Synthesis of D‑Ring Annulated Pyridosteroids from β‑Formyl Enamides and Their Biological Evaluations

Herein, we report the synthesis of a novel class of substituted androst­[17,16-b]­pyridines (pyridosteroids) from the reaction of β-formyl enamides with alkynes in high yields. The optimized reaction protocol was extended to acyclic and cyclic β-formyl enamides to afford nonsteroidal pyridines. Cell survival assay of all compounds were carried against prostate cancer PC-3 cells wherein 3-hydroxy-5-en-2′,3′-dicarbethoxy-androst­[17,16-b]­pyridine showed the highest cytotoxic activity. Phase contrast microscopy and flow cytometry studies exhibited marked morphological features characteristic of apoptosis in 3-hydroxy-5-en-2′,3′-dicarbethoxy-androst­[17,16-b]­pyridine and abiraterone treated PC-3 cells. The treatment of 3-hydroxy-5-en-2′,3′-dicarbethoxy-androst­[17,16-b]­pyridine induces G2/M phase cell cycle arrest in prostate cancer PC-3 cells. Enhancement of apoptotic inductions of PC-3 cells by 3-hydroxy-5-en-2′,3′-dicarbethoxy-androst­[17,16-b]­pyridine and abiraterone through the activation of caspases-6, -7, and -8 pathways were supported by qRT-PCR. In silico study of the compound 3-hydroxy-5-en-2′,3′-dicarbethoxy-androst­[17,16-b]­pyridine showed stable and promising interaction with the key caspase proteins. Our studies revealed that the pyridosteroid 3-hydroxy-5-en-2′,3′-dicarbethoxy-androst­[17,16-b]­pyridine, bearing pyridine-2,3-dicarbethoxy pharmacophore, facilitated initiation of caspase-8 and activates downstream effectors caspase-6 and caspase-7 and thereby triggering apoptosis of PC-3 cancer cells.