Transcriptional drug repositioning enhances the development of somatic cell nuclear transfer embryos by inhibiting CDK4/6 during the first cleavage of bovine pseudo-zygotes.

Since the birth of the first cloned sheep, "Dolly", somatic cell nuclear transfer (SCNT) has emerged as a valuable technique in agricultural and biomedical research. However, its low efficiency remains a challenge, partly due to errors in nuclear reprogramming. In this study, we employed a transcriptional drug repositioning approach to enhance SCNT efficiency. We generated a differential gene expression profile comparing SCNT embryos with in vitro fertilized (IVF) embryos, identifying key reprogramming-associated genes. Using this blueprint, we screened small molecules that target these genes, leading to the selection of Palbociclib, a selective CDK4/6 inhibitor, as a high-ranking candidate. We hypothesized that treating reconstructed oocytes during the first cleavage with Palbociclib could prolong the first cell cycle, favorably impacting reprogramming by extending the fibroblast cell's exposure to reprogramming factors in the oocyte. Treatment with 100 nM Palbociclib for 12 h significantly extended the first cleavage duration, increased cleavage rates at 72 h, and enhanced the developmental competence of SCNT embryos compared to IVF embryos. Furthermore, gene expression analysis of blastocysts revealed that Palbociclib-treated SCNT embryos upregulated the expression of key pluripotency markers (NANOG and POU5F1) and trophectoderm marker (TEAD4), indicating improved embryonic quality. This novel study reveals that a transcriptional drug repositioning strategy can effectively enhance bovine SCNT efficiency in vitro, highlighting a promising approach to improving reprogramming and embryo development.