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Objective: The objective of this study was to synthesize a series of 1,3,4-thiadiazole derivative and evaluate their possible in vitro and in vivo anticancer activity. Method: Compounds 7a-f have been synthesized by chlorination of 2-[bis(2-hydroxyethyl)amino]-N-[5- substituted phenyl)-1,3,4-thiadiazol-2-yl]acetohydrazide with phosphorous oxychloride and phosphorous pentachloride. Synthesis of the targeted compounds was (7a-f) confirmed through their MP, IR, 1HNMR and mass spectrum studies. Results: Compounds 7a and 7d showed considerable anticancer activity against Hep-2 cell lines and, therefore, selected for the in vitro studies and DLA cell line for the in vivo studies. Conclusion: It may be concluded that the presence of nitro and chloro phenyl substituted 1,3,4- thiadiazoles showed significant anticancer activity.