Design, synthesis, and preliminary antiproliferative evaluation of azetidine-functionalized pyrrolo[2,3-d]pyrimidines...

Azetidine-functionalized pyrrolo[2,3-d]pyrimidines were designed to examine the comparative influence of methyl-, propyl-, and butylsulfonyl substitution among final synthesized analogues on antiproliferative activity and structure-based behavior. Following in silico ADME filtering and docking-based prioritization, selected analogues were synthesized and characterized. Among the evaluated compounds, BCD17 showed the most favorable predicted docking profile toward 17β-hydroxysteroid dehydrogenase type 1 (3HB5), with a docking score of -9.130 kcal/mol and Glide energy of -69.236 kcal/mol. Molecular dynamics simulation and MM-GBSA analysis further supported the stable predicted interaction profile of BCD17, with a mean binding free energy of -109.49 ± 7.52 kcal/mol. DFT analysis indicated a balanced electronic profile compatible with its predicted structure-based performance. In vitro MTT evaluation confirmed antiproliferative activity in MCF-7 cells, with IC₅₀ values of 0.76, 0.75, and 1.83 µM for BCD1, BCD9, and BCD17, respectively. Although BCD1 and BCD9 showed slightly lower IC₅₀ values, BCD17 was prioritized based on its integrated computational stability, apoptosis-related response, and preliminary selectivity profile. MCF-7 cell-cycle and Annexin V/PI apoptosis analyses further supported the antiproliferative and pro-apoptotic effect of BCD17, although direct 17β-HSD1 target engagement was not experimentally confirmed. In MCF-10A cells, BCD9 and BCD17 showed comparatively lower toxicity than 5-FU. Overall, BCD17 emerged as a promising integrated lead candidate for further optimization and biological validation.