Thienopyrimidine derivatives are well-known in various pharmaceutical and biomedical domains. In this work, a series of efficacious and novel strategies for the synthesis of thienopyrimidines starting from 2-amino-3-carboethoxy-4,5,6,7-tetrahydrobenzo[b]thiophene has been developed. The regioselectivity of these synthetic strategies was investigated using density functional theory computation and spectroscopic analysis. In addition, the chemical structures of the new thienopyrimidines and their reaction mechanisms, as well as the biological activity of selected compounds were studied.