Επιγενετικές τροποποιήσεις στα γονίδια των ανοσοσφαιρινών στη χρόνια λεμφοκυτταρική λευχαιμία

Abstract

Until recently it was believed that CLL is a homogeneous disease, but now several lines of evidencedemonstrate, beyond doubt, that CLL cases can be subdivided into subgroups with distinct biologicalfeatures, extending from genomic aberrations to immune signaling via receptor molecules of bothinnate (e.g. Toll-like receptors, TLRs) and adaptive nature (i.e. the B cell receptor, BcR), with the latterso far thought to play perhaps a more pivotal role. In CLL, subsets of patients with stereotyped BcRs,callled stereotyped subsets, account for one-third of the cohort. Increasing evidence suggest that casesassigned to the same subset can share similar biological and clinical features. Consequently, the greatheterogeneity observed among different CLL cases stimulates the exploration of the implicatedmechanisms. Two primary, complementary and interconnected mechanisms which regulate geneexpression are: i) microRNAs (miRNAs) and ii) DNA methylation. Aberrant miRNA and DNAmethylation profiles are observed in different types of cancer, including CLL. Considering all the above,we investigated the expression profile of certain miRNAs and DNA methylation profile in subgroups ofCLL patients defined by their distinct BcR molecular features. More specifically, we investigatedwhether miRNAs could be implicated in the control of TLR and BcR signaling in CLL, both at basallevel and after ex vivo stimulation. At basal level we profiled the expression of 33 miRNAs relevant toCLL biology using RT-qPCR arrays. Distinct miRNA expression profiles were observed among CLLsubgroups. Differentially expressed miRNAs were functionally implicated in the biology of CLL cells,since they affected BcR and TLR signaling molecules expression levels, while they led to EZH2methyltransferase overexpression in ‘’bad prognosis’’ CLL cases. Next, we performed ex vivostimulation of CLL cells, belonging to cases assigned to specific stereotyped subsets, through the BcRand/or TLRs, and studied the expression levels of the 33 miRNAs described above. We observeddifferent miRNA expression regulation after stimulation of the same TLR in different stereotypedsubsets, while there are hints for the existence of a regulatory circuit between the immunomodulatorycluster miR-17~92 and certain signaling molecules of the BcR and TLR pathways. Finally, we studiedthe DNA methylation profile in bad prognosis CLL stereotyped subsets #6 and #8 and searched forepigenetic modifications that could be linked with their distinct clinicobiological features. Stereotypedsubsets #6 and #8 both express unmutated BcRs yet exhibit different clinical behaviour. So far, noexplanation is available for this difference, thus prompting investigation into the underlyingmechanisms. Aberrant DNA methylation is increasingly recognized as relevant for CLL. With this inmind, we profiled DNA methylation with the Infinium HumanMethylation450 BeadChip array whichallows interrogation of approximately 485,000 CpG sites. We found distinct DNA methylation profiles,affecting the transcriptional programming, between the two CLL stereotyped subsets. The observeddifferential DNA methylation and differential gene expression may be implicated in differences thatshape the biologic make-up and eventual clinical behaviour of each subset.