Megakaryocytes (MKs) in adult marrow produce platelets that play important roles in blood coagulation and hemostasis. Monoallelic mutations of the master transcription factor gene RUNX1 lead to familial platelet disorder (FPD) characterized by defective MK and platelet development. However, the molecular mechanisms of FPD remain unclear.

Inhibition of NOTCH4 by genetic approach or chemical inhibitors enhances MK production from human iPSCs and cord-blood CD34⁺ cells. In a study published in Blood, the research group led by Dr. WANG Qianfei from Beijing Institute of Genomics of Chinese Academy of Sciences, and Dr. CHENG Linzhao from Johns Hopkins University School of Medicine, revealed a previously unappreciated role of NOTCH4 in human MK regeneration.

The study reported that NOTCH4 is a RUNX1 direct target whose expression is negatively regulated by RUNX1 during human megakaryopoiesis. It also suggested that human iPSCs with monogenic mutations have the potential to serve as an invaluable resource for discovery of novel druggable targets. The medical use of the new target NOTCH4 and its inhibitor has been applied for domestic and international patents.

Production of MKs from the FPD-derived induced pluripotent stem cells (iPSCs) was reduced, and targeted correction of the RUNX1 mutation restored MK production. In this study, Dr. WANG’s lab utilized isogenic pairs of FPD-iPSCs and the MK differentiation system, combining integrative genomic analysis, to identify two gene sets whose transcription is either up- or downregulated by RUNX1 in mutation-corrected iPSCs.

Notably, NOTCH4 expression was negatively controlled by RUNX1 via a novel regulatory DNA element within the locus, and specific inactivation of NOTCH4 by an improved CRISPR-Cas9 system in human iPSCs enhanced megakaryopoiesis. Moreover, small molecules known to inhibit Notch signaling promoted MK generation from normal human iPSCs and postnatal CD34+ hematopoietic stem and progenitor cells.

This work has potential applications in regeneration of MKs or platelets in vitro. MKs and platelets play important roles in clinical infusion and treatment of a variety of blood coagulation dysfunction diseases, including thrombocytopenia, post-chemotherapy, surgery, trauma, etc.

This study was supported by grants from the External Cooperation Program of BIC, Chinese Academy of Sciences, the National Natural Science Foundation of China and so on.

Figure: Identify new target NOTCH4 for improving megakaryocyte regeneration ex vivo. (Image by WANG's group)