To explore the contribution of the carboxylic group to the antiproliferative activity of 5-aminosalicylic-4-thiazolinone-based hybrids, multiple derivatives were synthesized, including free carboxylic, hydroxamic acid, and amidation with glycine or 2-amino-5-nitrothiazole. The antiproliferative activity of these derivatives was compared with that of methyl ester, aliphatic, and aromatic amides of the hybrid against eight cancer cell lines and one normal cell line using the sulforhodamine B (SRB) assay. Among the newly synthesized compounds, glycinate amide 9b exerted good to moderate antiproliferative activity against five cancer cell lines., It displayed superior activity against the Jurkat and Lymphoma cell lines with IC₅₀ values of 4.74 µM and 7.16 µM, The overall results show that the methyl ester derivative of the hybrids is the most active among the tested derivatives against all tested cell lines with low micromolar to nanomolar potency. While hybrids with free or amidated carboxylic groups are much less potent. ADME study revealed that the difference in physicochemical properties might rationalize the activity pattern. This study demonstrates that the nature of the carboxylic group in these hybrids critically influences their antiproliferative activity. The findings pave the way for developing optimized anticancer salicylate drugs by focusing on modifications that improve key physicochemical properties.