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The firs method to make human iPSC used a retroviral vector delivery system, carrying the risk of transgene reactivation and insertional mutagenesis [1]. Since then many other groups have used the same methods to reprogram cells to pluripotency [2-4] among others. After that research efforts focused on searching different ways to induce pluripotency without suffering genetic changes in order to prevent transgenes from reactivating and avoiding the risk of genomic recombination or insertional mutagenesis. Some of these methods are; non-integration adenoviruses [5], expression plasmids [6], episomal vectors [7], piggy Bac transposition [8], Sendai virus [9], direct delivery of reprogramming proteins [10], synthetic modified mRNAs [11], chemical compounds [12] and synthetic self-replicative RNA replicons [13]. All though a wide range of methods have successfully reprogrammed somatic cells to a pluripotent state only three methods appear to be appropriate for reprogramming patient cells for cellular therapy: Sendai virus, mRNA and episomal vectors. (1) Sendai virus is a powerful and transient gene expression vector, with the advantages of wide host specificity and low pathogenicity, but with the worrying disadvantage of strong immunogenicity response [9]. (2) Direct transfection of synthetic modified mRNA is a strategy that administrates mRNA modified to overcome innate antiviral immune responses. It presents the best option for future clinical applications because you can control the dose and has transient expression over 48 hours [11]. (3) Episomal vector reprogramming consists on introducing episomal genes that are expressed and replicate when the host cell divides and the episome is naturally lost when the iPSCs multiply [7].
Interestingly, the first human clinical study approved for iPSC transplantation therapies (RPE cells) in Japan, headed by the stem cell researcher Masayo Takahashi, uses the episomal vector strategy. All three methods eliminate the risk of genomic integration and insertional mutagenesis, are conceivable from the technical, scientific and ethical point of view. In recent years there has been more of a focus to make the reprogramming procedure more efficient rather than improving the quality of iPSC. Recent work by Rais et al has solved the challenge of inefficiency of reprogramming with the discovery that the knockdown of the epigenetic modifier, methyl-binding protein 3 (Mbd3) in combination with provision of the four factors Oct4, Sox2, Klf4 and c-Myc results in almost 100% of somatic cells reprogramming into iPSC [14]. However, as pointed out by the authors, whether Mbd3 knockdown increases the quality of iPSC remains to be tested. The quality of differentiated cell types to be used for cell replacement therapy is dependent on the quality of the starting material. To develop high quality iPSC, the best method will most likely be to use modified RNA transfection methods using Oct4 and Sox2 with new pluripotency factors in a cell type that removes the use of oncogenic Klf4 and c-Myc, and using defined media that are xeno free, with GMP grade cell culture conditions [15-17]. The development of a protocol to make safe GMP-grade iPSC does not exist at the time of writing this review.
Citation: Requena J, Palomo ABA, Sal MF, Christodoulou J, Canals JM, et al. (2014) The Future Challenges for the Clinical Application of Reprogrammed Cells. Human Genet Embryol 4:120. doi: 10.4172/2161-0436.1000120
