Abstract

T cells used in immune cell therapy, represented by T cell receptor therapy (TCR-T), are usually activated and proliferated in vitro and are induced to a terminally differentiated phenotype, with limited viability after transfusion back into the body. T cells exhibited a robust proliferative potential and in vivo viability in the early stages of progressive differentiation. In this study, we identified microRNAs that regulate T cell differentiation. After microRNA sequencing of the four subsets: Naïve T cells (T N ), stem cell-like memory T cells (T SCM ), central memory T cells (T CM ）, and effector memory T cells (T EM ), miR-150 was identified as the most highly expressed miRNA among the four subsets and was lowly expressed in the T SCM cells. We predicted the target genes of miR-150 miRNA and performed Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes analyses. We observed that the target genes of miR-150 were enriched in pathways associated with T -cell differentiation. FOXP1 and RC3H1 were identified as key target genes of miR-150 in the regulation of T -cell function. We examined the effects of miR-150 on the differentiation and function of healthy donor T -cells. We observed that miR-150 overexpression promoted T -cell differentiation to effector T -cells and effector memory T -cells, enhanced apoptosis, inhibited cell proliferation and increased secretion of pro-inflammatory cytokines such as IFN-γ and TNF-α. In addition, the expressions of early differentiation-related genes ( ACTN1 , CERS6 , BCL2 , and EOMES ), advanced differentiation-related genes ( KLRG1 ), and effector-function-related genes ( PRF1 and GZMB ) were significantly decreased after overexpression of miR-150. Collectively, our results suggested that miR-150 can promote progressive differentiation of T cells and the downmodulation of miR-150 expression while performing a