Esbjorn Paul

Association of ABCB1 polymorphisms with survival and in vitro cytotoxicty in de novo acute myeloid leukemia with normal karyotype

Overexpression of the multi-drug transporter P-glycoprotein, encoded by the ABCB1 gene, is a clinically relevant problem in acute myeloid leukemia (AML). Polymorphisms in ABCB1 might contribute to cancer risk and therapeutic response. We therefore investigated the influence of polymorphisms G1199A, C1236T, G2677T/A and C3435T on cancer susceptibility, in vitro cytotoxicity and overall survival in 100 de novo AML patients with normal karyotype. Patients with 1236C/C or 2677G/G genotypes showed poorer survival than patients with other genotypes (P=0.03 and P=0.02, respectively). Both these genotypes were significant factors for survival in multivariate analysis, along with age, NPM1 and FLT3 mutation status. In vitro cytotoxicity studies demonstrated that leukemic cells from 1236T/T and 2677T/T patients were significantly more susceptible to mitoxantrone (P=0.02), and tended to be more susceptible to etoposide and daunorubicin (P=0.07–0.09), but not to cytarabine. No significant difference in allele frequencies was found between patients and healthy volunteers (n=400).

Decreased survival in normal karyotype AML with single-nucleotide polymorphisms in genes encoding the AraC metabolizing enzymes cytidine deaminase and 5'-nucleotidase.

De novo acute myeloid leukemia with normal karyotype (NK‐AML) comprises a large group of patients with no common cytogenetic alterations and with a large variation in treatment response. Single‐nucleotide polymorphisms (SNPs) in genes related to the metabolism of the nucleoside analogue AraC, the backbone in AML treatment, might affect drug sensitivity and treatment outcome. Therefore, SNPs may serve as prognostic biomarkers aiding clinicians in individualized treatment decisions, with the aim of improving patient outcomes. We analyzed polymorphisms in genes encoding cytidine deaminase (CDA 79A>C rs2072671 and −451C>T rs532545), 5′‐nucleotidase (cN‐II 7A>G rs10883841), and deoxycytidine kinase (DCK 3′UTR 948T>C rs4643786) in 205 de novo NK‐AML patients. In FLT3‐internal tandem duplication (ITD)‐positive patients, the CDA 79C/C and −451T/T genotypes were associated with shorter overall survival compared to other genotypes (5 vs. 24 months, P < 0.001 and 5 vs. 23 months, P = 0.015, respectively), and this was most pronounced in FLT3‐ITD‐positive/NPM1‐positive patients. We observed altered in vitro sensitivity to topoisomerase inhibitory drugs, but not to nucleoside analogues, and a decrease in global DNA methylation in cells carrying both CDA variant alleles. A shorter survival was also observed for the cN‐II variant allele, but only in FLT3‐ITD‐negative patients (25 vs. 31 months, P = 0.075). Our results indicate that polymorphisms in genes related to nucleoside analog drug metabolism may serve as prognostic markers in de novo NK‐AML. Am. J. Hematol. 88:1001–1006, 2013.

Mutations in the isocitrate dehydrogenase 2 gene and IDH1 SNP 105C > T have a prognostic value in acute myeloid leukemia

BackgroundThe isocitrate dehydrogenase (IDH1/IDH2) genes are metabolic enzymes, which are frequently mutated in acute myeloid leukemia (AML). The enzymes acquire neomorphic enzymatic activity when they mutated.MethodsWe have investigated the frequency and outcome of the acquired IDH1/IDH2 mutations and the IDH1 SNP 105C > T (rs11554137) in 189 unselected de novo AML patients by polymerase chain reaction amplification followed by direct sequencing. The survival are presented in Kaplan Meier curves with log rank test. Multivariable survival analysis was conducted using Cox regression method, taking age, risk group, treatment, IDH1/2 mutations and IDH1 SNP105 genotype into account.ResultsOverall, IDH1/2 mutations were found in 41/187 (21.7%) of the AML patients. IDH1 codon 132 mutations were present in 7.9%, whereas IDH2 mutations were more frequent and mutations were identified in codon 140 and 172 in a frequency of 11.1% and 2.6%, respectively. The SNP 105C > T was present in 10.5% of the patients, similar to the normal population. A significantly reduced overall survival (OS) for patients carrying IDH2 codon 140 mutation compared with patients carrying wild-type IDH2 gene (p < 0.001) was observed in the intermediate risk patient group. Neither in the entire patient group nor subdivided in different risk groups, IDH1 mutations had any significance on OS compared to the wild-type IDH1 patients. A significant difference in OS between the heterozygous SNP variant and the homozygous wild-type was observed in the intermediate risk FLT3 negative AML patients (p = 0.004).ConclusionsOur results indicate that AML-patients with IDH2 mutations or the IDH1 SNP 105C > T variant can represent a new subgroup for risk stratification and may indicate new treatment options.

Impact of ABCB1 single nucleotide polymorphisms 1236C>T and 2677G>T on overall survival in FLT3 wild‐type de novo AML patients with normal karyotype

Drug resistance is a clinically relevant problem in the treatment of acute myeloid leukaemia (AML). We have previously reported a relationship between single nucleotide polymorphisms (SNPs) of ABCB1, encoding the multi‐drug transporter P‐glycoprotein, and overall survival (OS) in normal karyotype (NK)‐AML. Here we extended this material, enabling subgroup analysis based on FLT3 and NPM1 status, to further elucidate the influence of ABCB1 SNPs. De novo NK‐AML patients (n = 201) were analysed for 1199G>A, 1236C>T, 2677G>T/A and 3435C>T, and correlations to outcome were investigated. FLT3 wild‐type 1236C/C patients have significantly shorter OS compared to patients carrying the variant allele; medians 20 vs. 49 months, respectively, P = 0·017. There was also an inferior outcome in FLT3 wild‐type 2677G/G patients compared to patients carrying the variant allele, median OS 20 vs. 35 months, respectively, P = 0·039. This was confirmed in Cox regression analysis. Our results indicate that ABCB1 1236C>T and 2677G>T may be used as prognostic markers to distinguish relatively high risk patients in the intermediate risk FLT3 wild‐type group, which may contribute to future individualizing of treatment strategies.

Low p14ARF expression in de novo acute myeloid leukemia with normal karyotype is associated with poor survival

The p14ARF protein activates the p53 tumor suppressor by binding to and inhibiting its negative regulator HDM-2. We have studied the prognostic impact of p14ARF in acute myeloid leukemia (AML). Leukemic cells from 57 adult patients with normal karyotype de novo AML were analyzed for p14ARF mRNA expression level using real-time polymerase chain reaction (RT-PCR). We also tested the effect of conventional anti-leukemic drugs and the mutant p53-targeting small molecule PRIMA-1 in vitro. Patients whose cells expressed more p14ARF mRNA than the 75th percentile (0.26) had significantly better survival compared with those expressing lower levels, 61 vs. 30% 3-year survival (p = 0.046). The difference remained significant also when NPM1/FLT3 status was considered. The mean effects of all the tested conventional anti-leukemic drugs were greater in leukemic cell samples expressing p14ARF mRNA ≥ 0.26, but the differences were not statistically significant. In contrast, PRIMA-1 had a significantly greater effect on leukemic cell samples with low levels of p14ARF mRNA. We conclude that low levels of p14ARF mRNA in leukemic cells from patients with normal karyotype AML is associated with poor prognosis. Treatment with drugs targeting p53 may be a future possibility to improve outcome for these patients.

Impact ofABCB1single nucleotide polymorphisms 1236C>T and 2677G>T on overall survival inFLT3wild-typede novoAML patients with normal karyotype

Drug resistance is a clinically relevant problem in the treatment of acute myeloid leukaemia (AML). We have previously reported a relationship between single nucleotide polymorphisms (SNPs) of ABCB1, encoding the multi‐drug transporter P‐glycoprotein, and overall survival (OS) in normal karyotype (NK)‐AML. Here we extended this material, enabling subgroup analysis based on FLT3 and NPM1 status, to further elucidate the influence of ABCB1 SNPs. De novo NK‐AML patients (n = 201) were analysed for 1199G>A, 1236C>T, 2677G>T/A and 3435C>T, and correlations to outcome were investigated. FLT3 wild‐type 1236C/C patients have significantly shorter OS compared to patients carrying the variant allele; medians 20 vs. 49 months, respectively, P = 0·017. There was also an inferior outcome in FLT3 wild‐type 2677G/G patients compared to patients carrying the variant allele, median OS 20 vs. 35 months, respectively, P = 0·039. This was confirmed in Cox regression analysis. Our results indicate that ABCB1 1236C>T and 2677G>T may be used as prognostic markers to distinguish relatively high risk patients in the intermediate risk FLT3 wild‐type group, which may contribute to future individualizing of treatment strategies.

Impact of ABCB1 single nucleotide polymorphisms 1236C greater than T and 2677G greater than T on overall survival in FLT3 wild-type de novo AML patients with normal karyotype

Drug resistance is a clinically relevant problem in the treatment of acute myeloid leukaemia (AML). We have previously reported a relationship between single nucleotide polymorphisms (SNPs) of ABCB1, encoding the multi‐drug transporter P‐glycoprotein, and overall survival (OS) in normal karyotype (NK)‐AML. Here we extended this material, enabling subgroup analysis based on FLT3 and NPM1 status, to further elucidate the influence of ABCB1 SNPs. De novo NK‐AML patients (n = 201) were analysed for 1199G>A, 1236C>T, 2677G>T/A and 3435C>T, and correlations to outcome were investigated. FLT3 wild‐type 1236C/C patients have significantly shorter OS compared to patients carrying the variant allele; medians 20 vs. 49 months, respectively, P = 0·017. There was also an inferior outcome in FLT3 wild‐type 2677G/G patients compared to patients carrying the variant allele, median OS 20 vs. 35 months, respectively, P = 0·039. This was confirmed in Cox regression analysis. Our results indicate that ABCB1 1236C>T and 2677G>T may be used as prognostic markers to distinguish relatively high risk patients in the intermediate risk FLT3 wild‐type group, which may contribute to future individualizing of treatment strategies.

Correlation between cytidine deaminase single nucleotide polymorphisms and in vitro drug sensitivity, DNA methylation and outcome in normal karyotype acute myelogenous leukemia

Correlation between cytidine deaminase single nucleotide polymorphisms and in vitro drug sensitivity, DNA methylation and outcome in normal karyotype acute myelogenous leukemia

Abstract 2757: NT5C2 single nucleotide polymorphisms affects survival and response in de novo AML patients with normal karyotype

The nucleoside analog cytosine arabinoside (Cytosar®, AraC) is one of the corner stones in the treatment of acute myeloid leukemia (AML). Activated AraC is metabolized to inactive compounds by several intracellular enzymes. The levels of some of these enzymes have been correlated to effect of AraC and clinical outcome in patients with AML. This study was designed to evaluate if single nucleotide polymorphisms (SNPs) in genes coding for enzymes involved in the inactivation of AraC affects the therapeutic response in AML-patients treated with AraC in combination with anthracyclines. Three SNPs were investigated in 96 de novo AML patients with normal karyotype and correlated to response and overall survival. Two SNPs were located in the cytidine deaminase gene (CDA, A79C, rs2072671 and C-451T, rs532545), deaminating and inactivating AraC to its corresponding uridine derivative, AraU. The third SNP (A7G, rs10883841) was in the NT5C2 gene coding for the cytosolic 5′-nucleotidase II (cN-II), responsible for dephosphorylation and hence inactivation of AraC monophosphate. All SNPs were in Hardy-Weinberg equilibrium and the allele frequencies were 34.4, 34.4 and 18.8% for CDA A79C and C-451T, and NT5C2 A7G, respectively. Patients that were wild type (A/A) for NT5C2 A7G were found to have a borderline significant higher response rate (51 complete responders of 60 patients with A/A vs 20 complete responders of 30 with A/G or G/G, p=0.06) and longer survival (median survival 1.66 years compared to 0.87 years, log-rank p=0.08) compared to the rest of the patients. The two CDA SNPs (A79C, C-451T) were found to be in linkage disequilibrium. Although statistically not significant, patients that were found to be homozygous for the alternate genetic variant of the CDA SNP had a shorter survival compared to patients carrying the wild type or heterozygous genetic variants. In conclusion, our findings suggest that certain NT5C2 (cN-II) genotypes might affect the survival after chemotherapy and may provide useful information for treatment strategies, classification and individualized chemotherapy, and eventually used as prognostic marker. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2757.