Epicardial Cell Regeneration Using p21 Inhibitors

Professor Yoshida’s team discovered that the inhibition of p21, a protein regulating the cell cycle also known ascyclin-dependent kinase inhibitor 1 (CDKN1A), promotes the regenerative ability of epicardial cells. The researchers designed a siRNA sequence targeting the CDKN1A gene to inhibit its expression. They showed that CDKN1A inhibition resulted in increased proliferation of human epicardial cells and expression of regeneration-related genes suggesting its therapeutic potential for the heart.

To assess the efficacy of CDKN1A inhibitor,Professor Yoshida’s team used human epicardial cells differentiated from iPSC cells as described inJunghof et al. 2022. Following the siRNA transfection, the researchers confirmed decreased expression of CDKN1A mRNA using RT-PCR (Fig.1a) and of the protein using Western blot (Fig.1b). Next, a significant increase in the proliferative capacity and viability of the epicardial cells was observed (Fig.1c,d). Finally, CDKN1A inhibition improved wound healing (Fig.1e) and decreased the expression of quiescence-associated genes.

Figure 1. Effects ofsiRNACDKN1A inhibition on hiPSC-derived epicardial (hiPSC-EPI) cells. a) Expression change of CDKN1A in cells treated with siRNA relative to control measured by RT-qPCR b) Western blots probed with antibodies against p21, WT1, and b-actin in hiPSC-EPI cells c) Cell growth curve assay of hiPSC-EPI cells d) Colony formation assay of hiPSC-EPI cells e) Wound healing assay assessing the migratory capacity of the cells showing the wound closure (%) after 24 hours of scratching a confluent cellular monolayer.

In addition toin vitro studies, the researchers performedex vivo explant assay of CDKN1A knockout mice. Compared to controls, epicardial explants obtained from CDKN1A knockout mice showed faster growth rates and almost twice the explant length, suggesting a higher regenerative potential. Therefore, the researchers believe that inhibiting either the mRNA expression or the protein level of CDKN1A can enhance the regenerative capacity of theepicardium. The former could be achieved by delivering siRNA, shRNA, or CRISPR-Cas system using viral vectors, while antibodies or p21-binding compounds such asflavopiridol could be used to achieve the latter. Professor Yoshida’s team is looking for biotech or pharmaceutical companies to develop therapeutics based on their discovery.

Intellectual property status

Granted Patent

Patent number: WO/2023/210713

Current development status

Experimental technologies

Desired business relationship

Patent licensing