Comparative analysis of IL6 and IL6 receptor gene polymorphisms in mastocytosis

Mastocytosis is a rare disease with reported high interleukin-6 (IL6) levels influencing disease severity. The present study investigated polymorphisms within the genes that encode IL6 and its receptor (IL6R) in relation to mastocytosis development in a case-control design. Analysis of the IL6R Asp358Ala polymorphism showed that carriers of the AA genotype had a 2.5-fold lower risk for mastocytosis than those with the AC or CC genotypes. No association with mastocytosis was found for the IL6-174G/C polymorphism, however, it may influence the effect of IL6R polymorphism. To the best of our knowledge this is the first study analysing IL6/IL6R polymorphisms in mastocytosis

Roles of genetic polymorphisms in the folate pathway in childhood acute lymphoblastic leukemia evaluated by bayesian relevance and effect size analysis.

In this study we investigated whether polymorphisms in the folate pathway influenced the risk of childhood acute lymphoblastic leukemia (ALL) or the survival rate of the patients. For this we selected and genotyped 67 SNPs in 15 genes in the folate pathway in 543 children with ALL and 529 controls. The results were evaluated by gender adjusted logistic regression and by the Bayesian network based Bayesian multilevel analysis of relevance (BN-BMLA) methods. Bayesian structure based odds ratios for the relevant variables and interactions were also calculated. Altogether 9 SNPs in 8 genes were associated with altered susceptibility to ALL. After correction for multiple testing, two associations remained significant. The genotype distribution of the MTHFD1 rs1076991 differed significantly between the ALL and control population. Analyzing the subtypes of the disease the GG genotype increased only the risk of B-cell ALL (p = 3.52x10(-4); OR = 2.00). The GG genotype of the rs3776455 SNP in the MTRR gene was associated with a significantly reduced risk to ALL (p = 1.21x10(-3); OR = 0.55), which resulted mainly from the reduced risk to B-cell and hyperdiploid-ALL. The TC genotype of the rs9909104 SNP in the SHMT1 gene was associated with a lower survival rate comparing it to the TT genotype (80.2% vs. 88.8%; p = 0.01). The BN-BMLA confirmed the main findings of the frequentist-based analysis and showed structural interactional maps and the probabilities of the different structural association types of the relevant SNPs especially in the hyperdiploid-ALL, involving additional SNPs in genes like TYMS, DHFR and GGH. We also investigated the statistical interactions and redundancies using structural model properties. These results gave further evidence that polymorphisms in the folate pathway could influence the ALL risk and the effectiveness of the therapy. It was also shown that in gene association studies the BN-BMLA could be a useful supplementary to the traditional frequentist-based statistical method

Candidate gene association study in pediatric acute lymphoblastic leukemia evaluated by Bayesian network based Bayesian multilevel analysis of relevance

Background: We carried out a candidate gene association study in pediatric acute lymphoblastic leukemia (ALL) to identify possible genetic risk factors in a Hungarian population. Methods: The results were evaluated with traditional statistical methods and with our newly developed Bayesian network based Bayesian multilevel analysis of relevance (BN-BMLA) method. We collected genomic DNA and clinical data from 543 children, who underwent chemotherapy due to ALL, and 529 healthy controls. Altogether 66 single nucleotide polymorphisms (SNPs) in 19 candidate genes were genotyped. Results: With logistic regression, we identified 6 SNPs in the ARID5B and IKZF1 genes associated with increased risk to B-cell ALL, and two SNPs in the STAT3 gene, which decreased the risk to hyperdiploid ALL. Because the associated SNPs were in linkage in each gene, these associations corresponded to one signal per gene. The odds ratio (OR) associated with the tag SNPs were: OR = 1.69, P = 2.22x10-7 for rs4132601 (IKZF1), OR = 1.53, P = 1.95x10-5 for rs10821936 (ARID5B) and OR = 0.64, P = 2.32x10-4 for rs12949918 (STAT3). With the BN-BMLA we confirmed the findings of the frequentist-based method and received additional information about the nature of the relations between the SNPs and the disease. E.g. the rs10821936 in ARID5B and rs17405722 in STAT3 showed a weak interaction, and in case of T-cell lineage sample group, the gender showed a weak interaction with three SNPs in three genes. In the hyperdiploid patient group the BN-BMLA detected a strong interaction among SNPs in the NOTCH1, STAT1, STAT3 and BCL2 genes. Evaluating the survival rate of the patients with ALL, the BN-BMLA showed that besides risk groups and subtypes, genetic variations in the BAX and CEBPA genes might also influence the probability of survival of the patients. Conclusions: In the present study we confirmed the roles of genetic variations in ARID5B and IKZF1 in the susceptibility to B-cell ALL. With the newly developed BN-BMLA method several gene-gene, gene-phenotype and phenotype-phenotype connections were revealed. We showed several advantageous features of the new method, and suggested that in gene association studies the BN-BMLA might be a useful supplementary to the traditional frequentist-based statistical method

Subgroups of Paediatric Acute Lymphoblastic Leukaemia Might Differ Significantly in Genetic Predisposition to Asparaginase Hypersensitivity.

L-asparaginase (ASP) is a key element in the treatment of paediatric acute lymphoblastic leukaemia (ALL). However, hypersensitivity reactions (HSRs) to ASP are major challenges in paediatric patients. Our aim was to investigate genetic variants that may influence the risk to Escherichia coli-derived ASP hypersensitivity. Sample and clinical data collection was carried out from 576 paediatric ALL patients who were treated according to protocols from the Berlin-Frankfurt-Munster Study Group. A total of 20 single nucleotide polymorphisms (SNPs) in GRIA1 and GALNT10 genes were genotyped. Patients with GRIA1 rs4958351 AA/AG genotype showed significantly reduced risk to ASP hypersensitivity compared to patients with GG genotype in the T-cell ALL subgroup (OR = 0.05 (0.01-0.26); p = 4.70E-04), while no such association was found in pre-B-cell ALL. In the medium risk group two SNPs of GRIA1 (rs2055083 and rs707176) were associated significantly with the occurrence of ASP hypersensitivity (OR = 0.21 (0.09-0.53); p = 8.48E-04 and OR = 3.02 (1.36-6.73); p = 6.76E-03, respectively). Evaluating the genders separately, however, the association of rs707176 with ASP HSRs was confined only to females. Our results suggest that genetic variants of GRIA1 might influence the risk to ASP hypersensitivity, but subgroups of patients can differ significantly in this respect

Genome-Wide DNA Methylation Analysis Reveals Epigenetic Deregulation of Key Biological Pathways in Therapy-Related Myeloid Neoplasms

Abstract Therapy-related myeloid neoplasms (tMN), including therapy-related myelodysplastic syndromes (tMDS) and therapy-related acute myeloid leukemia (tAML) are associated with resistance to chemotherapy and a more unfavorable prognosis than their de novo counterparts. While many studies have explored the epigenome of de novo MDS, little is known about the epigenetic status of tMN. Recently, we reported that tMN showed distinct mutational profiles whether they arose after an initial solid tumor or a lymphoproliferative disorder, indicating that aside from the type of therapy received it is possible the type of primary neoplasm might also help determine the phenotype of t-MN. Therefore, we sought to perform a comprehensive epigenomic study of tMN that would help improve our current understanding of epigenetic deregulation in this disorder. Using next-generation bisulfite sequencing we analyzed the DNA methylation status at ~3M CpG sites of 15 cases, including 5 de novo MDS (dnMDS), 5 tMN with a prior history of Hodgkin lymphoma (HL-tMN) treated with MOPP/BEACOPP-based regimens, and 5 tMN arising after breast cancer (BC-tMN) treated with combined chemo-radiotherapy. There were no significant differences in median age, median bone marrow blast percentage, or median latency to t-MN for the HL and BC groups. Karyotype mainly consisted of abnormalities of chromosomes 5 or 7, or complex karyotypes in all groups. None of the patients had balanced translocations or abnormalities of the 11q23 locus. We first sought to determine whether tMDS and dnMDS are epigenetically distinct. For this we compared the DNA methylation profiles of the tMDS (n=6) vs. dnMDS (n=5). We identified 1504 statistically significant (FDR<0.1 and mean methylation difference ≥25%) differentially methylated regions (DMRs). Relative to dnMDS, tMDS showed a markedly hypomethylated profile. Pathway analysis using DAVID revealed that these DMRs were significantly enriched for genes in the Wnt signaling (FDR=3.8x10-6) and cadherin pathways (FDR=5.3x10-7), indicating that these key pathways maybe play different roles in these two forms of MDS. Next we asked whether among tMNs the type of neoplasm preceding the development of tMN might also influence the epigenome. To address this we performed a direct comparison of the DNA methylation profiles for the HL-tMN (n=5) vs. BC-tMN (n=5), which identified 457 statistically significant DMRs, the vast majority of which localized to intronic and intergenic regions and were hypomethylated in HL-tMN relative to BC-tMN. Finally, in order to determine the role that epigenetic deregulation may play in different forms of AMLs not arising de novo, we compared the epigenetic profiles of tAML (n=4) to a cohort of AMLs arising after progression from a myeloproliferative neoplasm (MPN) (n=5). Notably, despite their shared features of chemoresistance and poor prognosis, the epigenetic profiles of these two entities were vastly different, with 14,887 statistically significant DMRs detected. These DMRs were depleted at promoter regions (DMRs 4% vs. Background [BG] 12%, Fisher p-value[p]=0.038) and CpG islands (DMRs 9% vs. BG 30%, p=0.0003), but were enriched at regions outside of CpG islands (DMRs 73% vs. BG 54%, p=0.008). Gene ontology analysis showed these DMRs were significantly enriched for genes involved transcriptional regulation (FDR=0.02), nucleotide binding (FDR=0.00025), regulation of RNA metabolic process (FDR=0.02), and protein kinase activity (FDR=0.009). In addition, these DMRs were significantly enriched at enhancer regions (p<0.0001), indicating they may play key regulatory roles. In summary, we have completed the first comprehensive analysis of the epigenome-wide abnormalities associated with tMN. Our findings demonstrate that tMN are epigenetically distinct from dnMDS and that these abnormalities target biological pathways known to play a key role in self-renewal and differentiation of hematopoietic stem and cancer cells, as well in MDS pathogenesis. Moreover, we demonstrate that similar to our findings at the mutational level, epigenetic deregulation in tMN also has distinct profiles strongly correlated with the type of first malignancy preceding the tMN. Finally, AML arising as a progression of MPN or after prior chemotherapy show robust epigenetic differences that clearly distinguish these two entities. Disclosures Voso: Celgene: Consultancy

Associations of novel genetic variations in the folate-related and ARID5B genes with the pharmacokinetics and toxicity of high-dose methotrexate in paediatric acute lymphoblastic leukaemia

High-dose methotrexate (HD-MTX) plays an important role in the consolidation therapy of acute lymphoblastic leukaemia (ALL) in many treatment regimens worldwide. However, there is a large interpatient variability in the pharmacokinetics and toxicity of the drug. We investigated the influence of single nucleotide polymorphisms (SNPs) in genes of the folate metabolic pathway, transporter molecules and transcription proteins on the pharmacokinetics and toxicity of MTX and 7-hydroxy-methotrexate (7-OH-MTX). 63 SNPs of 14 genes were genotyped and a total of 463 HD-MTX courses (administered according to the ALL-BFM 95 and ALL IC-BFM 2002 protocols) were analysed. Haematological, hepatic and renal toxicities, estimated by routine laboratory parameters were evaluated. Random forest and regression trees were used for variable selection and model building. Linear mixed models were established to prove the significance of the selected variables. SNPs (rs4948502, rs4948496, rs4948487) of the ARID5B gene were associated with the serum levels of MTX (P < 0.02), serum levels and area under the curve of 7-OH-MTX (P < 0.02) and with hypoproteinaemia (P = 0.004). SLCO1B1 rs4149056 also showed a significant association with serum MTX levels (P < 0.001). Our findings confirm the association of novel genetic variations in folate-related and ARID5B genes with the serum MTX levels and acute toxicity

In interaction with gender a common CYP3A4 polymorphism may influence the survival rate of chemotherapy for childhood acute lymphoblastic leukemia

CYP3A4 has an important role in the metabolisms of many drugs used in acute lymphoblastic leukemia (ALL) therapy; still, there are practically no publications about the role of CYP3A4 polymorphisms in ALL pharmacogenomics. We genotyped eight common single-nucleotide polymorphisms (SNPs) in the CYP3A4 and CYP3A5 genes in 511 children with ALL and investigated whether they influenced the survival of the patients. We involved additional 127 SNPs in 34 candidate genes and searched for interactions with respect to the survival rates. Significant association between the survival rates and the common rs2246709 SNP in the CYP3A4 gene was observed. The gender of the patients and the rs1076991 in the MTHFD1 gene strongly influenced this effect. We calculated new risk assessments involving the gender-rs2246709 interaction and showed that they significantly outperformed the earlier risk-group assessments at every time point. If this finding is confirmed in other populations, it can have a considerable prognostic significance.The Pharmacogenomics Journal advance online publication, 30 September 2014; doi:10.1038/tpj.2014.60

Illustration of different dependency relations between relevant SNPs and hyperdiploid ALL susceptibility.

<p>Top panel: An interactional map including hyperdiploid ALL susceptibility (red node) and strongly relevant SNPs with respect to it (other nodes). The width of the edges is proportional to their a posteriori probability. The directed edges represent only probabilistic relationships between the variables which are not necessary causal. Bottom panel: The posterior probability of strong relevance (blue columns), edge (direct strong relevance, red columns), pure interaction (green columns) and association (purple columns) of the variables to hyperdiploid ALL susceptibility according to the BN-BMLA method.</p

List of polymorphisms investigated in this study.

1<p>Position according to NCBI Genome Build 36.0;</p>2<p>Function according to the UCSC Genome Browser <a href="http://genome.ucsc.edu/(Accessed" target="_blank">http://genome.ucsc.edu/(Accessed</a>: 2013 Jun 20.);</p>3<p>RFC = SLC19A1;</p>4<p>SLCO1B1 = SLC21A6.</p