Presented in Stockholm, Sweden June 25, 2015 at the Annual Meeting of the International Society of Stem Cell Researchers
A PRIMITIVE GROWTH FACTOR IS SUFFICIENT TO INDUCE STABLE NAÏVE STATE HUMAN PLURIPOTENCY VIA A LIF- AND FGF-INDEPENDENT PATHWAY; REPROGRAMMING IN THIS NOVEL GROWTH FACTOR CONFERS SUPERIOR DIFFERENTIATION
Scientists have generated human stem cells that mimic mouse naïve cells, but their dependence on the addition of several extrinsic agents, and their propensity to develop abnormal karyotype calls into question their resemblance to a naturally occurring ‘naïve’ state in humans. Here we report the discovery of a novel cancer-testis antigen that is a stem cell growth factor. This naturally occurring growth factor alone is sufficient to convert human primed state stem cells to a naïve state that is karyotypically stable, without the use of inhibitors, transgenes, LIF or FGF2. In addition, reprogramming somatic cells with transient ectopic expression of OSK or OSKM in the presence of this growth factor, and in the absence of any other growth factor or inhibitor, results in increased efficiency of hiPSC generation. Both hESCs and hiPSCs cultured in this novel growth factor are naïve, have two active X chromosomes, can be passed as single cells, can be clonally expanded and have stable karyotype for more than 30 passages. These naïve cells differentiate down all three germlines as demonstrated by teratoma formation. Further, directed differentiation to cardiomyocytes, neuronal sub-types and mature hepatocytes was superior to that of FGF-grown cells in terms of yield, expression of markers of the desired cell type. In the case of cardiomyocytes, these naïve stem cells had greater strength of contraction, increased beating frequency and synchronized beating for more than 300 days. Transcriptome analysis revealed that a group of genes that are master regulators of differentiation is down-regulated in these naïve stem cells and is regulated by super-enhancers. None of these super-enhancer regulated genes was down-regulated in the other naïve stem cells generated using FGF or LIF based methods. This suggests that in our naïve cells, a master differentiation program is turned off but, upon removal of our novel growth factor, can be rapidly activated to execute a key cell fate decision, such as whether to grow pluripotently or differentiate. RNA SEQ suggests the existence of two naïve states: an early naïve state driven by our novel growth factor, which cannot limit self-replication and a later naïve state regulated by NME1, which limits self-replication when its multimerization state shifts from the active dimer to the inactive hexamer.