Novel thienoquinoline carboxamide-chalcone derivatives were prepared via the cyclization of acylated chalcones and 2-mercaptoquinoline-3-carbaldehyde in DMF with K2CO3. Thienoquinolines 9a-f, h exhibited promising antiproliferative effect against all the tested cell lines and gave a significant activity as EGFR inhibitors, with IC50 values ranging from 0.5 and 3.2 µM, and compounds 9e and 9f being the most active of the series. They also showed better activity than Erlotinib against melanoma cancer cell line A375. Moreover, compound 9f influenced pre G1 apoptosis and cell cycle arrest at G2/M phase. The binding mode of the best EGFR inhibitor 9e in the EGFR active site revealed that the thienoquinoline ring occupied the ATP-binding site while the chalcone moiety is located in the allosteric site and is responsible for the enhanced activity of these compounds.