Multi Centers, Open-trial Phase II Study Evaluating 5-azacytidine (Vidaza®) + Valproic Acid (Depakine ®) Before Administration of Retinoic Acid (Vesanoid®) in Patients With Acute Myelogenous Leukemia and High Risk Myelodysplasia.

Chromatin Demethylation Apart from histone acetylation deacetylation, promoter hypermethylation is another important and relevant mechanism involved in gene transcription regulation (reviewed in Herman, 2003). Chromatin remodeling might thus be also targeted using nucleoside analogues, such as 5 azacytidine or decitabine, which reactivate gene transcription through DNA demethylation (Silverman, 2001). Recently, in in VITRO studies, induction of gene expression by 5 AzaC has been obtained in primary AML and MDS cells by DNA methylation dependent and independent mechanisms (SCHMELZ, 2005)

Again, AzaC has been demonstrated as capable to induce clinical hematological responses in patients with MDS. A controlled study conducted by the US Cancer and Leukemia Group B (CALBG)has reported a higher response rate, a lower incidence of leukemic transformation and a prolonged survival as compared with supportive care alone in these patients Silverman, 2002. Another confirmatory Phase 3 study is ongoing.

AzaC, in combination with valproic acid, in leukemic cell line (HL60 and MOLT4, has demonstrated a synergistic activity to induce gene reexpression (reactivation of p21 CIP1) and a synergistic effect in terms of growth inhibition, induction of apoptosis (Yang H, 2005).

Histone Acetylation Numerous investigator groups have tried to elucidate the molecular mechanisms underlying the ATRA induced differentiation in NB4 cells, fresh APL cells, APL mice, or APL patients (Melnick 1999). One of the main issue was to understand the crucial role of the PML RARα fusion protein in the differentiation response to RA. It was observed first that therapeutic concentrations of ATRA resulted in the reformation of PML nuclear bodies associated with a cleavage of the PML RARα fusion protein. Disappearance of this fusion product which acts as a dominant negative regulator of RA target genes transcription gave an explanation for the rerun of the differentiation process. The dominant negative role of PML RARα was secondly explained by the association of the fusion protein to the N-CoR-SMRT-Sin3 corepressor complex, leading to histone deacetylase (HDAC) activities recruitment and to the lack of target genes transcription (REDNER, 1999). Of interest, a similar recruitment of corepressor HDAC activities has been reported in other fusion gene leukemia, including PLZF RARα ,AML1 ETO, CBFß MYH11, and TEL AML1 acute leukemia. In PML RARα APL cells, therapeutic concentrations of ATRA allow the release of corepressor HDAC activities, histone acetylation, chromatin remodeling, and transcription of target genes potentially responsible for terminal granulocytic differentiation (REDNER, 1999; DILWORK, 2001). From this point of view, ATRA therapy of APL is the first example of a gene targeted therapy which specifically targets pathogenic genetic abnormalities in a human leukemia.

In VITRO and in vivo resistance to ATRA-induced differentiation observed in patients with PLZF RARα leukemia has been related to a more potent recruitment of corepressor HDAC activities in this APL subset, as compared to classical PML- RARα APL (two corepressor binding sites on PLZF instead of one on PML). Very interestingly, it has been recently demonstrated that PLZF RARα leukemic cells are not actually completely resistant to differentiation induction, especially if appropriate COSTIMULI are given. First, these cells can differentiate in the presence of higher concentration of ATRA (3 microM instead of 1 microM).