Article Figures & Data
Figures
- Fig. 1.
Effect of various PF-06952229 dosing regimens on in vivo efficacy in the syngeneic MC38 colon carcinoma mouse model. Female C57BL/6 mice bearing established MC38 tumors (mean volume ∼100 mm3) were sorted into four groups (n = 10 mice/group) and dosed by oral gavage with vehicle (group 1, 0.5% methyl cellulose) or PF-06952229 at 10 (group 2) or 30 (group 3) mg/kg twice a day using an intermittent (7-day on/7-day off/cycle) schedule and at 30 (group 4) mg/kg twice a day using a daily dosing schedule. One measure of treatment efficacy was MTV on day 18, the last day all evaluable animals were alive in group 1. The figure shows the progression of MTV over 18 days in the various treatment groups. Data were analyzed using one-way ANOVA followed by Dunnett’s multiple comparison test. **P < 0.001 compared with MTV in the vehicle group.
- Fig. 2.
Effect of various PF-06952229 dosing regimens on survival over 63 days in the syngeneic MC38 colon carcinoma mouse model using the Kaplan–Meier survival curve. Female C57BL/6 mice bearing established MC38 tumors (mean volume ∼100 mm3) were sorted into four groups (n = 10 mice/group) and dosed by oral gavage with vehicle (group 1, 0.5% methyl cellulose) or PF-06952229 at 10 (group 2) or 30 (group 3) mg/kg twice a day using an intermittent (7-day on/7-day off cycle) schedule and at 30 (group 4) mg/kg twice a day using a daily dosing schedule. The figure shows the Kaplan–Meier survival curve (% survival) over 63 days.
- Fig. 3.
PF-06952229-mediated target engagement (pSMAD2 inhibition) in CD1 mouse splenocytes. A pSMAD2 time course was determined using WB analysis in CD-1 mice splenocytes. CD-1 mice (three mice/time point) were orally dosed with vehicle (two time point spleen collection, 0.5 and 24 h postdose) or PF-06952229 at 10 or 30 mg/kg and spleens were collected over a period of 24 h (at 0.5, 1, 2, 4, 8, 12, and 24 h) postdose. PF-06952229-mediated pSMAD2 and SMAD2 inhibitions were compared with vehicle-treated control values; pSMAD2 and SMAD levels were determined by densitometry and image quantitation.
- Fig. 4.
Representative photomicrographs of the femur physeal cartilage at the end of dose in the GLP rat study. SD rats (both sexes) were dosed with vehicle (left) or PF-06952229 (right, 20 mg/kg twice a day, 40 mg/kg/d) on an intermittent dosing schedule, 5-day on/5-day off cycle (5 cycles + 3 dosing days; 28 dosing days and 25 off days; end of dose on day 53). Hematoxylin and eosin staining of microscopic sections from female rats (also observed in males) demonstrate increased hypertrophy (width of the white arrow) in the femur physeal cartilages for the PF-06952229-treated (right) but not the vehicle-treated animals (10x magnification).
- Fig. 5.
Representative photomicrographs of the heart and proximal aorta at the end of dose in the GLP rat study. Sprague–Dawley rats (both sexes) were dosed with vehicle (left) or PF-06952229 (right, 20 mg/kg twice a day, 40 mg/kg/d) on an intermittent dosing schedule, 5-day on/5-day off cycle (5 cycles + 3 dosing days; 28 dosing days and 25 off days; end of dose on day 53). Hematoxylin and eosin staining of microscopic sections from the heart and proximal aorta in female rats (similar observation in males) show no adverse findings (valvulopathy or arteriopathy) in the vehicle (left) or PF-06952229-treated (right) rats (2x magnification).
- Fig. 6.
Representative photomicrographs of the femur physeal cartilage at the end of dose in the GLP cynomolgus monkey study. Cynomolgus monkeys (both sexes) were dosed with vehicle (left) or PF-06952229 (right, 100 mg/kg twice a day, 200 mg/kg/d) on an intermittent dosing schedule, 5-day on/5-day off cycle (5 cycles + 3 dosing days; 28 dosing days and 25 off days; end of dose on day 53). Hematoxylin and eosin staining of microscopic sections from female monkeys (also observed in males) demonstrate increased hypertrophy (width of the white arrow) in the femur physeal cartilages for the PF-06952229-treated (right) but not the vehicle-treated animals (5x magnification).
- Fig. 7.
Representative photomicrographs of the heart and proximal aorta at the end of dose in the GLP Cynomolgus monkeys. Cynomolgus monkeys (both sexes) were dosed with vehicle (left) or PF-06952229 (right, 100 mg/kg twice a day, 200 mg/kg/d) on an intermittent dosing schedule, 5-day on/5-day off cycle (5 cycles + 3 dosing days; 28 dosing days and 25 off days; end of dose on day 53). Hematoxylin and eosin staining of microscopic sections from the heart and proximal aorta in female monkeys (similar observation in males) show no adverse findings (valvulopathy or arteriopathy) in the vehicle (left) or PF-06952229-treated (right) rats (2x magnification).
Tables
Kinases Tested for IC50 IC50 Values (compound, nM) Repeats (n) TGFβR1 (ALK5) 0.8 (0.2–4.2)a 3 ACVR1B (ALK4) 3.14b 1 MAP4K4 (HGK) 4.49 1 CEREP Secondary Pharmacology Screen Primary Profiling Follow-up Profiling Target with 50% Response in Primary Assay Assay Format Test Concentration
(μM)% Inhibition at Test Concentration Endpoint Endpoint Value (nM) Exposure Multiplesb GABAA Cl Ch (BZD site) Binding 10 61 Ki 730 16.6x PDE4D2 Enzymatic 10 76 IC50 340 7.7x P38α MAPK Enzymatic 10 66 IC50 610 13.9x VEGFR2 kinase Enzymatic 10 96 IC50 897 20.4x ACVR1B, activin A receptor type-IB; BZD, benzodiazepine; GABA ClCh, γ-aminobutyric acid chloride channel; MAPK, mitogen-activated protein kinase; PDE4D2, phosphodiesterase 4D2; VEGFR, vascular endothelial growth factor receptor.
↵a 95% geometric confidence interval of 0.2 to 4.2 nM (n = 3) as measured in a LanthaScreen competition binding assay using a recombinant catalytic kinase domain and ATP site targeted fluorescent tracer.
↵b Exposure multiple relative to the predicted unbound Cmax (44.3 nM) at the projected clinically efficacious dose of 225 mg twice a day, based on efficacious exposure in MC38 nonclinical tumor model.
Parameter Unit Mouse Rat Monkey Human Liver microsomes CLint,app μL/min/mg 62 43 168 34 CLint,u μL/min/mg 276 236 767 121 CLb (pred) mL/min/kg 11 7 13 6 Eh (pred) % 12% 10% 30% 30% Hepatocytes CLint,app μL/min/M cells 16 9 101 32 CLint,u μL/min/M cells 89 51 414 161 CLb (pred) mL/min/kg 10 4 17 7 Eh (pred) % 11% 6% 39% 35% CL, clearance; CLint,app, apparent intrinsic CL; CLb (pred), total blood CL scaled from CLint,u; CLint,u, unbound intrinsic CL; Eh, hepatic extraction ratio.
Species
(N)Dose
(mg/kg)Cmax
(ng/mL)Tmax
(h)AUCa
(ng·h/mL)CL
(mL/min/kg)Vss
(L/kg)t1/2
(h)F
(%)Mouse 2/IV — — 1940 17.2 2.56 1.7 — (3b) 10/PO 1230 1 7020 — — 1.9 73 Rat 2/IV — — 1370 24.3 3.46 1.6 — (4a) 10/PO 1220 4 7300 — — — 109 Monkey 1/IV — — 1180 15.1 3.52 3.3 — (3a) 5/PO 214 4 1800 — — — 40 Em-dash (—) indicates not reported.
AUC, area under the concentration time curve from zero to time of last quantifiable concentration (AUCt) or infinity (AUCinf); CL, clearance; F, oral bioavailability; IV, intravenous; PO, oral; t1/2, half-life (terminal); Tmax, time of the first occurrence of Cmax; Vss, volume of distribution.
↵a AUCinf for IV; AUCt for PO.
↵b Per time point.
Group N mg/kg Dosing Schedule MTV
Day 18%TGI
Day 18CR
Day 631 10 NA Twice daily to the end of the study 707 0 2 9 10 Twice daily 7/7 × 5 (day 1 one dose) 728 −3 1 3 10/9 30 Twice daily 7/7 × 5 (day 1 one dose) 100 86 7 4 10/6 30 Twice daily to end of study 106 85 5 PF-06952229 was orally dosed twice daily on a 7-day on/7-day off cycle for up to five cycles with the first day (day 1) of each cycle receiving one dose (twice daily 7/7 × 5, day 1, one dose), otherwise twice daily to the end of the study.
Day 18 analysis: [(MTV in vehicle – MTV in treatment groups)/MTV in vehicle)] × 100.
Day 63 analysis: criteria for group CR response: tumor volume < 13.5 mm3 for three consecutive measurements, weight loss < 20%, and treatment-related deaths ≤ 10% during the study.
Occurrence of Heart and Cartilage Findings in Rat Studies 12-Day DRF
(5-2-5)a21-Day DRF
(7-7-7)b53-Day GLP Pivotal
(5-5) + 3cAUC μM × h (sex combined mean) 11 15 — 15 31 32 49 10 26 59 142 Cmax μM (sex combined mean) 4 47 — 2 4 5 5 0.9 2.3 5 12 Dose (mg/kg twice daily) 15 50/30 150d 10 20 20e 30 2.5 5 10 20 Femur physeal cartilage √ √ √ √ √ √ √ Femur articular cartilage √ √ Trachea cartilage NO NO NO NO √ Heart valves √ √ √ Heart aorta √ NO, not observed.
↵a Dosing schedule: 5-day on/2-day off/5-day on.
↵b Dosing schedule: 7-day on/7-day off/7-day on.
↵c Dosing schedule: 5-day on/5-day off + 3 dosing days; 28 total dosing days.
↵d Euthanized on day 3.
e Dosed once a day; all doses were administered twice a day unless specified otherwise.
√ Findings were noted.
Data Supplement
- Supplemental Data -
Supplemental Materials and Methods.
Supplemental Table 1. Duration, Schedule and Dosing Regimen in The Repeat Dose Toxicity Studies.
Supplemental Table 2. PF-06952229-Related Clinical Pathology Findings in Rat 12-Day DRF Study.
Supplemental Table 3. Summary of Clinical Pathology Findings in Rat 21-Day DRF Study.
Supplemental Table 4. Summary of Clinical Pathology Findings in Rat GLP Pivotal Study.
Supplemental Table 5. Summary of Clinical Pathology Findings in Cynomolgus Monkey GLP Pivotal Study.
- Supplemental Data -
