Figure 6. Pharmacological inhibition of PTPN11 in mouse melanoma cells with NRAS ^Q61K induces tumor regression in part through regulation of GSK3 and cyclin D1.

Survival analysis of 2187 and 5037 (mouse melanoma cells with Tyr-rtTA; tetO-NRAS ^Q61K; Cdkn2a (Ink4a/Arf) ^−/−) following treatment with increasing concentrations of SHP099 at 72 hours (A). Decreased tumor volume following treatment of established subcutaneous allograft 5037 tumors with SHP099 (PTPN11 inhibitor, 100 mg/kg; po qd) compared to treatment with vehicle alone. Data shown as mean tumor volume ratio to day 0 +/− SEM (B) and final tumor weight at harvest (p=0.0003; two-tailed t-test) on day 8 (C). Waterfall plot demonstrating ratio of D8 to D0 tumor volume for tumors treated with vehicle or SHP099. SHP099 treated tumor segregated into (red) responders and partial-responders (blue) with and without regression, respectively (D). Western analysis of 5037 subcutaneous tumors with (+) or without (–) SHP099 treatment for 3 or 8 days (E). Western analysis of parental 5037 and a cell line established from a partial responding tumor (PR1; blue in panel D) treated with 0, 10, or 30 uM SHP099 for 3 hrs (F). Cell survival was assessed at 72 hrs of post-treatment as the mean +/−SD survival (relative to DMSO or GSK3β inhibitor CHIR-99021 alone) (G&H). The response of 5037 with vector control or GSK3β-Y216A mutant and PR1 cells to SHP099 (G) and that of 5037 parental cells to increasing concentrations of SHP099 alone or in combination with 1μM or 3μM CHIR-99021 (H) were shown.