Stereoselective synthesis of (Z)-methyl 2-[(Z)-3-substituted-2-(2-(2,4-dinitrophenyl)hydrazono)-4-oxothiazolidine-5-ylidine)]acetates and (Z)-methyl 2-[(Z)-3-substituted-2-(2-(tosylhydrazono)-4-oxothiazolidine-5ylidine)]acetates via the cyclization of N-substituted-2-arylhydrazinocarbothioamides with dialkyl acetylenedicarboxylates in absolute ethanol is reported. The structures were confirmed by spectroscopic data as well as single crystal X-ray analyses. The mechanism of nucleophilic addition and the role of electronic factors were discussed. The anticancer activity of the newly synthesized compounds was evaluated against a panel of 60 cell lines derived from nine different types of cancers including, leukemia, melanoma, lung, colon, CNS, ovarian, renal, prostate, and breast cancers and the results revealed that the cyclohexyl derivative has a significant broad anticancer activity especially against different leukemia and colon cancer cell lines . MCCM_Template_Vers 4