properties, and/or solid state stability.

WO 2006/016995 PCT/US2005/022408 -9 The polymorphic behavior of drugs can be of crucial importance in pharmacy and pharmacology. The differences in physical properties exhibited by polymorphs affect pharmaceutical parameters such as storage stability, compressibility and density (important in formulation and product manufacturing), 5 and dissolution rates (an important factor in determining bio-availability). Differences in stability can result from changes in chemical reactivity (e.g. differential oxidation, such that a dosage form discolors more rapidly when it is one polymorph than when it is another polymorph) or mechanical changes (e.g. tablets crumble on storage as a kinetically favored polymorph converts to 10 thermodynamically more stable polymorph) or both (e.g. tablets of one polymorph are more susceptible to breakdown at high humidity). In addition, the physical properties of the crystal may be important in processing: for example, one polymorph might be more likely to form solvates or might be difficult to filter and wash free of impurities (i.e. particle shape and size distribution might be different 15 between one polymorph relative to other). In certain specific embodiments, the present invention is directed to crystalline polymorphs of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl) 3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide. Four such crystalline polymorphs described herein are designated as crystalline forms A, B, C, and D, and 20 may be distinguished by their respective X-ray powder diffraction patterns. Crystalline forms A, B, C, and D of the present invention may exist in anhydrous forms as well as hydrated and solvated forms, all of which are intended to be encompassed within the scope of the present invention. In one embodiment, X-ray powder diffraction patterns of the polymorphs are 25 measured on a Shimadzu XRD-6000 X-ray powder diffractometer using Cu Ka radiation. The instrument is equipped with a fine focus X-ray tube. The tube voltage and amperage ware set to 40 kV and 40 mA, respectively. The divergence and scattering slits are set at 10 and the receiving slit is set at 0.15 mm. Diffracted radiation is detected by Nal scintillation detector. A theta-two theta continuous scan 30 at 3 '/min (0.4 sec/0.02' step) from 2.5 to 40 '20 is used. A silicon standard is analyzed to check the instrument alignment. Samples were prepared for analysis by placing them in an aluminium holder with silicon insert. Some shifting in peak WO 2006/016995 PCT/US2005/022408 - 10 positions can occur due to differences in sample height. As defined herein, the 2 theta position has an error of +/- 0.20. Further, some shifting in peak positions can occur due to differences in the sample, such as, for example, particle size and crystal density. 5 In one specific embodiment, the present invention is directed to crystalline form A of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide, produced by slow evaporation from a solution thereof as described in Example 1, characterized by the following X ray powder diffraction pattern (also presented in Figure 1) expressed in terms of 20, 10 d-spacings and relative intensities: Peak 2Theta d Relative Intensity No. (deg) (A) 1 2.6 33.7 3 2 2.9 30.6 4 3 4.0 22.34 3 4 4.7 18.8 3 5 7.1 12.4 3 6 9.3 9.5 8 7 16.1 5.5 4 8 16.6 5.3 4 9 17.0 5.2 100 10 17.6 5.0 6 11 19.4 4.6 9 12 20.1 4.4 3 13 20.4 4.4 8 14 20.6 4.3 4 15 21.5 4.1 10 16 22.9 3.9 7 17 23.1 3.9 6 WO 2006/016995 PCT/US2005/022408 -11 18 23.3 3.8 6 19 23.5 3.8 9 20 24.1 3.7 4 21 25.4 3.5 4 22 26.2 3.4 37 23 26.5 3.4 4 24 27.1 3.3 6 25 27.4 3.3 5 26 29.8 3.0 6 27 30.0 3.0 4 28 30.7 2.9 4 29 30.9 2.9 3 30 31.5 2.8 5 31 33.3 2.7 18 32 34.7 2.6 4 33 35.1 2.6 3 34 37.2 2.4 4 35 37.3 2.4 5 36 37.5 2.4 4 37 37.9 2.4 3 38 38.1 2.4 3 39 38.4 2.3 4 40 38.8 2.3 4 Table 1: Peak Data List for crystalline form A of (3R)-1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide 5 In another embodiment, the present invention is directed to crystalline form B of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide, produced by slow evaporation from a solution thereof as described in Example 2, characterized by the following X- WO 2006/016995 PCT/US2005/022408 -12 ray powder diffraction pattern (also presented in Figure 2) expressed in terms of 20, d-spacings and relative intensities: Peak 2Theta D Relative Intensity No. (deg) (A) 1 8.1 10.9 8 2 9.0 9.8 4 3 9.6 9.2 8 4 10.7 8.3 4 5 13.6 6.5 8 6 14.3 6.2 12 7 16.3 5.4 40 8 17.5 5.1 12 9 18.0 4.9 12 10 19.2 4.6 68 11 20.1 4.4 56 12 21.4 4.2 20 13 23.0 3.9 100 14 24.0 3.7 16 15 24.6 3.6 4 16 25.3 3.5 12 17 26.2 3.4 100 18 27.0 3.3 52 19 29.8 3.0 16 20 31.5 2.8 32 21 33.1 2.7 16 22 34.1 2.6 12 23 35.3 2.5 8 24 37.4 2.4 4 25 38.6 2.3 8 WO 2006/016995 PCT/US2005/022408 - 13 Table 2: Peak Data List for crystalline form B of (3R)-1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide 5 In another embodiment, the present invention is directed to crystalline form C of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide, produced by slow evaporation from a solution thereof as described in Example 3. In another embodiment, the present invention is directed to crystalline form 10 D of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide, produced by slow evaporation from a solution thereof as described in Example 5, characterized by the following X ray powder diffraction pattern (also presented in Figure 3) expressed in terms of 20, d-spacings and relative intensities: 15 Peak 2Theta d Relative Intensity No. (deg) (A) 1 9.4 9.4 3 2 9.9 9.0 24 3 10.2 8.7 76 4 10.6 8.3 15 5 11.2 7.9 55 6 11.7 7.6 8 7 13.8 6.4 24 8 14.3 6.2 43 9 14.6 6.1 9 10 15.1 5.9 24 11 16.7 5.3 24 12 17.4 5.1 18 13 17.9 5.0 23 WO 2006/016995 PCT/US2005/022408 -14 14 18.2 4.9 15 15 18.7 4.7 53 16 19.8 4.5 40 17 20.6 4.3 100 18 21.1 4.2 5 19 21.6 4.1 9 20 21.8 4.1 10 21 22.5 4.0 7 22 23.4 3.8 56 23 23.8 3.7 6 24 24.9 3.6 9 25 25.8 3.5 13 26 26.4 3.4 6 27 27.5 3.2 11 28 27.7 3.2 21 29 28.8 3.1 13 30 30.0 3.0 14 31 30.4 3.0 18 32 31.3 2.9 13 33 31.9 2.8 6 34 32.9 2.7 4 35 33.6 2.7 18 36 34.1 2.6 9 37 34.4 2.6 4 38 35.5 2.5 7 39 36.8 2.4 10 40 37.4 2.4 11 Table 3: Peak Data List for crystalline form D of (3R)-1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide WO 2006/016995 PCT/US2005/022408 - 15 Further, the present invention is directed to a process for the preparation of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide which comprises crystallization of amorphous (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 5 piperidinecarboxylic acid 1,2,2-trimethylhydrazide. In one embodiment, the method broadly includes crystallization of a solute from a solvent or slurry under conditions which yield crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide. Thus, for example, crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3 10 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide may be prepared by crystallization of the amorphous form, from a solution or from a slurry thereof in a solvent. In one embodiment, the solution or slurry of amorphous (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide in the solvent is formed by combining the (3R)- 1 -(2 15 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide with the solvent. In another preferred embodiment, a solution of amorphous (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide in the solvent is formed by dissolving the (3R)- 1 -(2 20 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide in the solvent. Suitable solvents are those from which (3R)- 1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide can be crystallized by the methods described herein. Examples of suitable solvents 25 include those selected from the group consisting of alcohols (e.g., methanol, ethanol, n-propanol, isopropanol, butanol), polar organic solvents (e.g., dimethyl sulfoxide and ethylene glycol) and water. In a preferred embodiment, the solvent is selected from the group comprising: methanol, ethanol, n-propanol, isopropanol, butanol, and mixtures 30 thereof. In a more preferred embodiment, the solvent is methanol. In a most preferred embodiment, the solvent is a mixture of methanol and water containing, WO 2006/016995 PCT/US2005/022408 -16 for example, between 5% and 95% v/v methanol, between 20% and 80 % v/v methanol, preferably between 40% and 60% v/v methanol. In another embodiment, crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide can be 5 prepared from an aqueous-based solvent that includes, for example, between 5% and 95% v/v ethanol, between 20% and 80 % v/v ethanol, and preferably between 40% and 60% v/v ethanol. In a further embodiment, crystalline (3R)-1-(2-methylalanyl-D-tryptophyl) 3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide can be 10 prepared from an aqueous-based solvent that includes, for example, between 5% and 95% v/v isopropanol, between 20% and 80 % v/v isopropanol, and preferably between 40% and 60% v/v isopropanol. In one embodiment, the solvent may be heated to an elevated temperature. As used herein an "elevated temperature" is higher than ambient temperature, such 15 as for example, between 40 'C and 100 *C, between 50 'C and 80 *C, or preferably between 65 *C* and 75 *C. In another embodiment, the solvent is heated to an elevated temperature prior to combining of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide with the solvent. In another embodiment, the solvent is heated to an elevated temperature 20 after combining (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide with the solvent. In one embodiment, crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide is precipitated from the solvent. In one embodiment, precipitation is induced by concentration of 25 the solution of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide in the solvent by reduction of the volume of the solvent. The solvent may be reduced, for example, by evaporation of the solvent. In another embodiment, precipitation may be induced by allowing the solution of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 30 piperidinecarboxylic acid 1,2,2-trimethylhydrazide in the solvent to cool. In another embodiment, precipitation may be induced by rapidly cooling the solution, or by slowly cooling the solution. In a specific embodiment, precipitation WO 2006/016995 PCT/US2005/022408 -17 may be induced by slowly cooling the solution to about 20 *C at a rate ranging from about 0.2 *C/hr to about 20 "C/hr, preferably from about 1 'C/hr to about 10 'C/hr. In a preferred embodiment, the solution is cooled from the initial elevated temperature to 60 "C at a rate of 20 "C/h, then from 60 "C at a rate of 5 *C/h, and 5 then from 45 "C to ambient temperature at a rate of 15 *C/h. The temperature and the cooling rate can be appropriately decided depending on the choice of solvent and the volume thereof. For the purpose of this invention ambient temperature is from about 20 *C to about 250C. In one embodiment, the precipitated crystalline (3R)-1-(2-methylalanyl-D 10 tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide is isolated from the solvent. As used herein the term "isolated" encompasses crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide which is separated away from other material, such as the materials contained in the medium in which it was 15 produced. In one embodiment, the isolated crystals are essentially free of chemicals or contaminating solvents from the source from which the crystals were derived or produced. In one embodiment, the crystals are isolated from the solvent by filtration of the solvent. In a preferred embodiment, the crystalline form is isolated from the solvent by filtration of the solvent after precipitation of the crystals. 20 In one embodiment, the isolated crystals are washed with, for example, a methanol/water mixture and dried in an oven. The crystals are preferably dried in an oven, at a temperature of between about ambient temperature and about 70 "C, preferably from about 40 "C to about 60 "C. Preferably, the drying takes place under vacuum. 25 In a further embodiment, "seeds" of the desired polymorph are added to the solvent in order to promote formation of a particular polymorph in the solvent. In one embodiment, the amorphous (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide is added to the solvent, and after cooling has begun the solvent is seeded with crystals of the desired 30 polymorph. Alternatively, the seed crystals can be added once the solvent is saturated with amorphous of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide.

WO 2006/016995 PCT/US2005/022408 - 18 In one embodiment the crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide is formed from 1-[(1R)-2-](3R)-3-Benzyl-3-(N,N',N'-trimethylhydrazinocarbonyl)piperidin-1-yl-] 1 -(1H-indol-3-ylmethyl)-2-oxo-ethylcarbamoyl]- 1 -methylethyl} carbamic acid tert 5 butyl ester. The tert-butoxycarbonyl group on 1-[(1R)-2-](3R)-3-Benzyl-3-(NN',N' trimethylhydrazinocarbonyl)piperidin-1-yl-]-l-(1H-indol-3-ylmethyl)-2-oxo ethylcarbamoyl]- 1 -methylethyl} carbamic acid tert-butyl ester is removed to form the free amine in the presence of an acid. In one embodiment the acid is trifluoroacetic acid, hydrofluoric acid, hydrochloric acid, hydrobromic acid or 10 sulfuric acid. In a preferred embodiment the acid is methanesulfonic acid. Once the tert-butoxycarbonyl group is removed, the mixture is neutralized using a base such as, for example, potassium hydroxide, sodium hydroxide, calcium hydroxide, ammonium hydroxide or potassium carbonate. In a preferred embodiment, the mixture is neutralized with a mixture of water and a base such as, for example, 15 potassium hydroxide, sodium hydroxide, calcium hydroxide, ammonium hydroxide or potassium carbonate. The remainder of the crystallization process is carried out as described above. In one embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 20 trimethylhydrazide produced by the method described above. Variations in the methods of crystallization may produce variations in the crystals, including, for example, variations in the particle size and crystal density, which may result in variations in the XRPD patterns between crystal forms such as, for example, peak splitting, crystals produced by such methods are also encompassed by the present 25 invention In one embodiment, form C of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide can be produced by the methods described in Example 4 or Example 6, wherein a different XRPD spectrum is generated than by crystallization of form C (3R)-1-(2 30 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide by slow evaporation from a solution thereof as described in Example 3.

WO 2006/016995 PCT/US2005/022408 - 19 "Slow evaporation" as used herein, means evaporation under atmospheric conditions or under an inert atmosphere of a solvent or mixture of solvents from a solution or slurry thereof with the compound to be crystallized. In one embodiment, the present invention is a crystalline form C of (3R)-1 5 (2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide produced by the methods described in Example 4. In one embodiment, the present invention is a crystalline form C of (3R)-1 (2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide produced by the methods described in Example 6. In one 10 embodiment crystalline form C of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide is a monohydrate. In a preferred embodiment, crystalline form C of (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide is a dihydrate 15 In another embodiment, the present invention is directed to crystalline form C dihydrate of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the methods described in Example 8 characterized by the following X-ray powder diffraction pattern (also presented in Figure 4) measured on a Shimadzu XRD-6000 X-ray 20 powder diffractometer using Cu Ka radiation. Samples are placed on Si zero- return micro samples holders. The tube voltage and amperage are set to 40 kV and 40 mA, respectively. The divergence and scattering slits are set at 1 and the receiving slit is set at 0.30 mm. A theta-two theta continuous scan at 2 */min from 3.0 to 45 020, with a sampling pitch of 0.02 deg and a preset time of 0.60 seconds. The X-ray 25 powder diffraction pattern is expressed in terms of 20, d-spacings and relative intensities: Peak 2Theta d Relative Intensity No. (deg) (A) 1 9.6 9.2 8 WO 2006/016995 PCT/US2005/022408 -20 2 10.1 8.8 100 3 10.8 8.2 4 4 11.1 8.0 25 5 13.8 6.4 7 6 14.6 6.1 4 7 15.2 5.8 6 8 16.7 5.3 6 9 17.3 5.1 9 10 17.6 5.0 18 11 18.2 4.9 12 12 20.0 4.4 18 13 20.5 4.3 7 14 20.8 4.3 12 15 22.1 4.0 8 16 23.5 3.8 13 17 25.8 3.5 4 18 27.5 3.2 5 19 30.5 3.0 4 20 35.0 2.6 3 Table 4: Peak Data List for crystalline form C of (3R)-1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide dihydrate. 5 In another embodiment, crystalline form C dihydrate may be characterized by Differential Scanning Calorimetry (DSC) or by any other method known to persons skilled in the art. In one embodiment, the present invention is directed to form C (3R)-1-(2 10 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide dihydrate characterized by differential scanning calorimetry WO 2006/016995 PCT/US2005/022408 -21 analysis (Figure 5) which showed a broad endotherm centered at 68.5 'C, followed by the melt endotherm with onset of 110 'C and peak of 116.8 'C. In another embodiment, the present invention is crystalline (3R)- 1 -(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 5 trimethyihydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0 and 20.8. In yet another embodiment, the present invention is a crystalline composition, having an X-ray powder diffraction pattern wherein at least one, at 10 least two, at least three, at least four or at least five 20 values measured using Cu KQ radiation are selected from the group listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 9.6, 17.3, 18.2, 22.1 and 23.5. In another embodiment, the present invention is crystalline (3R)-1-(2 15 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3. In yet another embodiment, the present invention is a crystalline 20 composition, having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 9.3, 23.3, 20.4, 22.9, and 23.5. 25 In another embodiment, the present invention is crystalline (3R)-1-(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethyihydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 19.2, 20.1, 23.0, 26.2, 30 and 27.0. In yet another embodiment, the present invention is a crystalline composition, having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu K, WO 2006/016995 PCT/US2005/022408 - 22 radiation are selected from the group listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 16.3, 21.4, 24.0, 29.8, and 31.5. In another embodiment, the present invention is crystalline (3R)-1-(2 5 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethyihydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about 10.2, 11.2, 18.7, 20.6, and 23.4. In yet another embodiment, the present invention is a crystalline 10 composition, having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 9.9, 13.8, 14.3, 16.7, and 19.8. 15 In another embodiment, the present invention is crystalline (3R)- 1 -(2 methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2 trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, or at least four 20 values measured using Cu Ka radiation is selected from the group consisting of about: 10.1, 17.0, 20.6, and 23.0, and 20 optionally at least two, at least three, or at least four additional 20 values measured using Cu Ka radiation is selected from the group consisting: 17.6, 19.2, 23.0, 23.4, 26.2, and 33.3, or optionally at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, or at least nine additional 20 value measured using Cu Ka radiation is selected from the group consisting of about: 9.5, 16.7, 17.5, 25 17.9, 20.0, 21.5, 23.5, 23.9, and 27.5. Further, the crystalline forms of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide of the present invention are useful as pharmaceutical compositions. In one embodiment, the present invention is a pharmaceutical composition 30 comprising crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide as described herein and at least one pharmaceutically acceptable carrier or diluent.

WO 2006/016995 PCT/US2005/022408 - 23 Pharmaceutical compositions containing a polymorph of the present invention may be prepared by conventional techniques, as described in U.S. Patent 6,576,648, the entire contents of which are incorporated herein by reference. In another embodiment, the present invention relates to a pharmaceutical 5 composition comprising, as an active ingredient, a polymorph of the present invention together with a pharmaceutically acceptable carrier or diluent. The pharmaceutical carrier or diluent employed may be a conventional solid or liquid carrier. The carriers, diluents, preparation, composition, dosage and administration of the pharmaceutical compositions of the present invention are as 10 described in U.S. Patent 6,576,648, the entire contents of which are incorporated herein by reference. The compound of the present invention is expected to possess the ability to release endogenous growth hormone in vivo. The compound may therefore be used in the treatment of conditions which require increased plasma growth hormone 15 levels such as in growth hormone deficient humans or in elderly patients or livestock. Thus, in one embodiment, the present invention is directed to a pharmaceutical composition for stimulating the release of growth hormone from the pituitary of a mammal, the composition comprising, as an active ingredient, a 20 polymorph of the present invention together with a pharmaceutically acceptable carrier or diluent. In one embodiment, the mammal, is a human, canine, murine, feline, bovine, ovine, swine or caprine. In a preferred embodiment, the mammal is a human. Further the present invention is a method of stimulating the release of growth 25 hormone from the pituitary of a mammal in need thereof, the method comprising administering to the mammal a therapeutically effective amount of a polymorph of the present invention. As used herein, "a therapeutically effective amount" refers to an appropriate amount of active ingredient to obtain therapeutic or prophylactic effect and can be 30 determined by standard pharmaceutical procedures in cell cultures or experimental animals.

WO 2006/016995 PCT/US2005/022408 -24 In one embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 5 1,2,2-trimethylhydrazide. In another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 10 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0 and 20.8. In yet another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the 15 method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group 20 listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 9.6, 17.3, 18.2, 22.1 and 23.5. In another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising 25 administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 17.0, 19.4, 21.5, 30 26.2, and 33.3. In yet another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount WO 2006/016995 PCT/US2005/022408 - 25 of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu K. radiation are selected from the group 5 listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu K, radiation are selected from the group consisting of about: 9.3, 23.3, 20.4, 22.9, and 23.5. In another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising 10 administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about: 19.2, 20.1, 23.0, 15 26.2, and 27.0. In yet another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder 20 diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu K, radiation are selected from the group consisting of about: 16.3, 21.4, 24.0, 29.8, and 31.5. 25 In another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least 30 one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group consisting of about 10.2, 11.2, 18.7, 20.6, and 23.4. In yet another embodiment, the present invention is a method of WO 2006/016995 PCT/US2005/022408 - 26 stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder 5 diffraction pattern wherein at least one, at least two, at least three, at least four or at least five 20 values measured using Cu Ka radiation are selected from the group listed above and at least one, at least two, at least three, at least four or at least five 20 values measured using Cu K,, radiation are selected from the group consisting of about: 9.9, 13.8, 14.3, 16.7, and 19.8. 10 In another embodiment, the present invention is a method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R) 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least 15 one, at least two, at least three, or at least four 20 values measured using Cu K" radiation is selected from the group consisting of about: 10.1, 17.0, 20.6, and 23.0, and optionally at least two, at least three, or at least four additional 20 values measured using Cu Ka radiation is selected from the group consisting: 17.6, 19.2, 23.0, 23.4, 26.2, and 33.3, or optionally at least two, at least three, at least four, at 20 least five, at least six, at least seven, at least eight, or at least nine additional 20 value measured using Cu Ka radiation is selected from the group consisting of about: 9.5, 16.7, 17.5, 17.9, 20.0, 21.5, 23.5, 23.9, and 27.5. To those skilled in the art, it is well known that the current and potential uses of growth hormone in humans are varied and multitudinous. Thus, the polymorph 25 of the present invention can be administered for purposes stimulating release of growth hormone from the pituitary and would then have similar effects or uses as growth hormone itself. The polymorphs of the present invention are expected to be useful, for example, for stimulation of growth hormone release in the elderly, prevention of catabolic side effects of glucocorticoids, prevention and treatment of 30 osteoporosis, treatment of chronic fatigue syndrome (CFS), treatment of acute fatigue syndrome and muscle loss following elective surgery, stimulation of the immune system, acceleration of wound healing, accelerating bone fracture repair, WO 2006/016995 PCT/US2005/022408 - 27 accelerating complicated fractures, e.g. distraction osteogenesis, treatment of wasting secondary to fractures, treatment of growth retardation, treating growth retardation resulting from renal failure or insufficiency, treatment of cardiomyopathy, treatment of wasting in connection with chronic liver disease, 5 treatment of thrombocytopenia, treatment of growth retardation in connection with Crohn's disease, treatment of short bowel syndrome, treatment of wasting in connection with chronic obstructive pulmonary disease (COPD), treatment of complications associated with transplantation, treatment of physiological short stature including growth hormone deficient children and short stature associated 10 with chronic illness, treatment of obesity and growth retardation associated with obesity, treatment of anorexia, treating growth retardation associated with the Prader-Willi syndrome and Turner's syndrome; increasing the growth rate of a patient having partial growth hormone insensitive syndrome, accelerating the recovery and reducing hospitalization of burn patients; treatment of intrauterine 15 growth retardation, skeletal dysplasia, hypercortisolism and Cushing's syndrome; induction of pulsatile growth hormone release; replacement of growth hormone in stressed patients, treatment of osteochondrodysplasias, Noonan's syndrome, schizophrenia, depressions, Alzheimer's disease, delayed wound healing and psychosocial deprivation, treatment of catabolism in connection with pulmonary 20 dysfunction and ventilator dependency; treatment of cardiac failure or related vascular dysfunction, treatment of impaired cardiac function, treatment or prevention of myocardial infarction, lowering blood pressure, protection against ventricular dysfunction or prevention of reperfusion events; treatment of adults in chronic dialysis; attenuation of protein catabolic responses after major surgery, 25 reducing cachexia and protein loss due to chronic illness such as cancer or AIDS; treatment of hyperinsulinemia including nesidioblastosis, adjuvant treatment for ovulation induction; stimulation of thymic development and prevention of the age related decline of thymic function, treatment of immunosuppressed patients; treatment of sarcopenia, treatment of wasting in connection with AIDS; 30 improvement in muscle strength, mobility, maintenance of skin thickness, metabolic homeostasis and renal homeostasis in the frail elderly, stimulation of osteoblasts, bone remodelling and cartilage growth; regulation of food intake; stimulation of the WO 2006/016995 PCT/US2005/022408 - 28 immune system in companion animals and treatment of disorder of aging in companion animals, promoting growth in livestock and stimulation of wool growth in sheep, increasing milk production in livestock, treatment of metabolic syndrome (syndrome X), treatment of insulin resistance, including NIDDM, in mammals, e.g. 5 humans, treatment of insulin resistance in the heart, improvement of sleep quality and correction of the relative hyposomatotropism of senescence due to high increase in REM sleep and a decrease in REM latency, treatment of hypothermia, treatment of frailty associated with ageing, treatment of congestive heart failure, treatment of hip fractures, treatment of immune deficiency in individuals with a depressed T4/T8 10 cell ratio, treatment of muscular atrophy, treatment of musculoskeletal impairment in elderly, enhancing the activity of protein kinase B (PKB), improvement of the overall pulmonary function, treatment of sleep disorders, treatment of growth retardation in connection with asthma, treatment of growth retardation in connection with juvenile rheumatic arthritis, and treatment of growth retardation in connection 15 with cystic fibrosis. Optionally, the pharmaceutical composition of the invention may comprise a polymorph of the present invention combined with one or more compounds exhibiting a different activity, e.g., an antibiotic or other pharmacologically active material. 20 Apart from the pharmaceutical use of a polymorph of the present invention, it may be useful in vitro tools for investigating the regulation of growth hormone release. The polymorphs of the present invention may also be a useful in vivo tool for evaluating the growth hormone releasing capability of the pituitary. For example, 25 serum samples taken before and after administration of the polymorph to humans can be assayed for growth hormone. Comparison of the growth hormone in each serum sample would directly determine the ability of the patient's pituitary to release growth hormone. The polymorph of the present invention may be administered to 30 commercially important animals to increase their rate and extent of growth, and to increase milk production.

WO 2006/016995 PCT/US2005/022408 - 29 A further use of the polymorph of the present invention is in combination with other secretagogues such as GHRP (2 or 6), QHRH and its analogues, growth hormone and its analogues or somatomedins including IGF-1 and IGF-2. The invention will now be described more specifically by the examples. 5 EXAMPLE 1: Crystallization of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form A. 0.0103 g of (3R)-1 -(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide was dissolved in methanol (0.1 10 mL) in a glass vial. The glass vial was then covered with PARAFILM* (thermoplastic film) which was perforated with a single hole. The solvent was then allowed to evaporate under ambient conditions. An X-ray diffraction pattern showed the compound was crystalline (Figure 1). 15 EXAMPLE 2: Crystallization of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form B. 0.0124 g of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide was dissolved in methanol:water (2:1) mixture (0.15 mL) in a glass vial. The glass vial was then covered with 20 PARAFILM* (thermoplastic film) which was perforated with a single hole. The solvent was then allowed to evaporate under ambient conditions. An X-ray diffraction pattern showed the compound was crystalline (Figure 2). EXAMPLE 3: Crystallization of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 25 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C. 0.1003 g of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide was added to methanol (2 mL) in a glass vial. The mixture was filtered into a clean vial. The glass vial was then covered with PARAFILM* (thermoplastic film) which was perforated with a single 30 hole. The solvent was then allowed to evaporate under ambient conditions. XRPD analysis of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- WO 2006/016995 PCT/US2005/022408 -30 piperidinecarboxylic acid 1,2,2-trimethylhydrazide indicated that the molecule was crystalline [data not shown]. EXAMPLE 4: Crystallization of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 5 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C. { 1-[(1R)-2-](3R)-3-Benzyl-3-(NN',N' trimethylhydrazinocarbonyl)piperidin-1-yl-]-1-(1H-indol-3-ylmethyl)-2-oxo ethylcarbamoyl]- 1 -methylethyl} carbamic acid tert-butyl ester (4.72 kg) was dissolved in methanol (5 L/kg) and heated to 55 +/- 5 'C. Methanesulfonic acid (1.5 10 equiv) was added dropwise over 45 minutes while maintaining a temperature of 55 +/- 5 'C. After the addition was complete, the batch temperature was increased and maintained at 60 +/- 5 'C. The progress of the reaction was monitored using HPLC. Upon the completion of the deprotection, the batch was heated to 70 +/- 5 *C and a solution of potassium hydroxide (3 equiv) in water (5 L/kg of Boc-protected 15 compound) was added dropwise over 2 hours while maintaining the temperature. The heat was then removed from the vessel and the batch was allowed to cool to 22 C over 41 hours. The batch was filtered and the cake was washed with 50% aqueous methanol followed by water. The filter cake was then slurried in water at 50 +/- 5 *C for 24 hours, filtered, and washed with water. The filter cake was again 20 slurried in water at 50 +15 C for 17 hours, filtered, and washed with water. The product was then loaded onto trays and dried under vacuum at 70 +/- 5 *C. The final yield of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide was 3.25 kg (81%) with a purity of 99.8 area% by HPLC. Moisture of this product was determined using Karl 25 Fischer in two separate measurements to be 2.4% and 3.2%, which indicates that the sample is in a monohydrate form In an alternative embodiment the steps of slurring the filter cake in purified water at 50 +/- 5 'C for 24 hours, refiltering, and washing with purified water, and repeating the slurry/filter/wash process can be omitted. 30 EXAMPLE 5: Preparation of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form D.

WO 2006/016995 PCT/US2005/022408 -31 15g of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide was produced by the method described in Example 4 except that the steps in which the filter cake was reslurried in water at 50 +/- 5 C were omitted and the product was dried at room temperature 5 in a vacuum oven. This sample was charged to a vessel with 76 mL (5 vol) water. The mixture was heated to 50 C under moderate stirring overnight (16 h). The still warm mixture was filtered through Whatman #1 filter paper, washed with water (15 mL; 1 vol) and dried in a vacuum oven at 60 'C for 54 hours and then dried at 70 'C for 120 hours. An X-ray diffraction pattern showed the compound was crystalline 10 (Figure 3). EXAMPLE 6: Crystallization of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C. (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 15 piperidinecarboxylic acid 1,2,2-trimethylhydrazide amorphous was suspended in methanol (5 L/kg) and the mixture was heated to 65 'C while being stirred. Water (4 L/kg) was then added slowly while the temperature was maintained between 65 68 C. After the addition was complete the mixture was cooled to 60 *C at a rate of 20 'C/h, then from 60 'C to 45 0 C at a rate of 5 'C/h, and then from 45 C to 20 ambient temperature at a rate of 15 'C/h. The crystals were filtered and washed with methanol-water (5:4 1.5 L/kg) and dried in vacuo. Using this protocol approximately 175g of (3R)-1-(2-methylalanyl-D tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide with a purity of 99.3% area was isolated under traceable conditions from crude (3R) 25 1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide (approximately 90 area%). XRPD analysis of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl) 3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide indicated that the molecule was crystalline [data not shown]. KF analysis showed that the sample contained 30 2.1% water. OVI results showed that the product contained methanol of 1.9%.

WO 2006/016995 PCT/US2005/022408 - 32 EXAMPLE 7: Hygroscopicity study of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide (form C). The hygroscopicity of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C 5 produced by the method disclosed in Example 4 (monohydrate) was evaluated using Dynamic Vapor Sorption analysis (DVS) and Thermogravimetric analysis (TGA). Dynamic Vapor Sorption Analysis (DVS) The samples were analyzed using a Hiden IGAsorp vapor sorption analyzer. 10 Each sample was placed in a sample pan and exposed to drying at 25 *C in a dry nitrogen stream for two hours. This assured that the samples being analyzed were at a constant weight prior to the start of the analysis. The samples were then analyzed at 25.0 *C. The adsorption isotherm was conducted between: 10-90 % RH with a step size of 10 %. The desorption isotherm was conducted between 85-0 % RH with 15 a step size 10 %. After the isotherms were complete, each of the samples were heated to 125 'C until the weight loss curve reached an asymptote or for 4 hours. Thermogravimetric Analysis (TGA) The samples were analyzed using a Mettler 85 1e thermogravimetric 20 analyzer. Samples were weighed into a crucible. The samples were heated from 30 'C to 300.0 *C ramped at 10.00 'C/min with an air purge. The (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide initially contained 2.5% by weight bound water (shown by TGA analysis). Bound water was identified by a 25 step transition in the TGA thermogram. DVS analysis showed that during the adsorption isotherm the sample adsorbed water steadily until it contained 6% by weight at 35% RH which indicated the presence of a stable dihydrate. No other significant additional adsorption was observed up to 90% RH. During the desorption isotherm, the dihydrate form was observed down to 30% RH after which 30 desorption of moisture occurred. In conclusion DVS results showed that the dihydrate was formed above a relative humidity of 35%.

WO 2006/016995 PCT/US2005/022408 -33 EXAMPLE 8: Preparation of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C dihydrate and characterization of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 5 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C monohydrate and (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide form C dihydrate. One gram of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by 10 the method of Example 4 (monohydrate) was spread evenly on a Petri dish and equilibrated in a desiccator over a saturated solution of BaC 2 2H 2 0 for 2.5 days. The relative humidity in this environment was ~ 90 %. The conversion to the dihydrate was confirmed by sample weight measurement before and after equilibration. 15 The starting material (monohydrate) and dihydrate were then analyzed by DSC, and KF. XRPD was performed which investigated if moisture sorption altered the crystal lattice of the starting material. X-Ray Powder Diffraction (XRPD) 20 The X-ray powder diffraction patterns were measured on a Shimadzu XRD 6000 X-ray powder diffractometer using Cu Ka radiation. Samples were placed on Si zero- return micro samples holders. The tube voltage and amperage were set to 40 kV and 40 mA, respectively. The divergence and scattering slits were set at 1* and the receiving slit was set at 0.30 mm. A theta-two theta continuous scan at 2 25 */min from 3.0 to 45 020, with a sampling pitch of 0.02 deg and a preset time of 0.60 seconds. Differential Scanning Calorimetry Analysis (DSC) Samples were analyzed using a Mettler 821e Differential Scanning 30 Calorimeter DSC. Samples were weighed in pan, covered with a pierced lid and then crimped (sealed). Analysis conditions were 30-300.0 'C ramped at 10'C/min with a nitrogen purge.

WO 2006/016995 PCT/US2005/022408 - 34 XRPD analysis of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl) 3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide monohydrate indicated that the molecule was crystalline. DSC analysis showed an endothermic transition with onset of 109.5 *C and peak value of 116.4 'C. The extended onset was believed to 5 be due to dehydration of the material. KF analysis showed that the sample contained 2.4% water. This corresponded to an average molar ratio of water/sample of 0.8 or ~ 1, which would indicate the compound was a monohydrate. XRPD results of (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide dihydrate indicated that the 10 molecule was crystalline (Figure 4)and exhibited a similar pattern to the monohydrate starting material. DSC analysis showed a broad endotherm centered at 68.5 *C due to water loss followed by the melt endotherm with onset of 110 'C and peak of 116.8 *C (Figure 5). KF analysis showed that the sample contained 5.6% water. This corresponded to an average molar ratio of water/sample of 1.7 or ~ 2.0, 15 which would indicate the compound was a dihydrate. XRPD results indicated that both the monohydrate and its dihydrate were crystalline and exhibited similar powder patterns. DSC and KF results confirmed the formation of the dihydrate from the monohydrate. 20 EXAMPLE 9: Stability of crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide. Crystalline (3R)-1-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3 piperidinecarboxylic acid 1,2,2-trimethylhydrazide was placed in double plastic bags, closed under a nitrogen blanket, and placed inside a small fiberboard 25 container, the physical appearance, water content, purity and crystallinity were tested every three months for a period of 12 months under a variety of conditions. The results of this study can be seen in Table 5. The purity and water content of (3R)-1-(2-methylalanyl-D-tryptophyl)-3 (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide was analyzed 30 using HPLC and KF respectively before, during, and after the 12 month stability study.

WO 2006/016995 PCT/US2005/022408 -35/1 As can be seen from Table 5 the purity of crystalline (3R)- 1-(2-methylalanyl-D tryptophyl)-3 -(ph enylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethyihydrazide was relatively unchanged after 12 months. Karl Fischer Analysis found that the water content increased from 3.2 % to approximately 6 % after 12 months. 5 O N30190%RJ1 . _____ SIC 4002PJ 6;1-~~1 50urs~ Oi~l15 -M 6 40cnMR.R1 6.1%6 1% ri. St92Z~91a 9.~9~ 41 IIPLWh% rh a0 25'CWLCR 99 ljI; 93z;6 Mr. ~ u SIC917 99.71

.

99 .7 % 991 R* __ _ _ _ 4Deis,1EPi 491r. 99Y. qtr.. srk P 1W0..! Oristi2XC C9t!2TC onst 107.0 C riSC FST0%j .it 113 - OfistlfuW bssi11Wc SowdIou' C Drut 1.213Ct PeaX111r FskilrC Sap 9574) e&hII2.7-C Puat M'C 5OIut 892 'C Grst120.9CT GntrrslC OmthIl2VC .1'I3 HPe* 1349% PaU01.51C Pea*l7rIc *set157.l _____ PeaMO1S21C.1 SIC CI4a6= Crjih Ciysdio QrymmeO MU srrl A~~ry Fwwaiinow 25 CM.. pilI C~d CaW.J0n C y Crybn rwr-s Clissai JUIls9?g 351CM11 ____ ___40C65% R11 Cz %Ai" CrTsUa Cotdoc Table 5, 12 month stability study 10 EQUIVALENTS While this invention has been particularly shown and described with references to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the invention encompassed by the appended claims.

35/2 Throughout this specification, unless the context requires otherwise, the, word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers. Each document, reference, patent application or patent cited in this text is expressly incorporated herein in their entirely by reference, which means that it should be read and considered by the reader as part of this text. That the document, reference, patent application, or patent cited in this text is not repeated in this text is merely for reasons of conciseness. Reference to cited material or information contained in the text should not be understood as a concession that the material or information was part of the common general knowledge or was known in Australia or any other country.

Claims

CLAIMS What is claimed is:

1. A crystalline (3R)-l-(2-methylalanyl-D-tiyptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide.

2. The crystalline composition of Claim 1, wherein the crystalline composition is anhydrous.

3. The crystalline composition of Claim 1, wherein the crystalline composition is a hydrate.

4. The crystalline composition of Claim 3, wherein the crystalline composition is a dihydrate.

5. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0, and 20.8.

6. The crystalline composition of Claim 5, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0, and 20.8.

7. The crystalline composition of Claims 5 or 6, having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 9.6, 17.3, 18.2, 22.1, and 23.5.

8. The crystalline composition of Claim 7, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 9.6, 17.3, 18.2, 22.1, and 23.5.

9. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3.

10. The crystalline composition of Claim 9, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3.

11. The crystalline composition of Claims 9 or 10, having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 9.3, 23.3, 20.4, 22.9, and 23.5

12. The crystalline composition of Claim 11, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 9.3, 23.3, 20.4, 22.9, and 23.5.

13. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 19.2, 20.1, 23.0, 26.2, and 27.0.

14. The crystalline composition of Claim 13, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 19.2, 20.1, 23.0, 26.2, and 27.0.

15. The crystalline composition of Claims 13 or 14, having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 16.3, 21.4, 24.0, 29.8, and 31.5

16. The crystalline composition of Claim 15, having an X-ray powder diffraction pattern wherein at least two 2Θ values measured using Cu K_α radiation are selected from the group consisting of about: 16.3, 21.4, 24.0, 29.8, and 31.5

17. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 10.2, 11.2, 18.7, 20.6, and 23.4.

18. The crystalline composition of Claim 17, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 10.2, 11.2, 18.7, 20.6, and 23.4.

19. The crystalline composition of Claims 17 or 18, having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 9.9, 13.8, 14.3, 16.7, and 19.8.

20. The crystalline composition of Claim 19, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K₀ radiation are selected from the group consisting of about:9.9, 13.8, 14.3, 16.7, and 19.8.

21. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 10.1, 17.0, 20.6, and 23.0.

22. The crystalline composition of Claim 21, having an X-ray powder diffraction pattern including at least one of the following 2Θ values measured using Cu

K_α radiation, about: 17.6, 19.2, 23.0, 23.4, 26.2, and 33.3.

23. The crystalline composition of Claims 21 , or 22 having an X-ray powder diffraction pattern including at least one of the following 2Θ values measured using Cu K_α radiation, about: 9.5, 16.7, 17.5, 17.9, 20.0, 21.5, 23.5, 23.9, and

27.5.

24. A process for preparing crystalline (3R)- 1 -(2-methylalanyl-D-tryptophyl)-3 - (phenylmethyl)-3 -piperidinecarboxylic acid 1 ,2,2-trimethylhydrazide comprising the steps of: a) combining (3R)-I -(2-methylalanyl-D-tryptophyl)-3 -(phenylmethyl)- 3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide with a solvent; b) precipitating the crystals from the solvent; and c) isolating the crystals.

25. The process of Claim 24, wherein the solvent includes at least one member selected from the group consisting of water, methanol, ethanol, n-propanol, isopropanol and butanol and mixtures thereof.

26. The process of Claim 25, wherein the solvent is a mixture of methanol and water.

27. The process of Claim 26, wherein the solvent includes between 40% v/v methanol and 60 % v/v methanol.

28. The process of Claim 24, wherein the solvent is at an elevated temperature while combining with (3R)-l-(2-methylalanyl-D-tryptophyl)-3- (phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide in step a).

29. The process of Claim 24, wherein the solvent is heated to an elevated temperature after combining with (3R)-I -(2-methylalanyl-D-tryptophyl)-3 -

(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide in step a).

30. The process of Claim 28 or 29, wherein the elevated temperature is between 65 ⁰C and 75 °C.

31. The process of Claim 24, wherein the crystals are precipitated in step b) by cooling the solvent.

32. The process of Claim 31 , wherein the solvent is cooled to about ambient temperature.

33. The process of Claim 24, wherein the crystals are isolated in step c) by filtering the crystals from the solvent.

34. The process of Claim 24, further including the step of drying the isolated crystals.

35. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method of Claim 24.

36. A process for preparing crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3- (phenylmethyl)-3 -piperidinecarboxylic acid 1 ,2,2-trimethylhydrazide comprising the steps of: a) combining {l-[(lR)-2-](3R)-3-Benzyl-3-(N,N',N'- trimethylhydrazinocarbonyl)piperidin- 1 -yl-]- 1 -(I H-indol-3 - ylmethyl)-2-oxo-ethylcarbamoyl] - 1 -methylethyl} carbamic acid tert- butyl ester with a solvent; b) combining the mixture from step a) with an acid; c) neutralizing the mixture formed in step b); d) precipitating the crystals from the solvent; and e) isolating the crystals.

37. The process of Claim 36, wherein the solvent in step a) is methanol, and the mixture is neutralized in step c) with a mixture of potassium hydroxide and water.

38. The process of Claim 36, wherein the acid is methanesulfonic acid.

39. The process of Claim 36, wherein the solvent is at an elevated temperature while combining with {l-[(lR)-2-](3R)-3-Benzyl-3-(N,N',N'- trimethylhydrazinocarbonyl)piperidin- 1 -yl-] - 1 -( 1 H-indol-3 -ylmethyl)-2-oxo- ethylcarbamoyl]-l -methylethyl} carbamic acid tert-butyl ester in step a).

40. The process of Claim 36, wherein the solvent is heated to an elevated temperature after combining with {l-[(lR)-2-](3R)-3-Benzyl-3-(N,N',N'- trimethylhydrazinocarbonyl)piperidin- 1 -yl-] - 1 -( 1 H-indol-3 -ylmethyl)-2-oxo- ethylcarbamoyl]-l-methylethyl}carbamic acid tert-butyl ester in step a).

41. The process of Claim 39 or 40, wherein the elevated temperature is between 50 ⁰C and 75 ⁰C.

42. The process of Claim 36, wherein the crystals are precipitated in step d) by cooling the solvent.

43. The process of Claim 42, wherein the solvent is cooled to about ambient temperature.

44. The process of Claim 36, wherein the crystals are isolated in step e) by filtering the crystals from the solvent.

45. The process of Claim 36, further including the step of drying the isolated crystals.

46. A crystalline (3R)-l-(2-methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3- piperidinecarboxylic acid 1,2,2-trimethylhydrazide produced by the method of Claim 36.

47. A pharmaceutical composition comprising crystalline (3R)-I -(2- methylalanyl-D-tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2-trimethylhydrazide and at least one pharmaceutically acceptable carrier or diluent.

48. A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-l-(2-methylalanyl-D- tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- trimethylhydrazide.

49. A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-l-(2-methylalanyl-D- tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0 and 20.8.

50. The method of Claim 49, having an X-ray powder diffraction pattern wherein at least two 2Θ values measured using Cu K₀ radiation are selected from the group consisting of about: 10.1, 11.1, 17.6, 20.0 and 20.8.

51. The method of Claims 49 or 50, having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_n radiation is selected from the group consisting of about: 9.6, 17.3, 18.2, 22.1 and 23.5

52. The method of Claim 51 , having an X-ray powder diffraction pattern wherein at least two 2Θ values measured using Cu K_α radiation are selected from the group consisting of about: 9.6, 17.3, 18.2, 22.1 and 23.5.

53. A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-l-(2-methylalanyl-D- tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu KQ radiation is selected from the group consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3.

54. The method of Claim 53, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 17.0, 19.4, 21.5, 26.2, and 33.3.

55. The method of Claims 53 or 54, having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 9.3, 23.3, 20.4, 22.9, and 23.5.

56. The method of Claim 55, having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 9.3, 23.3, 20.4, 22.9, and 23.5.

57. A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-l-(2-methylalanyl-D- tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 19.2, 20.1, 23.0, 26.2, and 27.0.

58. The method of Claim 57, having an X-ray powder diffraction pattern wherein at least two 2Θ values measured using Cu K_α radiation are selected from the group consisting of about: 19.2, 20.1, 23.0, 26.2, and 27.0.

59. The method of Claims 57 or 58, having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 16.3, 21.4, 24.0, 29.8, and 31.5

60. The method of Claim 59, having an X-ray powder diffraction pattern wherein at least two 2Θ values measured using Cu K_α radiation are selected from the group consisting of about: 16.3, 21.4, 24.0, 29.8, and 31.5

61. A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)- 1 -(2-methylalanyl-D- tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 20 value measured using Cu K_α radiation is selected from the group consisting of about: 10.2, 11.2, 18.7, 20.6, and 23.4.

62. The method of Claim 61 , having an X-ray powder diffraction pattern wherein at least two 20 values measured using Cu K_α radiation are selected from the group consisting of about: 10.2, 11.2, 18.7, 20.6, and 23.4.

63. The crystalline composition of Claims 61 or 62, having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 9.9, 13.8, 14.3, 16.7, and 19.8.

64. The crystalline composition of Claim 63, having an X-ray powder diffraction pattern wherein at least two 2Θ values measured using Cu K_α radiation are selected from the group consisting of about:9.9, 13.8, 14.3, 16.7, and 19.8.

65. A method of stimulating the release of growth hormone from the pituitary of a mammal, the method comprising administering to said mammal a therapeutically effective amount of crystalline (3R)-l-(2-methylalanyl-D- tryptophyl)-3-(phenylmethyl)-3-piperidinecarboxylic acid 1,2,2- trimethylhydrazide having an X-ray powder diffraction pattern wherein at least one 2Θ value measured using Cu K_α radiation is selected from the group consisting of about: 10.1, 17.0, 20.6, and 23.0.

66. The method of Claim 65, having an X-ray powder diffraction pattern including at least one of the following 2Θ values measured using Cu K_α radiation, about: 17.6, 19.2, 23.0, 23.4, 26.2, and 33.3.

67. The crystalline composition of Claims 65, or 66 having an X-ray powder diffraction pattern including at least one of the following 20 values measured using Cu K_a radiation, about: 9.5, 16.7, 17.5, 17.9, 20.0, 21.5, 23.5, 23.9, and 27.5.