在现代合成方法和先进的生物评估技术时代，考虑到肝细胞癌（HCC）的臭名昭著历史，对于新型生物活性化合物的期望大幅增长。在药物发现研究中广泛应用的多种主题包括异喹啉和噻吩[2,3-b]吡啶。本文中，将这两种主题分子合并，形成噻吩[2,3-c]异喹啉，作为一种新型抗增生的生物活性化合物，并针对HCC进行了深入研究。因此，合成了4、5、7和8号化合物系列，并对HepG2细胞系进行了生物评估。通过生物研究，成功确定了C7-Ac/C8-OH取代基、C8-C9不饱和度、C1-NH2的1H-吡咯-1-基环闭合和C6-Ph p-卤代取代基的作用，并成功得到了安全的5b先导物。进一步，通过流式细胞术和Annexin V-FITC / PI凋亡生物研究，发现5b显著抑制G2 / M期细胞周期，并且凋亡增加了60倍。采用DFT构象研究和分子对接技术，结合分子力学/广义Born表面积评分发现，5b在可可碱结合位点具有潜在的微管目标作用，实验证明了这一点（可可碱抑制性IC50 = 71µM，而可可碱为14µM）。因此，在保留C7-乙酰和优化卤位置的同时，保留[6S，7R]-立体化学对于最佳结合微管的可可碱结合位点至关重要。 ©2023 Wiley Periodicals LLC.

In the era of modern synthetic methodology and advanced bio-evaluation techniques and considering the notorious history of hepatocellular carcinoma (HCC), hopeful expectations regarding novel bioactive chemotypes have grown dramatically. Among the widely versatile motifs in drug discovery studies are isoquinoline and thieno[2,3-b]pyridine. Herein, the molecular merging of both motifs evoked thieno[2,3-c]isoquinoline as a novel antiproliferative chemotype being hardly studied against HCC. Accordingly, compound series 4, 5, 7 and 8 were synthesized and bioevaluated against the HepG2 cell line. The role of C7-Ac/C8-OH substituents, C8-C9 unsaturation, 1H-pyrrol-1-yl ring closure at C1-NH2 and C6-Ph p-halo-substitution were biologically studied and successfully furnished the lead 5b while showing safe profile against Vero cells. Further, flow cytometric and Annexin V-FITC/PI apoptotic bio-investigations of 5b unveiled remarkable cell cycle arrest at the G2/M phase besides a 60-fold increase in apoptosis. The use of a DFT conformational study followed by Molecular docking and molecular mechanics/generalized born surface area scoring evoked potential tubulin-targeting activity of 5b at colchicine-binding site, which was confirmed by experimental evidence (Tub Inhib IC50  = 71 µM vs. 14 µM for colchicine). Accordingly, preserving C7-acetyl and optimizing halogen position while preserving [6S,7R]-stereochemistry is crucial for optimum binding to colchicine binding site of tubulin.© 2023 Wiley Periodicals LLC.