[Show abstract][Hide abstract] ABSTRACT: Background: The response definitions proposed by the European LeukemiaNet (ELN) have been recently modified. We evaluated the new criteria for de novo imatinib (400 mg/d) chronic phase chronic myeloid leukemia (CP-CML) patients.Methods: Response status according to the 2009 and 2013 criteria were determined in 180 unselected patients. Outcome of the subgroups of patients were then compared.Results: The 180 patients were classified as optimal responders (OR2009) (n=113, 62.7%), suboptimal responders (SOR2009) (n=47, 26.1%) and failures (FAIL2009) (n=20, 11.1%) according to the 2009 ELN criteria and optimal responders (OR2013) (n=77, 42.7%), warnings (WAR2013) (n=59, 32.7%) and failures (FAIL2013) (n=44, 24.4%) according to the 2013 ELN criteria. No difference in terms of outcome was observed between OR2009 patients who became WAR2013 when compared to OR2013 patients. When compared to FAIL2009 patients, SOR2009 patients who became WAR2013 had better EFS, FFS, PFS and OS. No difference was observed in PFS or OS in SOR2009 patients who became FAIL2013.Conclusions: The 2013 ELN response status criteria have improved patients classification in terms of response status. However in our patient population this improvement is related to a better definition of failure rather than that of optimal response for CP-CML patients treated with IM frontline therapy.
American Journal of Hematology 10/2014; · 3.48 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Sustained imatinib treatment in chronic myeloid leukaemia patients can result in complete molecular response allowing discontinuation without relapse. We set out to evaluate the frequency of complete molecular response in imatinib de novo chronic-phase chronic myeloid leukaemia patients, identify baseline and under-treatment predictive factors of complete molecular response in patients achieving complete cytogenetic response and assess if complete molecular response is associated with a better outcome. Unselected patients on frontline imatinib therapy (n=266) were considered for inclusion. Complete molecular response was confirmed and defined as MR4.5 with undetectable BCR-ABL transcript levels. Median follow-up was 4-43 years (range, 0.79-10.8 years). Sixty-five patients (24%) achieved complete molecular response within a median time of 32.7 months. Absence of spleen enlargement at diagnosis, achieving complete cytogenetic response before 12 months of therapy and major molecular response during the year following complete cytogenetic response was predictive of achieving further complete molecular response. Patients who achieved complete molecular response had better event-free and failure-free survivals than those with complete cytogenetic response irrespective of major molecular response status (95.2% vs. 64.7% vs. 27.7% p=.00124; 98.4% vs. 82.3% vs. 56%; p=.0335). Overall survival was identical in the 3 groups. In addition to complete cytogenetic response and major molecular response, further deeper molecular response is associated with better event-free and failure-free survivals, and complete molecular response confers the best outcome.
[Show abstract][Hide abstract] ABSTRACT: Myelodysplastic syndromes (MDS) are pre-leukaemic haematopoietic stem cell disorders. Among them, 10-20% occur after chemotherapy and/or radiotherapy, and are called 'therapy-related MDS' (t-MDS). The aim of this study was to identify genetic markers in t-MDS.
A prospective cohort of 59 MDS patients (39 de novo MDS, 20 t-MDS) was studied. A total of 384 single nucleotide polymorphisms (SNP) selected among genes involved in DNA repair, drug metabolism and transport, signal transduction and oncogenesis, were genotyped using a custom-made SNP chip.
Two non-synonymous SNPs present in the methylguanine methyltransferase (MGMT) gene, in complete linkage disequilibrium, were significantly associated with t-MDS: rs2308321 and rs2308327, with a raw p value of 7.4×10(-5) and a corrected p value after Benjamini-Hochberg correction of 0.014. Other associations tested between clinical and cytogenetic features and SNP chip gene variants gave corrected p values above 0.05. A validation cohort was separately constituted of 43 patients (24 de novo MDS, 19 t-MDS) and the two MGMT SNPs were genotyped; it confirmed a significant association between the variant allele of MGMT and t-MDS (p=0.038).
We thus identified a putative marker of the risk to develop MDS after cancer treatment.
European journal of cancer (Oxford, England: 1990) 11/2013; · 4.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Chronic myeloid leukemia disease (CML) found effective therapy by treating patients with tyrosine kinase inhibitors (TKI), which suppress the BCR-ABL1 oncogene activity. However, the majority of patients achieving remission with TKI still have molecular evidences of disease persistence. Various mechanisms have been proposed to explain the disease persistence and recurrence. One of the hypotheses is that the primitive leukemic stem cells (LSCs) can survive in the presence of TKI. Understanding the mechanisms leading to TKI resistance of the LSCs in CML is a critical issue but is limited by availability of cells from patients. We generated induced pluripotent stem cells (iPSCs) derived from CD34(+) blood cells isolated from CML patients (CML-iPSCs) as a model for studying LSCs survival in the presence of TKI and the mechanisms supporting TKI resistance. Interestingly, CML-iPSCs resisted to TKI treatment and their survival did not depend on BCR-ABL1, as for primitive LSCs. Induction of hematopoietic differentiation of CML-iPSC clones was reduced compared to normal clones. Hematopoietic progenitors obtained from iPSCs partially recovered TKI sensitivity. Notably, different CML-iPSCs obtained from the same CML patients were heterogeneous, in terms of BCR-ABL1 level and proliferation. Thus, several clones of CML-iPSCs are a powerful model to decipher all the mechanisms leading to LSC survival following TKI therapy and are a promising tool for testing new therapeutic agents.
PLoS ONE 08/2013; 8(8):e71596. · 3.53 Impact Factor
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[Show abstract][Hide abstract] ABSTRACT: Reliable detection of JAK2-V617F is critical for accurate diagnosis of myeloproliferative neoplasms (MPNs); in addition, sensitive mutation-specific assays can be applied to monitor disease response. However, there has been no consistent approach to JAK2-V617F detection, with assays varying markedly in performance, affecting clinical utility. Therefore, we established a network of 12 laboratories from 7 countries to systematically evaluate 9 different DNA-based quantitative PCR (qPCR) assays, including those in widespread clinical use. Seven quality control rounds involving over 21 500 qPCR reactions were undertaken using centrally distributed cell line dilutions and plasmid controls. The two best-performing assays were tested on normal blood samples (n=100) to evaluate assay specificity, followed by analysis of serial samples from 28 patients transplanted for JAK2-V617F positive disease. The most sensitive assay, which performed consistently across a range of qPCR platforms, predicted outcome following transplant, with the mutant allele detected a median of 22 weeks (range 6-85 weeks) prior to relapse. Four of 7 patients achieved molecular remission following donor lymphocyte infusion, indicative of a graft vs MPN effect. This study has established a robust, reliable assay for sensitive JAK2-V617F detection, suitable for assessing response in clinical trials, predicting outcome and guiding management of patients undergoing allogeneic transplant.Leukemia accepted article preview online, 17 July 2013. doi:10.1038/leu.2013.219.
Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 07/2013; · 10.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The BCR-ABL T315I mutation confers resistance to currently licensed tyrosine kinase inhibitors in chronic myelogenous leukemia. However, the impact on survival at early stages of disease, in chronic phase, has never been detailed. Using matched pair analysis, a cohort of 64 chronic phase patients harboring a T315I mutation and resistant to imatinib mesylate, was compared to a similar cohort of 53 chronic phase patients resistant to imatinib, but with no detectable T315I mutation, in the pre-ponatinib era. These patients were matched according to age at diagnosis, interval between disease diagnosis and imatinib, imatinib duration. Kaplan-Meier survival analyses demonstrate the significant negative impact of the presence of the T315I mutation on overall (since imatinib-resistance T315I+ 48.4 months versus not reached for T315I-, p=0.006) and failure-free survival (since imatinib-resistance: T315I+ 34.7 months versus not reached for T315I-, p=0.003). In addition, Cox proportional hazard models adjusted on overall survival demonstrate the negative influence of the T315I mutation (p=0.02, HR=2.54). These results confirm early assumptions concerning the poor prognosis of chronic phase chronic myelogenous leukemia patients with the T315I mutation who are not eligible for allogeneic transplant, and demonstrate the need for more therapeutic options.
[Show abstract][Hide abstract] ABSTRACT: Chromosomal rearrangements of the human MLL (mixed lineage leukemia) gene are associated with high-risk infant, pediatric, adult and therapy-induced acute leukemias. We used long-distance inverse-polymerase chain reaction to characterize the chromosomal rearrangement of individual acute leukemia patients. We present data of the molecular characterization of 1590 MLL-rearranged biopsy samples obtained from acute leukemia patients. The precise localization of genomic breakpoints within the MLL gene and the involved translocation partner genes (TPGs) were determined and novel TPGs identified. All patients were classified according to their gender (852 females and 745 males), age at diagnosis (558 infant, 416 pediatric and 616 adult leukemia patients) and other clinical criteria. Combined data of our study and recently published data revealed a total of 121 different MLL rearrangements, of which 79 TPGs are now characterized at the molecular level. However, only seven rearrangements seem to be predominantly associated with illegitimate recombinations of the MLL gene (B90%): AFF1/AF4, MLLT3/AF9, MLLT1/ENL,
[Show abstract][Hide abstract] ABSTRACT: Chromosomal translocations involving the immunoglobulin heavy chain locus (IGH@) are recurrent but rare in B-cell precursor acute lymphoblastic leukemia (BCP-ALL), and various partner genes have been described. Here, we report a new series of 29 cases of BCP-ALL with IGH@ translocations. The partner gene was identified by fluorescence in situ hybridization and/or molecular cloning in 20 patients. Members of the CEBP gene family (n = 11), BCL2 (n = 3), ID4 (n = 3), EPOR (n = 2), and TRA/D@ (n = 1) were identified and demonstrated by quantitative real-time reverse transcriptase-polymerase chain reaction to be markedly up-regulated. The present cases, added to those already reported, confirm the diversity of the partner genes, which, apart from BCL2, are specific to BCP-ALL. Collectively, patients with IGH@ translocations may represent a novel sub-group of BCP-ALL occurring in adolescents and young adults.
Cancer Genetics 05/2013; 206(5):162-173. · 2.42 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Refractory anaemia with ring sideroblasts (RARS) and marked thrombocytosis (RARS-T) is a provisional entity in the World Health Organisation 2008 classification and has previously been shown to have a high proportion of JAK2V617F (Janus Kinase 2) and SF3B1 (Splicing Factor 3B subunit 1) mutations. The purpose of the present study was to analyse the frequency of SF3B1 mutations in a large cohort of 111 patients with RARS-T and 33 patients with RARS and to explore the prognostic impact of SF3B1 mutational status on RARS-T. The frequency of SF3B1 mutations in RARS-T (96/111, 86.5%) and RARS (28/33, 84.8%) was similar. In RARS-T, median survival was better in SF3B1-mutated patients than in SF3B1-non-mutated patients (6.9 and 3.3 years, respectively, P=0.003). RARS can be differentiated from RARS-T by the frequency of JAK2V617F (0% vs 48.6%). In RARS-T patients, SF3B1 (P=0.021) and JAK2 mutations (P=0.016) were independent factors for a better prognosis. Altogether, our results confirm that RARS-T is an independent entity that should be recognised by the next World Health Organisation classification. The assessment of SF3B1 mutations is of prognostic interest in RARS-T patients. Younger age, JAK2V617F and SF3B1 mutations are the main predicting factors for survival in RARS-T.
[Show abstract][Hide abstract] ABSTRACT: Identification of chromosomal abnormalities is mandatory for classification of acute myeloid leukemia (AML), and the abnormalities have to be determined quickly, to allow patient enrollment in multicenter protocols and/or for selecting therapeutic strategies. Rapid AML molecular diagnosis is often difficult to achieve, however, because it is based on numerous different RT-PCR protocols. We developed a new RT-PCR method, one that does not require a nested step, to simultaneously detect all AML fusion transcripts from six major recurrent translocations found in adults: t(15;17)(q22;q12), inv(16)(p13.1q22) [t(16;16)(p13.1;q22)], t(8;21)(q22;q22), t(6;9)(p23;q34), t(9;22)(q34;q11), and t(10;11)(p13;q14). Specific primers for RT-PCR detection of the 24 fusion transcripts, along with two transcripts for controls, were designed for this 26-plex RT-PCR. Each PCR product had a different size and was separated by capillary electrophoresis. We also designed a multiplex positive control with 24 chimeric RNAs, corresponding to all chimeric RNAs tested. Compared with classical molecular biology protocols and cytogenetic analyses used as reference standards, results of the 26-plex RT-PCR method were concordant in all 204 (100%) cases of adult AML tested. Results were obtained in less than 24 hours. Because of the multiplex positive control, interpretation of the peaks was very easy, without any ambiguity. The tumor cell detection threshold was 1.5%.
The Journal of molecular diagnostics: JMD 01/2013; · 3.48 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Subclones homozygous for JAK2V617F are more common in polycythemia vera (PV) than essential thrombocythemia (ET), but their prevalence and significance remain unclear. The JAK2 mutation status of 6495 BFU-E, grown in low erythropoietin conditions, was determined in 77 patients with PV or ET. Homozygous-mutant colonies were common in patients with JAK2V617F-positive PV and were surprisingly prevalent in JAK2V617F-positive ET and JAK2 exon 12-mutated PV. Using microsatellite PCR to map loss-of-heterozygosity breakpoints within individual colonies, we demonstrate that recurrent acquisition of JAK2V617F homozygosity occurs frequently in both PV and ET. PV was distinguished from ET by expansion of a dominant homozygous subclone, the selective advantage of which is likely to reflect additional genetic or epigenetic lesions. Our results suggest a model in which development of a dominant JAK2V617F-homzygous subclone drives erythrocytosis in many PV patients, with alternative mechanisms operating in those with small or undetectable homozygous-mutant clones.
[Show abstract][Hide abstract] ABSTRACT: The PICALM-MLLT10 fusion gene, generated by the t(10;11)(p12-13;q14-21) translocation, is a rare but recurrent event in acute leukemias. In this study, we assessed the characteristics and outcome of 18 PICALM-MLLT10 AML patients. As compared with non PICALM-MLLT10 patients (n=72), PICALM-MLLT10 AML were characterized by more frequent extramedullary diseases, CD7 expression and higher platelet counts. Three out of four therapy-related PICALM-MLLT10 AMLs had been previously treated for diffuse large B-cell lymphoma. The complete response rate was 71% after intensive chemotherapy. PICALM-MLLT10 patients had a shorter median overall survival than patients with favorable cytogenetics (12months vs. not reached, p=0.07) but not significantly different from those of intermediate (26months, p=0.32) or unfavorable cytogenetic groups (8months, p=0.13). Long term responses were achieved in a subset of patients after allogeneic stem-cell transplantation but also after high-dose cytarabine.
Leukemia research 08/2012; 36(11):1365-9. · 2.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Congenital erythropoietic porphyria (CEP) is due to a deficiency in the enzymatic activity of uroporphyrinogen III synthase (UROS); such a deficiency leads to porphyrin accumulation and results in skin lesions and hemolytic anemia. CEP is a candidate for retrolentivirus-mediated gene therapy, but recent reports of insertional leukemogenesis underscore the need for safer methods. The discovery of induced pluripotent stem cells (iPSCs) has opened up new horizons in gene therapy because it might overcome the difficulty of obtaining sufficient amounts of autologous hematopoietic stem cells for transplantation and the risk of genotoxicity. In this study, we isolated keratinocytes from a CEP-affected individual and generated iPSCs with two excisable lentiviral vectors. Gene correction of CEP-derived iPSCs was obtained by lentiviral transduction of a therapeutic vector containing UROS cDNA under the control of an erythroid-specific promoter shielded by insulators. One iPSC clone, free of reprogramming genes, was obtained with a single proviral integration of the therapeutic vector in a genomic safe region. Metabolic correction of erythroblasts derived from iPSC clones was demonstrated by the disappearance of fluorocytes. This study reports the feasibility of porphyria gene therapy with the use of iPSCs.
The American Journal of Human Genetics 07/2012; 91(1):109-21. · 11.20 Impact Factor