Ricerc@Sapienza

Over time, an understanding of the molecular mechanisms of replication and differentiation of stem cells has enabled the development of various protocols and techniques to be applied with a regenerative scope, as discussed in previous chapters. Here, we will discuss the recent progress of regenerative medicine in chemical reprogramming and transdifferentiation based on the use of epi-drugs. The main epigenetic mechanisms regulating gene expression include DNA methylation and histone tails modifications. Briefly, DNA methyltransferases (DNMTs) are S-adenosyl methionine (SAM)-dependent enzymes catalyzing the addition of a methyl group at the C5 of the cytosine ring of DNA. DNA methylation is generally seen as a mark of gene silencing, which is often reinforced by other heterochromatin-associated marks. More recently, ten-eleven translocation (TET) enzymes, able to oxidize a 5-methyl-cytosine onto a 5-hydroxymethyl-cytosine, have been also described. Histones also undergo cell type-specific posttranslational modifications that influence the degree of chromatin condensation, and biologically modulate and stabilize transcriptional output by recruiting coactivators and repressors. The most important histone modifications include acetylation, phosphorylation, methylation, ubiquitination, and SUMOylation. Specifically, histone methyltransferases catalyze the mono-, di- or trimethylation of the terminal amino or guanidine group of lysines or arginines, respectively. Histone methylation levels are regulated by the activity of histone demethylases, and the transcriptional effect of this mark can vary based on the location of the methylated residue or on the degree of methylation. The balanced activity of histone deacetylases (HDACs) (including both Zn2+-dependent/classical HDACs and nicotinamide adenine dinucleotide (NAD+)-dependent HDACs/sirtuins) and HATs defines the levels of histone lysine acetylation, a mark of gene activation. A number of drugs targeting various epigenetic players are currently available, and some of them have entered clinical trials for the treatment of a number of disorders. Generally, protocols applying epigenetic modulators, kinases inhibitors, and cytokines in sequence and/or in specific combinations to best mimic the physiological environment have been applied. One of the main advantages of epigenetic drugs is that their action is reversible. Hence they can be used before transplantation of stem or progenitor cells as a functional boost without the risk of geneticmodification. Nevertheless their effect on the epigenome is global,whereas themodifications required for cell reprogramming are generally focused at certain loci. In this way, these epigenetic modulators could also activate genes with undesirable functions and generate a global epigenetic disturbance leading to malignant transformations (teratomas). More recently this issue has been overcome by using protocols to obtain various types of differentiated cells from other terminally differentiated cells by means of chemical transdifferentiationwithout passing through a pluripotent state. The use of chemical reprogramming in applying epi-drug treatment is an emerging strategy in regenerative medicine (only one clinical trial is running). Despite this, it appears quite promising. The use of epi-drugs in regenerative medicine holds the promise of an increasingly personalized medicine, pointing to a future where patient-derived cells can be specifically reprogrammed using the appropriate cocktail for the specific regenerative scope in question. In this chapter we will discuss the current knowledge regarding the use of epi-drugs in the regeneration of a variety of tissues and organs.