The ability to “reprogram” differentiated adult cells to a state that resembles embryonic stem cells has created wide-ranging opportunities for development of relevant in vitro disease models and patient-specific cell-replenishment therapies.
Initial efforts to generate human induced pluripotent stem cells (iPS cells) required simultaneous co-infection of cells with four separate retroviral expression vectors (Oct-4, Klf4, Sox-2, and c-Myc). Each vector carried one transcription factor, which resulted in a high number of genomic integrations.
Alternative approaches to iPS generation have included use of plasmids and nonintegrating adenovirus vectors to deliver the transcription factors. The rates at which cells convert to pluripotency using these methods, however, are far lower than those obtained using retroviral vectors.
Generation of human and mouse iPS cells is now routinely accomplished using a single, excisable polycistronic lentiviral vector that delivers all four Yamanaka transcription factors. EMD Millipore's STEMCCA™ lentiviral-based reprogramming yields cells that form multilayered tightly packed colonies with well-defined borders. These cells stain positive for alkaline phosphatase, express embryonic stem cell markers, form embryoid bodies, and differentiate into all three germ layers.
Use of a single vector significantly reduces the number of viral integrations required—in some cases, iPS clones possessing only a single viral integrant can be isolated.
Even with the use of a single vector, however, reprogramming human somatic cells remains a highly inefficient and time-consuming process. Small molecules targeting specific signaling pathways are being investigated for their ability to enhance reprogramming and/or replace the transcription factors required for reprogramming.
In this study, chemical compounds were screened for their effects on increasing the ratio of fully reprogrammed SSEA4+TRA-1-60+ Hoechst dim iPS cells versus reprogramming intermediates, increasing colony numbers, and reducing the time to establishment of full reprogrammed iPS cell colonies.