Reduction of phosphoinositide-phospholipase C beta1 methylation predicts the responsiveness to azacitidine in high-risk MDS

Abstract

Lipid signaling pathways are involved in cell growth, differentiation, and apoptosis, and could have a role in the progression of myelodysplastic syndromes (MDS) into acute myeloid leukemia (AML). Indeed, recent studies showed that phosphoinositide-phospholipase (PI-PL)Cbeta1 mono-allelic deletion correlates with a higher risk of AML evolution. Also, a single patient treated with azacitidine, a DNA methyltransferase inhibitor currently used in MDS, displayed a direct correlation between PI-PLCbeta1 gene expression and drug responsiveness. Consequently, we hypothesized that PI-PLCbeta1 could be a target for demethylating therapy. First, we analyzed the structure of PI-PLCbeta1 gene promoter, then quantified the degree of PI-PLCbeta1 promoter methylation and gene expression in MDS patients at baseline and during azacitidine administration. Indeed, PI-PLCbeta1 mRNA increased in responder patients, along with a reduction of PI-PLCbeta1 promoter methylation. Also, the molecular response correlated to and anticipated the clinical outcome, thus suggesting that PI-PLCbeta1 gene reactivation could predict azacitidine responsiveness. Our results demonstrate not only that PI-PLCbeta1 promoter is hypermethylated in high-risk MDS patients, but also that the amount of PI-PLCbeta1 mRNA could predict the clinical response to azacitidine, therefore indicating a promising new therapeutic approach. myelodysplastic syndromes ͉ real-time PCR ͉ signal transduction ͉ inositides ͉ epigenetic therapy

Fas expression at diagnosis as a biomarker of azacitidine activity in high-risk MDS and secondary AML

“…Indeed, azacitidine favors the re-expression of FAS in neoplastic mast cells or PI-PLCbeta1 genes in MDS or sAML. 7,12 Here we show that the in vivo use of the drug also favors the re-appearance of Fas receptor in sAML-derived CD45 lo /CD34 þ cells, together with FAS promoter demethylation (Supplementary Figure S1).…”

Synergistic induction of PI-PLCβ1 signaling by azacitidine and valproic acid in high-risk myelodysplastic syndromes

“…24,25 PI-PLCb1 has also been demonstrated to have a role both in the genetic 26 and in the epigenetic mechanisms of MDS progression to AML. 27,28 As for this latter, recent studies showed not only that PI-PLCb1 promoter gene is hypermethylated in high-risk MDS patients but also that it can be a specific target for AZA. Indeed, the amount of PI-PLCb1 messenger RNA could predict the clinical response to AZA, therefore indicating a promising new tool in the evaluation of response to demethylating therapies.…”

“…The degree of PI-PLCb1 promoter methylation was quantified by a standard SYBR-Green Real-Time methylation-specific PCR method using the glyceraldehyde-3-phosphate-dehydrogenase as a reference gene, as previously described. 28 …”

“…These two cell lines were used, respectively, as a positive and negative control for PI-PLCb1 methylation, as previously described. 28 In all in vitro experiments, AZA was administered at 5 mM for 24 h, whereas cells were exposed to VPA for 24 h at a final concentration of 0.3 mM.…”

“…Indeed, the amount of PI-PLCb1 messenger RNA could predict the clinical response to AZA, therefore indicating a promising new tool in the evaluation of response to demethylating therapies. 28 Moreover, it has been demonstrated that AZA efficacy on increasing PI-PLCb1 gene expression can be accompanied by an inverse correlation to the degree of Akt activation. 27 In this study, we further investigated the role of PI-PLCb1 signaling in epigenetic processes, focusing on the effect of the combination of AZA and VPA on signal transduction molecules.…”