COMPOSITIONS AND METHODS FOR TREATMENT OF ABNORMAL CELL GROWTH

Abstract

This invention relates to oral dosage forms and methods that are useful in the treatment of abnormal cell growth, such as cancer, in mammals, especially humans.

Claims

1-60. (canceled)

61. A method of treating cancer in a subject identified as having a mutation status in the gene selected from the group consisting of KRAS, INK4a/Arf, and p53, comprising administering to the subject an effective amount of VS-6063, or a pharmaceutically acceptable salt thereof.

62. The method of claim 61, wherein the cancer is lung cancer.

63. The method of claim 62, wherein the lung cancer is non-small cell lung cancer (NSCLC).

64. The method of claim 63, wherein the NSCLC is KRAS mutant NSCLC.

65. The method of claim 63, wherein the NSCLC is INK4a/Arf mutant NSCLC.

66. The method of claim 63, wherein NSCLC is p53 mutant NSCLC.

67. The method of claim 61, wherein the cancer is metastatic.

68. The method of claim 61, further comprising administering to the subject one or more anti-tumor or anti-cancer agents.

69. The method of claim 68, wherein the one or more anti-tumor or anti-cancer agents are selected from the group consisting of: mitotic inhibitors, alkylating agents, anti-metabolites, intercalating antibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes, topoisomerase inhibitors, biological response modifiers, antibodies, cytotoxics, anti-hormones, and anti-androgens.

70. The method of claim 61, wherein the VS-6063, or a pharmaceutically acceptable salt thereof, is administered 1, 2, 3, or 4 times per day.

71. The method of claim 61, wherein the VS-6063, or a pharmaceutically acceptable salt thereof, is administered 2 times per day.

72. The method of claim 61, wherein 100 to 800 mg per day of VS-6063, or a pharmaceutically acceptable salt thereof, is administered to the subject.

73. The method of claim 61, wherein 200 mg per day of VS-6063, or a pharmaceutically acceptable salt thereof, is administered to the subject.

74. The method of claim 61, wherein 400 mg per day of VS-6063, or a pharmaceutically acceptable salt thereof, is administered to the subject.

75. The method of claim 61, wherein VS-6063, or a pharmaceutically acceptable salt thereof, is administered to the subject as a pharmaceutical composition comprising: 10 to 30% by weight of VS-6063, or a pharmaceutically acceptable salt thereof, per weight of the composition, and a pharmaceutically acceptable excipient.

76. The method of claim 75, wherein the pharmaceutically acceptable excipient is a mixture of two fillers.

77. The method of claim 76, wherein one filler is microcrystalline cellulose and the other filler is lactose.

78. The method of claim 75, wherein the pharmaceutically acceptable excipient is a disintegrant.

79. The method of claim 78, wherein the disintegrant is modified starch.

80. The method of claim 75, wherein the pharmaceutically acceptable excipient is lubricant.

81. The method of claim 80, wherein the lubricant is magnesium stearate.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0125] FIG. 1 shows exemplary mean dissolution profiles of 100 mg VS-6063 IR tablet formulations

[0126] FIG. 2 shows exemplary mean dissolution profiles of 100 mg VS-6063 IR tablet formulations

[0127] FIG. 3 shows exemplary mean dissolution profiles of 100 mg VS-6063 IR tablet formulations

[0128] FIG. 4 shows exemplary mean dissolution profiles of 100 mg VS-6063 IR tablet formulations in 0.1M HCl

[0129] FIG. 5 shows exemplary mean (n=4) VS-6063 Concentrations in Plasma Following an Oral Gavage Dose of VS-6063 in Various Formulations to Male Beagle Dogs at 10 mg API/kg-Rectilinear Scale

[0130] FIG. 6 shows exemplary mean (n=4) VS-6063 Concentrations in Plasma Following an Oral Gavage Dose of VS-6063 in Various Formulations to Male Beagle Dogs at 10 or 20 mg API/kg-Rectilinear Scale

EXAMPLES

[0131] The disclosure is further described in the following examples, which do not limit the scope of the claims.

Example 1. Dry Granulation of VS-6063 Formulations

[0132] VS-6063 immediate release (IR) formulations were prepared using a dry granulation approach. The composition of the formulations is shown on Table 1:

TABLE-US-00001 TABLE 1 Composition of 100 mg VS-6063 IR tablet formulation HPMC-AS Formulation PVP VA/Soluplus Formulation g/per g/per Composition % w/w mg/tablet 200 g batch % w/w mg/tablet 200 g batch Intragranular blend VS-6063* 13.529 108.23 27.058 13.529 108.23 27.058 (free base) (100.00) (100.00) HPMC AS-HF 40.586 324.69 81.172 — — — PVP VA — — — 20.293 162.35 40.586 Soluplus — — — 20.293 162.35 40.586 Avicel PH 102 20.943 167.54 41.886 — — — Lactose Monohydrate 20.943 167.54 41.886 — — — (FastFlo 316) Pearlitol ® Flash — — — 34.885 279.08 69.770 Crospovidone — — — 10.000  80.00 20.000 Sodium starch glycolate 3.000  24.00  6.000 — — — Magnesium Stearate 0.500  4.00  1.000  0.500  4.00  1.000 Sub-total 99.50 796.00 199.00  99.50  796.01 199.00  Extragranular blend Magnesium stearate 0.500  4.00 Final blend for  0.500  4.00 Final blend for Total 100.00 800.00 tableting based on 100.00  800.00 tableting based on yield of granulate yield of granulate available available

[0133] The intragranular blend was prepared by screening VS-6063 and all excipients through 600 μm sieves before use. The required amount of the functional excipients and VS-6063 was weighed out and pre-blended for 1 minute using a large plastic spatula, followed by mixing on the Turbula mixer at a setting of 32 rpm for 5 minutes. The remaining screened components (except the magnesium stearate) were added to the pre-blend mixed for 1 minute using a large plastic spatula, followed by mixing on the Turbula mixer at a setting of 32 rpm for 15 minutes. Final blending on the Turbula mixer was conducted at a setting of 32 rpm for 3 minutes after addition of magnesium stearate. The intragranular blend was then roller compacted. The granule properties for two IR tablet formulations were characterized and compared to the direct compression batch of the same composition; shown in Table 2:

TABLE-US-00002 TABLE 2 Powder characterisation for 100 mg VS-6063 tablet formulations PVP PVP HPMC- HPMC-AS VA/Soluplus VA/Soluplus AS Dry Direct Dry Direct Test granulation Compression granulation Compression Bulk Density 0.522 0.282 0.572 Not tested (g/mL) Tapped Density 0.679 0.570 0.748 Not tested (g/mL) Hausner Ratio 1.30  2.02 1.31 Not tested Carr's Index (%) 23.20  50.60 23.49 Not tested Angle of Repose 17.54  38.37 36.49 Not tested (°)

[0134] The particle size analysis data indicate bi-modal particle size distribution with approximately 20% of the particle size distribution >710 μm and approximately 20% of the particle size distribution <75 μm.

[0135] The physical properties of the blends were determined and compared to the direct compression formulation (HPMC-AS formulation only); (Table 2). The Hausner ratio for the blends tested was >1.3 and the Carr's index was >23% indicating the blends may have poor powder flow, cohesiveness and high interparticulate friction. However, the angle of repose does indicate passable powder flow.

[0136] In comparison to a direct compression approach, a higher compression force may be required to achieve the desired tablet hardness for the dry granulated blends.

TABLE-US-00003 TABLE 3 Tablet compression data for 100 mg VS-6063 tablet formulations PVP PVP HPMC-AS HPMC-AS VA/Soluplus VA/Soluplus Dry Direct Dry Direct Test granulation Compression granulation Compression Average 12.7  13.3 11.7 11.2 Hardness % RSD 8.7 % RSD 3.8 % RSD 8.8 (kP) Min 11.9 Min 12.8 Min 10.4 Max 14.0 Max 14.1 Max 12.7 Average 805.4 804.7 793.0 797.1 Tablet % RSD 0.9 % RSD 0.7 % RSD2.1 % RSD 0.5 Weight  Min 797.8  Min 796.7  Min 763.9  Min 791.2 (mg)  Max 821.6  Max 814.4  Max 810.2  Max 803.2 Average 5.6  6.2 5.4 5.6 Tablet % RSD 0.3 % RSD 0.6 % RSD 1.7 % RSD 0.4 Thickness Min 5.5  Min 6.2  Min 5.2  Min 5.6  (mm) Max 5.6  Max 6.3  Max 5.5  Max 5.6  Compression 1800 800   10000 3500 Force (lbs) Disintegration 10.5  8*  20 8 time (mins)

[0137] FIG. 1 shows that for the HPMC-AS formulation, dry granulation may slow dissolution after a pH switch (e.g., gastric to intestinal pH) in comparison to the dissolution of tablets prepared using a direct compression approach.

[0138] For the PVP VA/Soluplus formulation, the dissolution profiles for tablets prepared via a dry granulation approach may be slower in comparison to tablets prepared using a direct compression approach. The dissolution profile generated was very similar to that for a VS-6063 reference product (e.g., unformulated product).

Example 2. Dry Granulation of VS-6063 Immediate Release Formulations

[0139] One batch of the VS-6063 IR formulations was prepared (Table 4), using a dry granulation approach:

TABLE-US-00004 TABLE 4 Composition of 100 mg VS-6063 tablet formulations PVP VA/Soluplus Composition % w/w mg/tablet g/per 200 g batch Intragranular blend VS-6063* 13.529 108.23 27.058 (free base) (100.00) PVP VA 20.293 162.35 40.586 Soluplus 20.293 162.35 40.586 Pearlitol ® Flash 34.885 279.08 69.770 Crospovidone 10.000 80.00 20.000 Magnesium Stearate 0.500 4.00 1.000 Sub-total 99.50 796.01 199.00 Extragranular blend Magnesium stearate 0.500 4.00 Final blend for Total 100.00 800.00 tableting based on yield of granulate available

[0140] The intragranular blend was prepared by screening VS-6063 and all excipients through 600 μm sieves before use. The required amount of the functional excipients and VS-6063 was weighed out and pre-blended for 1 minute using a large plastic spatula, followed by mixing on the Turbula mixer at a setting of 32 rpm for 5 minutes. The remaining screened components (except the magnesium stearate) were added to the pre-blend, mixed for 1 minute using a large plastic spatula, followed by mixing on the Turbula mixer at a setting of 32 rpm for 15 minutes. Final blending on the Turbula mixer was conducted at a setting of 32 rpm for 3 minutes after addition of magnesium stearate. The intragranular blend was then roller compacted.

[0141] After completion of the dry granulation process, the compacted ribbons were passed through 1.4 mm, 1.18 mm and 850 μm sieves. The tablet blend was then prepared by mixing the granules with magnesium stearate using the Turbula mixer at 32 rpm for 3 minutes.

TABLE-US-00005 TABLE 5 Powder characterisation for 100 mg VS-6063 tablet formulations PVP VA/ PVP VA/ PVP VA/ Soluplus Soluplus Soluplus Prior to dry Dry granulation Dry granulation Test granulation softer ribbons harder ribbons Bulk Density 0.431 0.529 0.572 (g/mL) Tapped Density 0.610 0.761 0.748 (g/mL) Hausner Ratio 1.41 1.44 1.31 Carr's Index (%) 29.29 30.49 23.49 Angle of Repose 22.39 35.83 36.49 (°)

[0142] The particle size analysis data indicate that a larger portion of the blend consisted of fines (approximately 45%<150 μm), in comparison to roller compacted PVP VA/Soluplus formulations (approximately 35%<150 μm), which generated harder ribbons.

[0143] The physical properties of the blend pre and post roller compaction was also determined and indicated that by using a dry granulation approach, the flowability may not be significantly improved (Table 5). The data also indicate that that by formation of softer ribbons, the powder flow properties may not be improved in comparison to granules formed from harder ribbons (Table 5).

[0144] In comparison to a direct compression approach, a higher compression force of 8000 lbs may be required to achieve the desired tablet hardness, for the dry granulation formulation. However, this may be lower in comparison to 10000 lbs compression force that may be required for manufacture of tablets from the granules prepared from harder ribbons.

TABLE-US-00006 TABLE 6 Tablet compression data for 100 mg VS-6063 tablet formulations. PVP PVP VA/ PVP VA/ VA/Soluplus Soluplus Soluplus Dry granulation Dry granulation Direct Test softer ribbons harder ribbons Compression Average Hardness 10.6 11.7 11.2 (kP) % RSD 2.7 % RSD 8.8  Min 10.4  Min 10.4  Max 10.8  Max 12.7 Average Tablet 799.8 793.0 797.1 Weight % RSD 1.2 % RSD 2.1 % RSD 0.5 (mg)  Min 779.1  Min 763.9  Min 791.2  Max 811.0  Max 810.2  Max 803.2 Average Tablet 5.4 5.4 5.6 Thickness % RSD 0.2 % RSD 1.7 % RSD 0.4 (mm) Min 5.4 Min 5.2 Min 5.6 Max 5.4 Max 5.5 Max 5.6 Compression Force 8000 10000 3500 (lbs) Disintegration time 13 20 8 (mins)

[0145] To determine whether addition of further Crospovidone to the formulation would improve the disintegration time, the Crospovidone content was further increased to a total disintegrant content of 13%, where 3% was added to the extragranular blend. Tablets were compressed using 18.97×10.41 mm oval tooling tablet tooling on the Carver tablet press. Compression force was adjusted to achieve a tablet hardness of 10-14 kP. Tablet compression data is shown in Table 6.

TABLE-US-00007 TABLE 7 Tablet compression data for 100 mg VS-6063 tablet formulations. Provisional PVP VA/Soluplus Dry Acceptance granulation with addition Test Criteria of 3% disintegrant Average Hardness 10-14 kP 11.6 (kP) % RSD 6.9 Min 10.8  Max 12.5  Average Tablet Target ± 5.0%, 800.2 Weight RSD ≤ 2% % RSD 2.2 (mg) Min 779.2 Max 835.4 Average Tablet Run and record 5.5 Thickness % RSD 2.0 (mm) Min 5.4  Max 5.7  Compression Force In-process control, not 8000 (lbs) part of the finished product specification Disintegration time Run and record 10.5 (mins)

[0146] Compared with direct compression, granules made by dry granulation may not improve compressibility or powder flow. While using softer ribbons may improve the disintegration time for the resultant tablets, the dissolution profile was not improved in comparison to the reference product (FIG. 2), indicating that the functionality of the precipitation inhibitors in the PVP VA/Soluplus formulation may be lost during dry granulation.

Example 3. Dry Granulation of VS-6063 Formulations

[0147] One batch of the VS-6063 IR formulations has been prepared (Table 8), using a dry granulation approach:

TABLE-US-00008 TABLE 8 Composition of 100 mg VS-6063 IR tablet formulation Composition % w/w mg/tablet g/per 300 g batch Intragranular blend VS-6063* 13.529 108.23 40.587 (free base) (100.00) HPMC AS-HF 40.586 324.69 121.758 Avicel PH 102 20.943 167.54 62.829 Lactose 20.943 167.54 62.829 Monohydrate (FastFlo 316) Sodium starch 3.000 24.00 9.000 glycolate Magnesium 0.500 4.00 1.500 Stearate Sub-total 99.501 796.00 298.503 Extragranular blend Magnesium 0.500 4.00 Final blend for stearate tableting based on Total 100.00 800.00 yield of granulate available

[0148] The intragranular blend preparation was conducted by screening VS-6063 and all excipients through 600 μm sieves before use. The required amount of the functional excipients and the API (e.g., VS-6063) was weighed out and pre-blended for 1 minute using a large plastic spatula, followed by mixing on the Turbula mixer at a setting of 32 rpm for 5 minutes. The remaining screened components (except the magnesium stearate) were added to the pre-blend, mixed for 1 minute using a large plastic spatula, followed by mixing on the Turbula mixer at a setting of 32 rpm for 15 minutes. Final blending on the Turbula mixer was conducted at a setting of 32 rpm for 3 minutes after addition of magnesium stearate. The intragranular blend was then roller compacted.

[0149] The granule properties for the IR tablet formulations are shown in Table 9 and compared to the direct compression batch of the same composition:

TABLE-US-00009 TABLE 9 Powder characterisation for 100 mg VS-6063 tablet formulations HPMC- HPMC- HPMC- HPMC- AS Dry AS Dry AS Dry AS granulation granulation granulation Direct Test batch 1 batch 2 batch 3 Compression Bulk Density Not tested 0.470 0.522 0.282 (g/mL) Tapped Density Not tested 0.640 0.679 0.570 (g/mL) Hausner Ratio Not tested 1.36 1.30 2.02 Carr’s Index (%) Not tested 26.56 23.20 50.60 Angle of Repose Not tested 29.87 17.54 38.37 (°)

[0150] The particle size analysis data indicated a bi-modal particle size distribution for both batches.

[0151] The physical properties of the blends were determined and compared to the direct compression formulation (HPMC-AS formulation only). The data indicated that by using a dry granulation approach the flowability may be improved (Table 9). However, the Hausner ratio for these experiments was >1.3 and the Carr's index was >23%, indicating possibly poor powder flow, cohesiveness and high interparticulate friction. Still the data for angle of repose indicates a passable powder flow.

[0152] For both dry granulation formulations in comparison to a direct compression approach, a higher compression force may be required to achieve the desired tablet hardness.

TABLE-US-00010 TABLE 10 Tablet compression data for 100 mg VS-6063 tablet formulations. HPMC-AS HPMC-AS HPMC-AS Dry Dry Dry HPMC-AS granulation granulation granulation Direct Test batch 1 batch 2 batch 3 Compression Average 12.0 12.3 12.7  13.3 Hardness % RSD 7.0 % RSD 3.3 % RSD 8.7 % RSD 3.8 (kP) Min 10.6  Min 11.9  Min 11.9  Min 12.8  Max 13.6  Max 12.8  Max 14.0  Max 14.1  Average 799.9 802.4 805.4 804.7 Tablet % RSD 0.8 % RSD 0.6 % RSD 0.9 % RSD 0.7 Weight Min 780.4 Min 787.6 Min 797.8 Min 796.7 (mg) Max 812.2 Max 812.9 Max 821.6 Max 814.4 Average 5.9 5.7 5.6  6.2 Tablet % RSD 0.9 % RSD 0.7 % RSD 0.3 % RSD 0.6 Thickness Min 5.7  Min 5.6  Min 5.5  Min 6.2  (mm) Max 6.0  Max 5.8  Max 5.6  Max 6.3  Compression 1300 1500 1800 800   Force (lbs) Disintegration Not measured Not measured 10.5  8*  time (mins)

[0153] The data from FIG. 3 suggests that for the HPMC-AS formulation by introducing a dry granulation approach, the dissolution profiles of all the batches after the pH switch are comparable to tablets prepared using a direct compression approach.

Example 4. In Vivo Assessment of the Pharmacokinetics of VS-6063 in Various Formulations

Abbreviations Used:

[0154] Relative bioavailability=AUC.sub.inf test formulation/AUC.sub.inf reference formulation [0155] C.sub.max Maximum concentration observed [0156] T.sub.max Time of maximum concentration [0157] t.sub.1/2 Terminal half life [0158] AUC.sub.inf Area under the concentration-time curve from time zero extrapolated to infinity [0159] AUC.sub.last Area under the concentration-time curve from time zero to the last quantifiable concentration

Study Design

[0160] 4 groups of 1 male beagle dog per group received an oral gavage dose of VS-6063 in various formulations in a 4-session crossover design with a minimum 3-day washout period between dose sessions. In each session, VS-6063, VS-6063-25% SDD in Soluplus/PVP-A, VS-6063-25% physical mixture with Soluplus/PVP-A, and VS-6063-10% SDD in EUDRAGIT L100-55 were administered at a target dose level of 10 mg API/kg. Blood samples (anticoagulant: K.sub.2EDTA) were scheduled for collection from each animal prior to dosing and at 0.25, 0.5, 1, 2, 4, 6, 8, 12 and 24 hours after dosing in each dose session. Blood samples were processed for plasma and the plasma was analyzed for VS-6063 by LC-MS/MS. Plasma concentration-time data for individual animals were analyzed to determine the pharmacokinetic profiles of VS-6063.

[0161] 4 groups of 1 male beagle dog per group received an oral gavage dose of VS-6063 in various formulations in a 4-session crossover design with a minimum 3-day washout period between dose sessions. In each session, VS-6063-25% physical mixture with Soluplus/PVP-A and VS-6063-25% SDD in Soluplus/PVP-A were administered at a target dose level of 20 mg API/kg and VS-6063-50% physical mixture with Soluplus/PVP-A was administered at target dose levels of 10 and 20 mg API/kg. Blood samples (anticoagulant: K.sub.2EDTA) were scheduled for collection from each animal prior to dosing and at 0.25, 0.5, 1, 2, 4, 6, 8, 12 and 24 hours after dosing in each dose session. Blood samples were processed for plasma and the plasma was analyzed for VS-6063 by LC-MS/MS. Plasma concentration-time data for individual animals were analyzed to determine the pharmacokinetic profiles of VS-6063.

Dose Formulation

[0162] VS-6063 was formulated (0.934 correction factor) at a target concentration of 10 mg API/mL in 0.5% (w/v) Methocel A4M in water with 0.1% (v/v) Tween 80. VS-6063-25% SDD in Soluplus/PVP-A was formulated (0.25 correction factor) at a target concentration of 10 mg API/mL in 0.5% (w/v) Methocel A4M in water. VS-6063-25% physical mixture with Soluplus/PVP-A was formulated (0.25 correction factor) at a target concentration of 10 mg API/mL in 0.5% (w/v) Methocel A4M in water. VS-6063-10% SDD in EUDRAGIT L100-55 was formulated (0.10 correction factor) at a target concentration of 10 mg API/mL in 0.5% (w/v) Methocel A4M in water.

[0163] VS-6063-25% physical mixture with Soluplus/PVP-A and VS-6063-25% SDD in Soluplus/PVP-A were formulated (0.25 correction factor) at a target concentration of 10 mg API/mL in 0.5% (w/v) Methocel A4M in water. VS-6063-50% Physical mixture with Soluplus/PVP-A was formulated (0.50 correction factor) at target concentrations of 5 mg API/mL and 10 mg API/mL in 0.5% (w/v) Methocel A4M in water.

Dose Administration

[0164] 4 groups of 1 male beagle dog per group received an oral gavage dose of VS-6063 in various formulations in a 4-session crossover design with a minimum 3-day washout period between dose sessions. In each session, VS-6063, VS-6063-25% SDD in Soluplus/PVP-A, VS-6063-25% physical mixture with Soluplus/PVP-A and VS-6063-10% SDD in EUDRAGIT L100-55 were administered at a target dose level of 10 mg API/kg. Animals were fasted overnight prior to dose administration; water was not withheld. Food was returned at 4 hours post-dose.

[0165] Prior to dosing, the body weight of each animal was recorded. Doses (rounded to the nearest 0.01 mL) were calculated based on the pretreatment body weight (kg) and a dose volume of 1 mL/kg. Following the oral gavage dose, the gavage tube was flushed with 10 to 20 mL of water prior to removal of the tube. Dosing syringes were weighed immediately prior to and immediately after dosing each animal, and the quantity of formulation administered to each animal was determined from the difference in syringe weights.

[0166] 4 groups of 1 male beagle dog per group received an oral gavage dose of VS-6063 in various formulations in a 4-session crossover design with a minimum 3-day washout period between dose sessions. In each session, VS-6063-25% physical mixture with Soluplus/PVP-A and VS-6063-25% SDD in Soluplus/PVP-A were administered at a target dose level of 20 mg API/kg and VS-6063-50% physical mixture with Soluplus/PVP-A was administered at target dose levels of 10 and 20 mg API/kg. Animals were fasted overnight prior to dose administration; water was not withheld. Food was returned at 4 hours post-dose.

[0167] Prior to dosing, the body weight of each animal was recorded. Doses (rounded to the nearest 0.01 mL) were calculated based on the pretreatment body weight (kg) and a dose volume of 2 mL/kg. Following the oral gavage dose, the gavage tube was flushed with 10 to 20 mL of water prior to removal of the tube. Dosing syringes were weighed immediately prior to and immediately after dosing each animal, and the quantity of formulation administered to each animal was determined from the difference in syringe weights.

VS-6063 Concentrations in Plasma

[0168] Mean (n=4) plasma concentration-time profiles are shown in FIG. 5 (rectilinear scale).

[0169] Following a single oral dose of VS-6063 at a target dose of 10 mg/kg over 4 sessions, plasma concentrations rose at a fairly rapid rate and maximum concentrations for individual animals were reached within 2 to 4 hours post-dose (mean 2.50 hours). Mean concentrations then declined in an apparent first-order manner. Concentrations of VS-6063 were quantifiable in all animals through the 24 hours post-dose (the final time point).

[0170] Following a single oral dose of VS-6063-25% SDD in Soluplus/PVP-A, VS-6063-25% physical mixture with Soluplus/PVP-A, and VS-6063-10% SDD in EUDRAGIT L100-55 at a target dose of 10 mg API/kg over 4 sessions, plasma concentrations rose at a fairly rapid rate and maximum concentrations for individual animals were reached within 0.5 to 4 hours post-dose (mean range of 1.75 to 1.88 hours). Mean concentrations then declined in an apparent first-order manner. Concentrations of VS-6063 were quantifiable in all animals through the 24 hours post-dose (the final time point).

Plasma Pharmacokinetics for VS-6063

[0171] Plasma pharmacokinetic parameters for VS-6063 are shown in Tables 13 and 14.

[0172] For VS-6063 at a target dose of 10 mg/kg over 4 sessions, the mean estimated half life was approximately 3.9 hours. Mean C.sub.max and mean T.sub.max were 614 ng/mL and 2.50 hours, respectively. Mean AUC.sub.last was 3100 ng*hr/mL and mean AUC.sub.inf was 3120 ng*hr/mL. (Table 17)

[0173] For VS-6063-25% SDD in Soluplus/PVP-A at a target dose of 10 mg API/kg over 4 sessions, the mean estimated half life was approximately 3.2 hours. Mean C.sub.max and mean T.sub.max were 1770 ng/mL and 1.75 hours, respectively. Mean AUC.sub.last was 8130 ng*hr/mL and mean AUC.sub.inf was 8170 ng*hr/mL. (Table 18)

[0174] For VS-6063-25% physical mixture with Soluplus/PVP-A at a target dose of 10 mg API/kg over 4 sessions, the mean estimated half life was approximately 3.7 hours. Mean C.sub.max and mean T.sub.max were 1460 ng/mL and 1.75 hours, respectively. Mean AUC.sub.last was 7320 ng*hr/mL and mean AUC.sub.inf was 7410 ng*hr/mL. (Table 19)

[0175] For VS-6063-10% SDD in EUDRAGIT L100-55 at a target dose of 10 mg API/kg over 4 sessions, the mean estimated half life was approximately 2.9 hours. Mean C.sub.max and mean T.sub.max were 1800 ng/mL and 1.88 hours, respectively. Mean AUC.sub.last was 8720 ng*hr/mL and mean AUC.sub.inf was 8740 ng*hr/mL. (Table 20)

[0176] Mean pharmacokinetic data suggests relative bioavailability was increased when comparing test formulations (VS-6063-25% SDD, VS-6063-25% physical mixture and VS-6063-10% SDD to the reference formulation (VS-6063), although variability among animals was significant. Mean relative bioavailability estimates were 2.97 (CV 72.7%) for VS-6063-25% physical mixture, 3.21 (CV 43.3%) for VS-6063-10% SDD, and 3.78 (CV 89.9%) for VS-6063-25% SDD. When comparing crossover data; Animal No. 4001 had the lowest exposure to the reference formulation (VS-6063) which contributed to overall variability among animals receiving the treatment (CV 52.8% for mean AUC.sub.inf) as well as contributing to the highest relative bioavailability among test formulations.

[0177] Although emesis was observed in three of the four subjects dosed with VS-6063-25% physical mixture, mean relative bioavailability was similar to the other test formulations when compared to the reference formulation.

VS-6063 Concentrations in Plasma

[0178] Mean plasma concentration-time profiles are shown in FIG. 6 (rectilinear scale).

[0179] Following a single oral dose of VS-6063-25% physical mix at a target dose of 20 mg API/kg over 4 sessions, plasma concentrations rose at a rapid rate and maximum concentrations were reached within 0.25 to 1 hour post-dose (mean 0.688 hr). Mean concentrations then declined in an apparent first-order manner. Concentrations of VS-6063 were quantifiable in all animals through the 24 hours post-dose (the final time point).

[0180] Following a single oral dose of VS-6063-25% SDD at a target dose of 20 mg API/kg over 4 sessions, plasma concentrations rose at a fairly rapid rate and maximum concentrations were reached within 0.25 to 2 hours post-dose (mean 1.31 hr). Mean concentrations then declined in an apparent first-order manner. Concentrations of VS-6063 were quantifiable in all animals through the 24 hours post-dose (the final time point).

[0181] Following a single oral dose of VS-6063-50% physical mixture at target doses of 10 mg API/kg and 20 mg API/kg over 4 sessions, plasma concentrations rose at a fairly rapid rate and maximum concentrations were reached within 0.5 to 2 hours post-dose (mean 1.38 hr) for the 10 mg API/kg dose and within 0.5 to 4 hours post-dose (mean 2.13 hr) for the 20 mg API/kg dose. Mean concentrations then declined in an apparent first-order manner. Concentrations of VS-6063 were quantifiable in all animals through the 24 hours post-dose (the final time point).

Plasma Pharmacokinetics for VS-6063

[0182] Plasma pharmacokinetic parameters for VS-6063 are shown in Tables 15 and 16.

[0183] For VS-6063-25% physical mixture with Soluplus/PVP-A at a target dose of 20 mg API/kg over 4 sessions, the mean estimated half life was approximately 3.0 hours. Mean C.sub.max and mean T.sub.max were 1670 ng/mL and 0.688 hours, respectively. Mean AUC.sub.last was 8670 ng*hr/mL and mean AUC.sub.inf was 8700 ng*hr/mL. (Table 21)

[0184] For VS-6063-25% SDD in Soluplus/PVP-A at a target dose of 20 mg API/kg over 4 sessions, the mean estimated half life was approximately 3.1 hours. Mean C.sub.max and mean T.sub.max were 1300 ng/mL and 1.31 hours, respectively. Mean AUC.sub.last was 5920 ng*hr/mL and mean AUC.sub.inf was 5940 ng*hr/mL. (Table 22)

[0185] For VS-6063-50% physical mixture with Soluplus/PVP-A at a target dose of 10 mg API/kg over 4 sessions, the mean estimated half life was approximately 3.0 hours. Mean C.sub.max and mean T.sub.max were 680 ng/mL and 1.38 hours, respectively. Mean AUC.sub.last was 3390 ng*hr/mL and mean AUC.sub.inf was 3400 ng*hr/mL. (Table 23)

[0186] For VS-6063-50% physical mixture with Soluplus/PVP-A at a target dose of 20 mg API/kg over 4 sessions, the mean estimated half life was approximately 3.5 hours. Mean C.sub.max and mean T.sub.max were 1160 ng/mL and 2.13 hours, respectively. Mean AUC.sub.last was 6900 ng*hr/mL and mean AUC.sub.inf was 6930 ng*hr/mL. (Table 24)

[0187] Observations of emesis corresponded with lower exposure while in other instances there was no correlation.

[0188] Pharmacokinetic data suggests that VS-6063-50% physical mixture at target doses of 10 mg API/kg and 20 mg API/kg are dose proportional: mean dose-normalized AUC.sub.inf and C.sub.max may be similar.

[0189] Comparing data from both studies, VS-6063-25% physical mixture (20 mg API/kg) and VS-6063-25% SDD (20 mg API/kg) were less than dose proportional relative to dose-normalized mean AUC.sub.inf and C.sub.max with VS-6063-25% physical mixture (10 mg API/kg) and VS-6063-25% SDD (10 mg API/kg) as summarized below in Table 11.

TABLE-US-00011 TABLE 11 Dose Level Mean C.sub.max/ Mean AUC.sub.inf/ Formulation mg API/kg Dose Dose 25% Physical Mixture 20 83.5 440 25% Physical Mixture 10 146 741 25% SDD 20 65.0 297 25% SDD 10 177 817

[0190] Mean pharmacokinetic data suggests relative bioavailability is similar when comparing test formulations VS-6063-50% physical mixture at target doses of 10 mg API/kg and 20 mg API/kg to reference formulation VS-6063 (10 mg/kg).

TABLE-US-00012 TABLE 12 Dose Level Mean Relative Formulation mg API/kg AUC.sub.inf/Dose Bioavailability VS-6063 (neat) 10 312 NA 50% Physical Mixture 10 340 1.09 50% Physical Mixture 20 347 1.11

CONCLUSIONS

[0191] Mean pharmacokinetic data suggests relative bioavailability may be increased when comparing VS-6063-25% SDD in Soluplus (10 mg API/kg), VS-6063-25% physical mixture with Soluplus (10 mg API/kg), and VS-6063-10% SDD in EUDRAGIT L100-55 (10 mg API/kg) to VS-6063 (10 mg/kg), although there was variability among animals. Mean relative bioavailability estimates were 2.97 (CV 72.7%) for VS-6063-25% physical mixture, 3.21 (CV 43.3%) for VS-6063-10% SDD, and 3.78 (CV 89.9%) for VS-6063-25% SDD.

[0192] Mean pharmacokinetic data suggests that VS-6063-50% physical mixture with Soluplus/PVP-A at target doses of 10 mg API/kg and 20 mg API/kg are dose proportional. Mean dose-normalized mean AUC.sub.inf and mean C.sub.max may be generally similar.

[0193] Comparing data from both studies, VS-6063-25% physical mixture (20 mg API/kg) and VS-6063-25% SDD (20 mg API/kg) are less than dose proportional when comparing dose-normalized mean AUC.sub.inf and mean C.sub.max with VS-6063-25% physical mixture (10 mg API/kg) and VS-6063-25% SDD (10 mg API/kg). Dose normalized mean pharmacokinetic data suggests relative bioavailability may be similar when comparing VS-6063-50% physical mixture at target doses of 10 mg API/kg and 20 mg API/kg to VS-6063 (10 mg/kg).

TABLE-US-00013 TABLE 13 Oral (Gavage) Dose Administration of VS-6063 in Various Formulations to Male Beagle Dogs Animal Formulation Protocol Dosing Session Test Article Group Animal Weight Admin Conc.sup.a Dose Administered Dose Variance No. Formulation No. No. (kg) (g) (mg/g) (mg) (mg/kg) (mg/kg) (%) 1 VS-6063 1 1001 8.80 7.986 10 79.86 9.075 10 −9.25 VS-6063-25% SDD in 2 2001 10.74 10.577 10 105.8 9.848 10 −1.52 Soluplus/PVP-A VS-6063-25% Phys Mix 3 3001 10.34 10.280 10 102.8 9.942 10 −0.58 with Soluplus/PVP-A VS-6063-10% SDD in 4 4001 9.14 8.688 10 86.88 9.505 10 −4.95 EUDRAGIT L100-55 2 VS-6063-10% SDD in 1 1001 9.22 8.872 10 88.72 9.623 10 −3.77 EUDRAGIT L100-55 VS-6063 2 2001 10.46 9.794 10 97.94 9.363 10 −6.37 VS-6063-25% SDD in 3 3001 11.10 10.951 10 109.51 9.866 10 −1.34 Soluplus/PVP-A VS-6063-25% Phys Mix 4 4001 9.62 9.516 10 95.16 9.892 10 −1.08 with Soluplus/PVP-A 3 VS-6063-25% Phys Mix 1 1001 8.50 8.459 10 84.59 9.952 10 −0.48 with Soluplus/PVP-A VS-6063-10% SDD in 2 2001 10.58 9.950 10 99.50 9.405 10 −5.95 EUDRAGIT L100-55 VS-6063 3 3001 10.48 10.462 10 104.6 9.983 10 −0.17 VS-6063-25% SDD in 4 4001 9.38 9.280 10 92.80 9.893 10 −1.07 Soluplus/PVP-A 4 VS-6063-25% SDD in 1 1001 8.820 8.800 10 88.00 9.977 10 −0.23 Soluplus/PVP-A VS-6063-25% Phys Mix 2 2001 10.800 10.692 10 106.9 9.900 10 −1.00 with Soluplus/PVP-A VS-6063-10% SDD in 3 3001 10.580 10.390 10 103.9 9.820 10 −1.80 EUDRAGIT L100-55 VS-6063 4 4001 9.380 8.864 10 88.64 9.450 10 −5.50 .sup.aAssumes a density of 1 g/mL.

TABLE-US-00014 TABLE 14 Mean (n = 4) Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063 in Various Formulations to Male Beagle Dogs at 10 mg/kg C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Relative Formulation (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) Bioavailability VS-6063 Mean 614 2.50 3.86 3120 3100 NA SD 302 1.00 0.872 1650 1640 NA % CV 49.2 40.0 22.6 52.8 53.0 NA VS-6063-25% SDD in Mean 1770 1.75 3.24 8170 8130 3.78 Soluplus/PVP-A SD 791 1.66 0.501 2490 2480 3.40 % CV 44.8 94.8 15.5 30.5 30.5 89.9 VS-6063-25% Phys Mix Mean 1460 1.75 3.71 7410 7320 2.97 With Soluplus/PVP-A SD 520 1.66 0.551 3400 3310 2.16 % CV 35.7 94.8 14.9 45.8 45.2 72.7 VS-6063-10% SDD in Mean 1800 1.88 2.93 8740 8720 3.21 EUDRAGIT L100-55 SD 495 1.55 0.351 3560 3540 1.39 % CV 27.6 82.6 12.0 40.7 40.7 43.3 Relative bioavailability = AUC.sub.inf test formulation/AUC.sub.inf reference formulation

TABLE-US-00015 TABLE 15 Oral (Gavage) Dose Administration of VS-6063 in Various Formulations to Male Beagle Dogs Animal Formulation Protocol Dosing Session Test Article Group Animal Weight Admin Conc.sup.a Dose Administered Dose Variance No. Formulation No. No. (kg) (g) (mg/g) (mg) (mg/kg) (mg/kg) (%) 1 VS-6063-25% Phys Mix 1 1001 9.76 19.44 10 194.35 19.91 20 −0.44 with Soluplus/PVP-A VS-6063-25% SDD in 2 2001 9.40 18.64 10 186.38 19.83 20 −0.86 Soluplus/PVP-A VS-6063-50% Phys Mix 3 3001 8.64 17.01 5 85.03 9.84 10 −1.59 with Soluplus/PVP-A VS-6063-50% Phys Mix 4 4001 9.68 19.26 10 192.58 19.89 20 −0.53 with Soluplus/PVP-A 2 VS-6063-50% Phys Mix 1 1001 10.08 19.98 10 199.75 19.82 20 −0.92 with Soluplus/PVP-A VS-6063-25% Phys Mix 2 2001 9.08 18.33 10 183.25 20.18 20 0.91 with Soluplus/PVP-A VS-6063-25% SDD in 3 3001 9.00 17.95 10 179.52 19.95 20 −0.27 Soluplus/PVP-A VS-6063-50% Phys Mix 4 4001 9.90 19.49 5 97.44 9.84 10 −1.58 with Soluplus/PVP-A 3 VS-6063-50% Phys Mix 1 1001 9.94 19.45 5 97.24 9.78 10 −2.18 with Soluplus/PVP-A VS-6063-50% Phys Mix 2 2001 8.68 17.08 10 170.83 19.68 20 −1.60 with Soluplus/PVP-A VS-6063-25% PHYS mix 3 3001 8.40 16.61 10 166.10 19.77 20 −1.13 with Soluplus/PVP-A VS-6063-25% SDD in 4 4001 9.80 18.96 10 189.62 19.35 20 −3.26 Soluplus/PVP-A 4 VS-6063-25% SDD in 1 1001 10.50 20.45 10 204.49 19.48 20 −2.62 Soluplus/PVP-A VS-6063-50% Phys Mix 2 2001 9.00 17.70 5 88.48 9.83 10 −1.69 with Soluplus/PVP-A VS-6063-50% Phys Mix 3 3001 8.92 17.65 10 176.47 19.78 20 −1.08 with Soluplus/PVP-A VS-6063-25% Phys Mix 4 4001 10.40 20.61 10 206.05 19.81 20 −0.94 with Soluplus/PVP-A .sup.aAssumes a density of 1 g/mL.

TABLE-US-00016 TABLE 16 Mean (n = 4) Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063 in Various Formulations to Male Beagle Dogs at 10 or 20 mg API/kg C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Formulation Dose (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) C.sub.max/Dose AUC.sub.inf/Dose VS-6063-25% Phys Mix Mean 1670 0.688 3.01 8700 8670 83.5 440 with Soluplus/PVP-A SD 580 0.375 0.156 3400 3390 29.0 170 20 mg API/kg % CV 34.7 54.5 5.2 39.1 39.1 34.7 39.1 VS-6063-25% SDD in Mean 1300 1.31 3.12 5940 5920 64.9 297 Soluplus/PVP-A SD 672 0.851 0.289 1640 1640 33.6 82.1 20 mg API/kg % CV 51.8 64.8 9.3 27.6 27.7 51.8 27.6 VS-6063-50% Phys Mix Mean 680 1.38 3.00 3400 3390 68.0 340 with Soluplus/PVP-A SD 190 0.750 0.0208 1150 1150 19.0 115 10 mg API/kg % CV 28.0 54.5 0.700 33.9 33.8 28.0 33.9 VS-6063-50% Phys Mix Mean 1160 2.13 3.51 6930 6900 57.9 347 with Soluplus/PVP-A SD 516 1.44 0.813 3500 3500 25.8 175 20 mg API/kg % CV 44.6 67.6 23.1 50.5 50.8 44.6 50.5

TABLE-US-00017 TABLE 17 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063 to Male Beagle Dogs at 10 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) 1 1001 537 2 3.49 2830 2820 2 2001 987 4 3.42 5200 5180 3 3001 671 2 3.39 3240 3230 4 4001 260 2 5.17 1200 1180 Mean 614 2.50 3.86 3120 3100 SD 302 1.00 0.872 1650 1640 % CV 49.2 40.0 22.6 52.8 53.0

TABLE-US-00018 TABLE 18 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063-25% SDD in Soluplus/PVP-A to Male Beagle Dogs at 10 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Relative Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) Bioavailability 4 1001 2390 0.5 3.58 9880 9840 3.49 1 2001 1280 4 2.58 7460 7450 1.43 2 3001 912 2 3.12 4950 4930 1.53 3 4001 2490 0.5 3.67 10400 10300 8.68 Mean 1770 1.75 3.24 8170 8130 3.78 SD 791 1.66 0.501 2490 2480 3.40 % CV 44.8 94.8 15.5 30.5 30.5 89.9

TABLE-US-00019 TABLE 19 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063-25% Physical Mixture With Soluplus/PVP-A to Male Beagle Dogs at 10 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Relative Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) Bioavailability 3 .sup. 1001.sup.a 1810 0.5 3.26 6420 6350 2.27 4 2001 1750 4 4.40 12000 11800 2.31 1 3001 691 2 3.92 3890 3850 1.20 2 4001 1580 0.5 3.27 7320 7300 6.12 Mean 1460 1.75 3.71 7410 7320 2.97 SD 520 1.66 0.551 3400 3310 2.16 % CV 35.7 94.8 14.9 45.8 45.2 72.7 .sup.aRegression R square value < 0.9

TABLE-US-00020 TABLE 20 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063-10% SDD in EUDRAGIT L100-55 to Male Beagle Dogs at 10 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Relative Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) Bioavailability 2 1001 2180 4 2.77 11400 11400 4.03 3 2001 2200 1 3.33 12200 12100 2.35 4 3001 1160 2 3.10 5700 5680 1.76 1 4001 1650 0.5 2.53 5640 5640 4.72 Mean 1800 1.88 2.93 8740 8720 3.21 SD 495 1.55 0.351 3560 3540 1.39 % CV 27.6 82.6 12.0 40.7 40.7 43.3

TABLE-US-00021 TABLE 21 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS- 6063-25% Physical Mixture With Soluplus/PVP-A to Male Beagle Dogs at 20 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) C.sub.max/Dose AUC.sub.inf/Dose 1 1001 952 1 3.15 4770 4750 47.6 240 2 2001 1570 1 3.13 7200 7180 78.5 360 3 3001 1810 0.5 2.87 10300 10300 90.5 520 4 4001 2350 0.25 2.87 12500 12500 120 630 Mean 1670 0.688 3.01 8700 8670 83.5 440 SD 580 0.375 0.156 3400 3390 29.0 170 % CV 34.7 54.5 5.2 39.1 39.1 34.7 39.1

TABLE-US-00022 TABLE 22 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS-6063-25% SDD in Soluplus/PVP-A to Male Beagle Dogs at 20 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) C.sub.max/Dose AUC.sub.inf/Dose 4 1001 2240 0.25 2.73 7870 7860 112 394 1 2001 650 2 3.15 3930 3920 32.5 197 2 3001 1200 2 3.43 6380 6350 60.0 319 3 4001 1100 1 3.18 5580 5560 55.0 279 Mean 1300 1.31 3.12 5940 5920 64.9 297 SD 672 0.851 0.289 1640 1640 33.6 82.1 % CV 51.8 64.8 9.3 27.6 27.7 51.8 27.6

TABLE-US-00023 TABLE 23 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS- 6063-50% Physical Mixture With Soluplus/PVP-A to Male Beagle Dogs at 10 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) C.sub.max/Dose AUC.sub.inf/Dose 3 1001 809 2 3.03 3970 3960 80.9 397 4 2001 398 0.5 3.01 1820 1810 39.8 181 1 3001 733 2 2.98 3350 3340 73.3 335 2 4001 778 1 3.00 4470 4450 77.8 447 Mean 680 1.38 3.00 3400 3390 68.0 340 SD 190 0.750 0.0208 1150 1150 19.0 115 % CV 28.0 54.5 0.700 33.9 33.8 28.0 33.9

TABLE-US-00024 TABLE 24 Plasma Pharmacokinetic Parameters for VS-6063 Following an Oral Gavage Dose of VS- 6063-50% Physical Mixture With Soluplus/PVP-A to Male Beagle Dogs at 20 mg API/kg Animal C.sub.max T.sub.max t.sub.1/2 AUC.sub.inf AUC.sub.last Session Number (ng/mL) (hr) (hr) (hr*ng/mL) (hr*ng/mL) C.sub.max/Dose AUC.sub.inf/Dose 2 1001 1210 0.5 3.32 6250 6230 60.5 312 3 2001 1710 2 2.88 11000 10900 85.5 549 4 3001 1250 2 3.14 7930 7900 62.5 396 1 4001 462 4 4.70 2570 2530 23.1 129 Mean 1160 2.13 3.51 6930 6900 57.9 347 SD 516 1.44 0.813 3500 3500 25.8 175 % CV 44.6 67.600 23.100 50.5 50.8 44.6 50.5

Example 5. Clinical Study Protocol: Phase I Study of Safety, Bioavailability, and Pharmacodynamics of Selected Formulations of VS-6063

Study Design

[0194] In a Phase I, open-label, single center, single dose, partially randomized trial, the safety, bioavailability, and pharmacodynamics of prototype and reference formulations of VS-6063 were investigated in healthy male subjects. The study was comprised of four periods, each of which adhered to the same study design and involved 24 healthy male subjects between 40 and 65 years of age with a body mass index between 18.0 and 35.0 kg/m.sup.2. Following a 28 day screening period, the subjects were admitted to the clinical unit the morning prior to dosing day (Day −1) and were dosed in the morning of Day 1. Subjects in Regimens A and B were administered VS-6063 following an overnight fast of about 10 hours, while subjects in Regimens C and D were administered VS-6063 within 30 minutes of starting a high fat breakfast, which was consumed within 25 minutes. Subjects remained on site until 36 hours after dosing. There was a minimum washout period of 7 days between the administration of each regimen. A follow-up phone call took place 7 to 10 days post final dose in periods 3 and 4 to ensure the ongoing well-being of the subjects.

TABLE-US-00025 TABLE 25 Summary of pharmacokinetic results Regimen A Regimen B Regimen C Regimen D Formulation Reference Prototype Prototype Reference Dose 200 mg 100 mg 100 mg 200 mg Fasted/Fed Fasted Fasted Fed Fed Parameter N = 24 N = 24 N = 23 N = 9 T.sub.lag.sup.a (h) 0.000 (0.00- 0.042 (0.00- 0.000 (0.00- 0.000 (0.00- 0.00) 0.50) 0.52) 1.00) T.sub.max.sup.a (h) 1.000 (0.50- 2.000 (1.00- 4.000 (2.00- 4.000 (2.00- 3.00) 4.00) 10.02) 6.00) C.sub.max (ng/mL)  212 (72.8%)  446 (32.3%)  262 (43.5%)  427 (66.7%) C.sub.12 (ng/mL)  6.1 (78.0%) 15.1 (88.8%) 22.7 (80.6%)  38.5 (174.0%) C.sub.24 (ng/mL) 3.30 (59.9%) 3.72 (63.4%) 4.45 (74.2%) 4.52 (93.2%)  [n = 23] AUC.sub.last (ng .Math. h/mL)  883 (69.2%) 1910 (36.6%)   1590 (44.6%)   2490 (75.8%)  AUC.sub.inf (ng .Math. h/mL)  755 (46.0%) 2350 (29.2%)   1790 (37.0%)   2970 (75.4%)  [n = 2] [n = 11] [n = 12] [n = 7] AUC.sub.% extrap (%) 1.581 (155.7%) 1.028 (41.8%)  1.232 (46.1%)  0.574 (69.3%)  [n = 2] [n = 11] [n = 12] [n = 7] lambda-z 0.16125 (214.7) 0.10403 (31.4%)   0.09156 (21.4%)   0.13372 (40.1%)   [n = 2] [n = 11] [n = 12] [n = 7] α t.sub.1/2 (h) 1.880 (18.9%)  1.989 (20.1%)  2.082 (21.4%)  2.154 (15.9%)  (Distributional Component) t.sub.1/2 (h) 4.299 (214.7%) 6.663 (31.4%)  7.571 (21.4%)  5.183 (40.1%)  [n = 2] [n = 11] [n = 12] [n = 7] .sup.aMedian (range)
Abbreviations: T.sub.lag.sup.a: the elapsed time from dosing at which VS-6063 was first quantifiable in a concentration vs time profile; C.sub.max: the maximum observed VS-6063 concentration; T.sub.max.sup.a: the time from dosing at which C.sub.max was present; C.sub.12: the observed VS-6063 concentration at 12 h; C.sub.24: the observed VS-6063 concentration at 24 h; AUC.sub.last: the area under the concentration vs time curve from time zero to the time of the last quantifiable concentration; AUC.sub.inf: the area under the concentration vs time curve from time zero extrapolated to infinity; AUC.sub.% extrap: the percentage of AUC.sub.(0-inf) accounted for by extrapolation; lambda-z: slope of the regression time passing through the apparent elimination phase in a concentration vs time plot; t.sub.1/2: the apparent elimination half-life; α t.sub.1/2: the distributional half-life of VS-6063 occurring over the time range between C.sub.max and approximately 12 h post-dose.

Statistical Methods

[0195] Formal statistical analysis was performed on the dose corrected pharmacokinetic (PK) parameters AUC.sub.last, AUC.sub.inf, and C.sub.max to assess relative bioavailability and the presence of food effects. The PK parameters were subjected to a natural logarithmic transformation and were analyzed using mixed model techniques. Adjusted geometric mean rations (GMRs) and 90% confidence intervals (CIs) for the adjusted GMRs were calculated for the comparisons of the different regimens. The CIs were based on the t-distribution and the variances estimated by the model using the Kenward-Roger method for calculating the degrees of freedom.

Study Results

[0196] Following administration of either a 200 mg VS-6063 reference formulation tablet (comprising VS-6063, microcrystalline cellulose PH 102, Fastflo 316, sodium starch glycolate, and magnesium stearate) or a 100 mg VS-6063 prototype formulation tablet (comprising VS-6063, HPMCAS-HF, microcrystalline cellulose PH 102, Fastflo 316, sodium starch glycolate, and magnesium stearate) in the fed and fasted states, plasma concentrations rose to a maximal level immediately following dosing. This was followed by a biphasic reduction in plasma levels over the 24 h sampling period, as VS-6063 was distributed and eliminated from the systemic circulation. No real difference in half-life could be determined between the reference and prototype formulations in either the fed or fasted states.

[0197] In the fasted state, the 100 mg prototype formulation administered in Regimen B lead to significantly higher exposure in comparison to the 200 mg reference formulation in Regimen A. The adjusted GMRs and 90% CIs for Regimen B/Regimen A based on C.sub.max and AUC.sub.k were 421.33 (348.4%, 509.5%) and 433.05 (366.8%, 511.3%), respectively, equating to a 4-fold increase in exposure as a result of the change in formulation. In addition to the increase in bioavailability, there was evidence of a decrease in variability observed upon administration of the prototype formulation associated with exposure parameters from approximately 70% to 30%.

[0198] Comparison of the time taken to reach maximum plasma concentrations of VS-6063 was 2 to 4 times longer in the fed state than in the fasted state (as assessed by C.sub.max and AUC.sub.last), indicating a food effect on the rate of absorption. In addition, determination of exposure following administration of the 200 mg VS-6063 reference formulation in both the fed and fasted state indicate a higher level of exposure in the fed state relative to the fasted state, which signifies the presence of a food effect for the reference formulation. Despite these effects, an increase in VS-6063 exposure was still observed when the prototype formulation was administered compared with the reference formulation in the fed state. The adjusted GMRs and 90% CIs for Regimen C/Regimen D based on C.sub.max and AUC.sub.last were 129.8 and 125.1, respectively.

[0199] Overall, the maximum exposure was achieved with the prototype formulation administered in the fasted state. Equivalent exposure in terms of AUC.sub.inf was also achieved with the prototype formulation in the fed state.

[0200] VS-6063 was well tolerated when administered in both the fed and fasted states in both the reference and prototype formulations. There were no serious adverse events (AEs) or severe AEs reported during the study, and no subject was withdrawn as a result of an AE. The overall incidence of total AEs was low, with 12 AEs reported by 9 subjects. The incidence of AEs was highest for Regimen C (5 subjects) compared to Regimens A, B, and D (3 subjects, 2 subjects, and 1 subject, respectively). The most common AE was headache, which was reported by 3 subjects. All other AEs were reported by 1 subject. No subject reported an AE that was related to VS-6063, and nearly all AEs were mild in severity. There were no clinically significant findings in any laboratory assessments, measurement of vital signs, ECGs, or physical examinations.