Novel 2,6-Bis(indolyl/ 5-bromoindolyl)dihydropyridine/pyridine hybrids as potential anti-tumor agents: Design, one-pot Green synthesis, molecular docking, and cytotoxicity evaluation

This work presents seventeen new bis(indolyl/5-bromoindolyl)dihydropyridine and bis(indolyl/5-bromoindolyl)pyridine derivatives synthesized using an eco-friendly, solvent-free, and catalyst-free protocol. The reaction occurred between different aldehydes and 3-(1H-indol-3-yl)-3-oxopropanenitrile (1a), or 3-(5-bromo-1H-indol-3-yl)-3-oxopropanenitrile (1b), or 1-(1H-indol-3-yl)ethan-1-one (4) in the presence of ammonium acetate via one-pot reaction cyclization. The structures of the synthesized compounds were confirmed by IR, ¹H NMR, ¹³C NMR, and elemental analysis. The synthesized compounds were evaluated for their anticancer activity against the colon (HCT-116) cancer cell line, the breast (MCF-7) cancer cell line, and normal epithelial cells (RPE-1) using the MTT assay. Compounds 2a, 2b, 2c, 2f, 2g, 2h, 2i, 2j, 2k, 2l, and 2m exhibited more antiproliferative activity (IC₅₀ values of 1.07–6.9 μM against the MCF-7 cell line and IC₅₀ values ranging from 2.69 to 6.37 μM against the HCT-116 cell line) than that of doxorubicin against the two tested cell lines (IC₅₀ = 9 μM and 7.1 μM against MCF-7 and HCT-116, respectively). Compounds 2a, 2b, 2c, 2e, 2f, 2 g, 2h, 2i, 2k, 2l, 2m, and 5 were found to be more selective against both cell lines than normal epithelial cells (RPE-1). Compounds 2b, 2f, 2i, 2j, 2k, and 2l, in addition to doxorubicin, were further subjected to an in vitro VEGFR-1 inhibition assay in the MCF-7 and HCT-116 cell lines. Compounds 2j and 2l demonstrated excellent VEGFR-1 inhibitory activity in both cell lines compared to doxorubicin (% inhibition in HCT-116 = 89.23 %, 88.9 %, and 53.85 %, respectively, and % inhibition in MCF-7 = 93.16 %, 94.89 %, and 86.9 %, correspondingly). Cell cycle analysis was performed for the most potent derivatives, compounds 2j and 2 l, which arrested MCF-7 cells at the G₀/G₁ phase with high percentages (94.18 % and 86.80 %, respectively). The expression levels of the caspase-3 gene induced by compound 2l treatment were statistically comparable to those of doxorubicin and greater than those of compound 2j. Finally, molecular docking studies were conducted to determine the essential amino acid interactions and free binding energy of these active derivatives. All our results led us to consider compound 2l as a potential lead for the battle against cancer.