| In Vitro |
In vitroactivity: ARS-1620 is a novel, potent, oral and covalent inhibitor of KRASG12C with high potency and selectivity for KRASG12C. It was identified from structure-based design and can achieve rapid and sustained in vivo target occupancy to induce tumor regression. KRASG12C was recently identified to be potentially druggable by allele-specific covalent targeting of Cys-12 in vicinity to an inducible allosteric switch II pocket (S-IIP). Success of this approach requires active cycling of KRASG12C between its active-GTP and inactive-GDP conformations as accessibility of the S-IIP is restricted only to the GDP-bound state. This strategy proved feasible for inhibiting mutant KRAS in vitro; however, it is uncertain whether this approach would translate to in vivo. ARS-1620 achieves rapid and sustained in vivo target occupancy to induce tumor regression. This study provides in vivo evidence that mutant KRAS can be selectively targeted and reveals ARS-1620 as representing a new generation of KRASG12C-specific inhibitors with promising therapeutic potential.
Kinase Assay: ARS-1620 covalently inhibits KRAS (G12C) activity with high potency and atropisomeric selectivity in p.G12C mutant cancer cells. ARS-1620 rapidly engaged G12C in a concentration- and time- dependent manner consistent with its covalent mechanism of inhibition. Across a panel of cell lines harboring the mutant allele, ARS-1620 exhibited a half maximal G12C target engagement (TE50) at ~0.3 渭M and near complete engagement at 3.0 渭M after 2 hr of treatment. RS-1620 inhibits RAS-GTP and the phosphorylation of MEK, ERK, RSK, S6, and AKT in a dose-dependent and selective manner in H358 (p.G12C) but not in negative control lung cancer cell lines lacking p.G12C (A549, H460, and H441). ARS-1620 elicits sub-micromolar allele-specific potency (IC50 = 0.3 渭M; IC90 = 1 渭M). The activity of ARS-1620 is specific to the G12C allele and mediated by the covalent modification of Cys-12.
Cell Assay: cells are seeded into 24 well ULA-plates and allowed to rest overnight. Cells are then treated with DMSO or ARS-1620. After 2 days of treatment, apoptosis and cell death is measured by staining with annexinV-APC and prodidium iodide or by 70% ethanol fixation followed by FxCycle Violet staining to measure DNA content (cell cycle) and percentage of sub-diploid events by flow cytometry |
| In Vivo |
Following a single oral dose or 5 consecutive daily doses, ARS-1620 yields average peak tumor concentrations of 1.5 渭M (50 mg/kg) and 5.5 渭M (200 mg/kg), respectively, that enables significant KRASG12C target occupancy (>=70% G12C-TE at 200 mg/kg) for>24 hr. In MIAPaCa2 xenografts (p.G12C), ARS-1620 significantly inhibits tumor growth (p<0.001) in a dose-dependent manner with marked regression at a dose of 200 mg/kg, given once daily. Across all tumor models employed, ARS-1620 is well tolerated over the entire 3-week treatment period. Moreover, there are no observed clinical signs or toxicity of ARS-1620 in CD-1 mice even at oral doses up to 1,000 mg/kg administered daily over a 7-day period |
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