Cyclizing-berberine A35 induces G2/M arrest and apoptosis by activating YAP phosphorylation (Ser127)

A35 is a novel synthetic cyclizing-berberine recently patented as an antitumor compound. Based on its dual targeting topoisomerase (top) activity, A35 might overcome the resistance of single-target top inhibitors and has no cardiac toxicity for not targeting top2β. In this study we further explored the biological effects and mechanisms of A35. Antitumor activity of A35 was evaluated by SRB and colony formation assay. G2/M phase arrest (especially M) and first damage of M-phase cells were investigated by flow cytometry, cytogenetic analysis, immunofluorescence, co-immunoprecipitation and WB. The key role of phospho-YAP (Ser127) in decreasing YAP nuclear localization, subsequent G2/M arrest and proliferation inhibition were explored by YAP1−/− cells, mutated Ser127 YAP construct (Ser127A) and TUNEL. G2/M arrest induced by A35 was independent of p53. M phase cells at low dose were firstly damaged and most damaged-cells accumulated in M phase, and that was a result of preferring targeting top2α by A35, as top2α is essential to push M phase into next phase, and targeting top2α induced cells arrested at M phase. A35 decreased YAP1 nuclear localization by activating YAP phosphorylation (Ser127) which subsequently regulated the transcription of YAP target genes associated with growth and cycle regulation to induce G2/M arrest and growth inhibition. Our studies suggested the mechanism of G2/M arrest induced by A35 and a novel role of YAP1 (Ser127) in G2/M arrest. As a dual topoisomerase inhibitor characterized by no cardiac toxicity, A35 is a promising topoisomerase anticancer agent and worthy of further development in future.