The mystery of titan hunter: Rationalized striking of the MAPK pathway via Newly synthesized 6-Indolylpyridone-3-Carbonitrile derivatives.

MAPK pathway sparkles with RTK activation, passes through subsequent downstream RAS-RAF-MEK-ERK signaling cascades, with consequent direct and indirect CDK4/6 signaling activation, and ends with cell survival, division, and proliferation. However, the emergence of anomalies such as mutations or overexpression in one or more points of the pathway could lead to cancer development and drug resistance. Therefore, designing small inhibitors to strike multitudinous MAPK pathway steps could be a promising synergistic strategy to confine cancer. In this study, twelve 6-indolylpyridone-3-carbonitrile candidates were synthesized and assessed in vitro for antineoplastic activity using four cancer cell lines. The initial antiproliferative screening revealed that compounds 3g, 3h, and 3i were the most potent candidates (GI% Avg = 70.10, 73.94, 74.33%, respectively) compared to staurosporine (GI% Avg = 70.99%). The subsequent safety and selectivity assessment showed that 3h exhibited sub-micromolar inhibition against lung cancer cells (HOP-92 GI50 = 0.75 μM) and 13.7 times selectivity toward cancerous cells over normal cells. As a result, 3h was nominated for deep mechanistic studies which evidenced that compound 3h impressively blocks multiple keystones of the MAPK pathway with nanomolar potency (EGFRWT IC50 = 281 nM, c-MET IC50 = 205 nM, B-RAFWT IC50 = 112 nM, and CDK4/6 IC50 = 95 and 184 nM, respectively). Surprisingly, 3h showed a remarkable potency against mutated EGFR and B-RAF, being 4 and 1.3 more selective to the mutated enzymes over the wild-type forms (EGFRT790M IC50 = 69 nM and B-RAFV600E IC50 = 83 nM). Ultimately, combined molecular docking and molecular dynamics (MD) calculations were executed to inspect the mode of binding and the complex stability of 3h towards the keystones of the MAPK pathway.