Exome Sequencing and Linkage Analysis Implicates Two Candidate Genes On Chromosome 3p in Familial Hodgkin Lymphoma

Abstract

53 Background It is well recognised that a genetic component exists in classical Hodgkin lymphoma. Higher rates are seen in first-degree relatives of affected individuals, with high concordance observed in monozygotic compared to dizygotic twins. Numerous associations with HLA alleles have been demonstrated in both Epstein-Barr virus-positive and negative forms of the disease and several non-HLA genes have now been implicated. However, these genetic factors are insufficient to account for all the observed inherited risk. Methods We report a family from North-East Scotland containing five related individuals with classical Hodgkin lymphoma. DNA was extracted from either whole peripheral blood or saliva. Affected individuals were genotyped using the Affymetrix 500Kb SNP array. Genome-wide linkage analysis was performed using easyLINKAGE plus. Targeted exome capture was performed on four affected individuals using the Agilent SureSelect 50Mb kit. Exome sequencing was performed on the Illumina HiSeq2000 to produce 101bp paired-end reads. Following quality control, raw fastq reads were aligned to the human genome (GRCh37) using bwa. SNP/indel calls were performed using Samtools. Variants identified were filtered by quality to include only those with a read coverage of ≥ 20 and phred score of ≥ 30. Given the high penetrance in this family, we hypothesised that a causative genetic variant would not have been described before so eliminated all SNPs present in dbSNP (version 132) or the 1000 genomes project. Finally, only SNPs affecting protein structure were considered further. SIFT and Polyphen-2 were used to predict impact of genetic variation upon protein structure/function. Results Karyotypic analysis was performed on one individual at diagnosis. This was normal, suggesting that a consitutional chromosomal disorder is not responsible for classical Hodgkin lymphoma in this family. The two regions exhibiting strongest linkage with disease were observed on chromosome 3p (LOD scores 1.5 & 1.4). Exome sequencing revealed seven novel, non-synonymous, heterozygous SNPs present in all four analysed individuals. Three were considered biologically plausible (FAM107A:A89S, ALS2CL:L440V, IGSF3:E414G) on the basis of known function and pattern of expression. Two of these three genes (FAM107A and ALS2CL) are on chromosome 3p, in regions similar to those identified in the linkage analysis, and have been implicated as candidate tumor suppressor genes in other malignancies. All three variants were located in evolutionary conserved regions. However, only the mutations in FAM107A and IGSF3 were predicted to disrupt protein function. Discussion Two genes on chromosome 3p represent candidate loci for causing familial classical Hodgkin lymphoma. FAM107A protein is downregulated in several malignant cell lines and primary tumor cells. Overexpression can result in suppression of tumor growth. A study of head and neck carcinoma identified frequent missense mutations and/or loss of heterozygosity at ALS2CL, suggesting that this gene can contribute to tumorigenesis. We are evaluating these genes further by assessing gene expression and protein levels in both Hodgkin cell lines and primary Reed-Sternberg cells. Finally, we intend to evaluate the presence of these candidate genetic variants in other, unaffected members of this family and sporadic cases. Disclosures: No relevant conflicts of interest to declare.

Functional multigenic variations associated with hodgkin lymphoma

“…More recent data also suggest contribution of non‐HLA genes to the inherited susceptibility of familial HL 12,13 . In addition, a recent exome sequencing and linkage analysis in one family identified two genes on 3p ( FAM107A and ALS2CL ) as causative candidates 14 . A strong HLA association for classical Hodgkin lymphoma (HL) risk is well established; however, our understanding of HL heritability has been transformed by recent genome‐wide association studies (GWAS), which have identified single‐nucleotide polymorphisms (SNPs) at seven non‐HLA loci influencing risk 15‐18 .…”

“…12,13 In addition, a recent exome sequencing and linkage analysis in one family identified two genes on 3p (FAM107A and ALS2CL) as causative candidates. 14 A strong HLA association for classical Hodgkin lymphoma (HL) risk is well established; however, our understanding of HL heritability has been transformed by recent genome-wide association studies (GWAS), which have identified single-nucleotide polymorphisms (SNPs) at seven non-HLA loci influencing risk. [15][16][17][18] Although projections indicate that additional risk variants for HL can be discovered by GWAS, 19 the statistical power of published studies is limited.…”