After myocardial infarction, a large number of cardiomyocytes die as a result of ischemia and hypoxia. Because the proliferative capacity of adult mammalian cardiomyocytes is extremely limited, the damaged myocardium is often replaced by fibrotic scar tissue, ultimately leading to impaired cardiac function and even heart failure. Therefore, how to reactivate the proliferative potential of adult cardiomyocytes and promote cardiac regeneration remains a major scientific challenge in the field of cardiovascular regenerative medicine. Small-molecule drugs, which offer advantages such as controllable biological effects and flexible routes of administration, provide a promising new avenue for activating endogenous myocardial regeneration.

Professor Xiong Jingwei’s team at School of Basic Medical Sciences previously identified a five-small-molecule combination, termed 5SM, through small-molecule drug screening. The combination consists of phenylephrine hydrochloride (PE), baricitinib, harmine (HM), VO-OHpic trihydrate, and AZD3965. The study showed that 5SM promotes cardiomyocyte proliferation, improves cardiac function, and reduces myocardial fibrosis in adult rat models of myocardial infarction and ischemia-reperfusion injury. The team subsequently published related findings in the Journal of Molecular and Cellular Cardiology, revealing that TGFβ/BMP-SMAD4 signaling is one of the key mechanisms through which 5SM promotes cardiomyocyte proliferation and cardiac regeneration.

Given the relatively large number of components in 5SM, which increased the complexity of mechanistic investigation and subsequent translational development, the team further simplified the cocktail and investigated the synergistic interactions among its components. Recently, the team published a research article in Cell Regeneration entitled “Small-molecule cocktail 2SM promotes heart regeneration via inducing embryonic-like cardiomyocyte state.” In this study, the researchers identified a two-small-molecule combination, 2SM, consisting of PE and HM, from the original 5SM cocktail and elucidated the mechanism by which the two compounds act synergistically to promote cardiomyocyte proliferation.

The study found that 2SM promotes cell-cycle re-entry and completion of cell division in neonatal mouse and rat cardiomyocytes, while also enhancing cell-cycle activity in human induced pluripotent stem cell-derived cardiomyocytes. In an adult mouse myocardial infarction model, 2SM promoted cardiomyocyte cytokinesis. In an adult rat myocardial infarction model, 2SM enhanced cell-cycle activity in cardiomyocytes in the infarct border zone, improved cardiac function, and reduced infarct size.

Mechanistic studies revealed that PE and HM act at distinct stages of the cell cycle, producing a temporally coordinated synergistic effect. PE induces a fetal-like transcriptional program and activates mTORC1 signaling, thereby promoting cardiomyocyte progression into the G1/S phase, while simultaneously causing G2/M arrest. HM, in turn, relieves this arrest by inhibiting DYRK1A and activating the MMB/FOXM1-MuvB-associated transcriptional program, enabling cardiomyocytes to complete mitosis and cytokinesis. Sequential treatment with PE followed by HM was more effective in promoting cardiomyocytes through a complete cell cycle.

Transcriptomic and metabolomic analyses further showed that 2SM-treated cardiomyocytes exhibited activation of cell-cycle programs, enhanced glycolysis, and reduced fatty acid metabolism. These findings indicate that 2SM induces an embryonic-like state in cardiomyocytes and promotes cardiomyocyte proliferation through the coordinated regulation of PE and HM at distinct stages of the cell cycle.

This study streamlined the original 5SM cocktail to a two-small-molecule combination comprising PE and HM. While retaining its ability to promote cardiomyocyte proliferation and cardiac regeneration, 2SM also enabled further elucidation of the mechanism by which the two small molecules synergistically regulate distinct stages of the cell cycle. These findings provide a new foundation for optimizing small-molecule-based strategies for cardiac regeneration.

Wang Zihao, doctoral student (already graduated) of Peking University, is the first author of the paper. Associate Research Professor Zheng Lixia and Professor Xiong Jingwei at the School of Basic Medical Sciences, the Second Affiliated Hospital, and the Institute of Biomedical Innovation at Nanchang University are the co-corresponding authors.

Original article: https://doi.org/10.1186/s13619-026-00301-0