Transient Naive Treatment (TNT) iPS cells do not feature Sendai virus expression: Response to Sendai virus persistence questions the transient naive reprogramming method for iPSC generation

SummaryThe Transient Naive Treatment (TNT) reprogramming method involves a brief period of culturing in naive media early in the reprogramming process. Through conducting extensive molecular, epigenetic, and functional analyses on numerous induced pluripotent stem cell lines, we have demonstrated that TNT reprogramming facilitates epigenetic memory erasure and produces induced pluripotent stem (iPS) cells more similar to embryonic stem (ES) cells than conventional reprogramming methods1. In 2024, De Los Angeles et al. posted a contradictory results preprint on bioRxiv titled Sendai virus persistence questions the transient naive reprogramming method for iPSC generation2. Their claims include: 1) that Sendai virus expression in naive cultured cells questions the TNT reprogramming method, 2) the possible selection of cells (clonal expansion) with Sendai virus expression during TNT reprogramming, 3) Sendai virus genes being expressed in control samples implying a potential sample mix-up. Here, we demonstrate these claims are not directly supported by the data, and emphasize that we assessed the possibility of cell sub-population selection experimentally in Buckberry et al. (2023). Moreover, our re-analyses of the data in the context of Sendai virus expression during reprogramming actually highlight the advantages of TNT reprogramming concerning the timing of the naive media treatment and the absence of detectable Sendai virus gene expression.