Alterazione di pathway epigenetici come meccanismo di resistenza ad Imatinib in una linea cellulare di CML

CML is a myeloproliferative disorder resulting from polyclonal stem cell expansion. The standard treatment is imatinib (IM). Despite the success of IM, within 18-24 months about 30% of patients develop secondary resistance. The aim of my project was to investigate possible genetic and epigenetic mechanisms (as the deregulation of miRNAs and aberrant DNA methylation), to determine how they can contribute to the resistance mechanism. Cell cultures of K562 resistant to IM (0.05 -3 μM) were set up. MiRNA, RNA and DNA were isolated. The miRNAs were analysed with a preformed tool to identify a profiling of the resistance process, while for mRNA alterations in the expression of the genes involved in the transport of drugs were sought. Regarding DNA, we analysed how methylation levels vary during the development of resistance by analyzing over 850,000 CpG sites. From the analysis of the drug transporters, it has emerged that many of the superfamily genes of the ABC transporters are overexpressed in the cells that have acquired resistance. Among these, worthy of note are ABCG2, ABCA3 and ABCC1. Comparing the miRNA expressions to the different concentrations with untreated, we observed that 6 miRNAs are significantly deregulated: miR-193b-3p, miR-486-5p, miR-512-3p, miR-517a-3p, miR-365a -3p, miR-372-3p. These miRNAs modulate genes belonging to the ErbB signaling pathway, involved in the processes of modulation of cell viability, apoptosis, and tumorigenesis mechanism. Regarding methylation, it has been observed that the number of methylated genes increases considerably and the PTPRF, TP73, ARHGEF10, FHDC1, DUSP6, PLD6 and MIR548H4 genes are significantly hypermethylated in resistant cells. Given the recent attention to the role of epigenetic mechanisms in the onset of resistance, it is possible that a genetic and epigenetic profiling, which takes into account how the efflux transporters, miRNAs and DNA methylation interact, can represent a carried out for target therapy.