Limited evidence that cancer susceptibility regions are preferential targets for somatic mutation

BackgroundGenome wide-association studies have successfully identified several hundred independent loci harboring common cancer susceptibility alleles that are distinct from the more than 110 cancer predisposition genes. The latter are generally characterized by disruptive mutations in coding genes that have been established as ‘drivers’ of cancer in large somatic sequencing studies. We set out to determine whether, similarly, common cancer susceptibility loci map to genes that have altered frequencies of mutation.ResultsIn our analysis of the intervals defined by the cancer susceptibility markers, we observed that cancer susceptibility regions have gene mutation frequencies comparable to background mutation frequencies. Restricting analyses to genes that have been determined to be pleiotropic across cancer types, genes affected by expression quantitative trait loci, or functional genes indicates that most cancer susceptibility genes classified into these subgroups do not display mutation frequencies that deviate from those expected. We observed limited evidence that cancer susceptibility regions that harbor common alleles with small estimated effect sizes are preferential targets for altered somatic mutation frequencies.ConclusionsOur findings suggest a complex interplay between germline susceptibility and somatic mutation, underscoring the cumulative effect of common variants on redundant pathways as opposed to driver genes. Complex biological pathways and networks likely link these genetic features of carcinogenesis, particularly as they relate to distinct polygenic models for each cancer type.     

ABC Transporters and the Proteasome Complex Are Implicated in Susceptibility to Stevens–Johnson Syndrome and Toxic Epidermal Necrolysis across Multiple Drugs

Stevens–Johnson syndrome (SJS) and Toxic Epidermal Necrolysis (TEN) represent rare but serious adverse drug reactions (ADRs). Both are characterized by distinctive blistering lesions and significant mortality rates. While there is evidence for strong drug-specific genetic predisposition related to HLA alleles, recent genome wide association studies (GWAS) on European and Asian populations have failed to identify genetic susceptibility alleles that are common across multiple drugs. We hypothesize that this is a consequence of the low to moderate effect size of individual genetic risk factors. To test this hypothesis we developed Pointer, a new algorithm that assesses the aggregate effect of multiple low risk variants on a pathway using a gene set enrichment approach. A key advantage of our method is the capability to associate SNPs with genes by exploiting physical proximity as well as by using expression quantitative trait loci (eQTLs) that capture information about both cis- and trans-acting regulatory effects. We control for known bias-inducing aspects of enrichment based analyses, such as: 1) gene length, 2) gene set size, 3) presence of biologically related genes within the same linkage disequilibrium (LD) region, and, 4) genes shared among multiple gene sets. We applied this approach to publicly available SJS/TEN genome-wide genotype data and identified the ABC transporter and Proteasome pathways as potentially implicated in the genetic susceptibility of non-drug-specific SJS/TEN. We demonstrated that the innovative SNP-to-gene mapping phase of the method was essential in detecting the significant enrichment for those pathways. Analysis of an independent gene expression dataset provides supportive functional evidence for the involvement of Proteasome pathways in SJS/TEN cutaneous lesions. These results suggest that Pointer provides a useful framework for the integrative analysis of pharmacogenetic GWAS data, by increasing the power to detect aggregate effects of multiple low risk variants. The software is available for download at https://sourceforge.net/projects/pointergsa/.     

Identification of common predisposing loci to hematopoietic cancers in four dog breeds

Histiocytic sarcoma (HS) is a rare but aggressive cancer in both humans and dogs. The spontaneous canine model, which has clinical, epidemiological, and histological similarities with human HS and specific breed predispositions, provides a unique opportunity to unravel the genetic basis of this cancer. In this study, we aimed to identify germline risk factors associated with the development of HS in canine-predisposed breeds. We used a methodology that combined several genome-wide association studies in a multi-breed and multi-cancer approach as well as targeted next-generation sequencing, and imputation We combined several dog breeds (Bernese mountain dogs, Rottweilers, flat-coated retrievers, and golden retrievers), and three hematopoietic cancers (HS, lymphoma, and mast cell tumor). Results showed that we not only refined the previously identified HS risk CDKN2A locus, but also identified new loci on canine chromosomes 2, 5, 14, and 20. Capture and targeted sequencing of specific loci suggested the existence of regulatory variants in non-coding regions and methylation mechanisms linked to risk haplotypes, which lead to strong cancer predisposition in specific dog breeds. We also showed that these canine cancer predisposing loci appeared to be due to the additive effect of several risk haplotypes involved in other hematopoietic cancers such as lymphoma or mast cell tumors as well. This illustrates the pleiotropic nature of these canine cancer loci as observed in human oncology, thereby reinforcing the interest of predisposed dog breeds to study cancer initiation and progression.     

Mining and validation of pyrosequenced simple sequence repeats (SSRs) from American cranberry (<Emphasis Type="Italic">Vaccinium macrocarpon</Emphasis> Ait.)

The American cranberry (Vaccinium macrocarpon Ait.) is a major commercial fruit crop in North America, but limited genetic resources have been developed for the species. Furthermore, the paucity of codominant DNA markers has hampered the advance of genetic research in cranberry and the Ericaceae family in general. Therefore, we used Roche 454 sequencing technology to perform low-coverage whole genome shotgun sequencing of the cranberry cultivar ‘HyRed’. After de novo assembly, the obtained sequence covered 266.3 Mb of the estimated 540–590 Mb in cranberry genome. A total of 107,244 SSR loci were detected with an overall density across the genome of 403 SSR/Mb. The AG repeat was the most frequent motif in cranberry accounting for 35% of all SSRs and together with AAG and AAAT accounted for 46% of all loci discovered. To validate the SSR loci, we designed 96 primer-pairs using contig sequence data containing perfect SSR repeats, and studied the genetic diversity of 25 cranberry genotypes. We identified 48 polymorphic SSR loci with 2–15 alleles per locus for a total of 323 alleles in the 25 cranberry genotypes. Genetic clustering by principal coordinates and genetic structure analyzes confirmed the heterogeneous nature of cranberries. The parentage composition of several hybrid cultivars was evident from the structure analyzes. Whole genome shotgun 454 sequencing was a cost-effective and efficient way to identify numerous SSR repeats in the cranberry sequence for marker development.     

Genetic Variants Associated with Colorectal Adenoma Susceptibility

BackgroundCommon low-penetrance genetic variants have been consistently associated with colorectal cancer risk.AimTo determine if these genetic variants are associated also with adenoma susceptibility and may improve selection of patients with increased risk for advanced adenomas and/or multiplicity (≥ 3 adenomas).MethodsWe selected 1,326 patients with increased risk for advanced adenomas and/or multiplicity and 1,252 controls with normal colonoscopy from population-based colorectal cancer screening programs. We conducted a case-control association study analyzing 30 colorectal cancer susceptibility variants in order to investigate the contribution of these variants to the development of subsequent advanced neoplasia and/or multiplicity.ResultsWe found that 14 of the analyzed genetic variants showed a statistically significant association with advanced adenomas and/or multiplicity: the probability of developing these lesions increased with the number of risk alleles reaching a 2.3-fold risk increment in individuals with ≥ 17 risk alleles.ConclusionsNearly half of the genetic variants associated with colorectal cancer risk are also related to advanced adenoma and/or multiplicity predisposition. Assessing the number of risk alleles in individuals within colorectal cancer screening programs may help to identify better a subgroup with increased risk for advanced neoplasia and/or multiplicity in the general population.     

Spectrum of HLA associations: the case of medically refractory pediatric acute lymphoblastic leukemia

Although studies of HLA and disease now date back some 50 years, a principled understanding of that relationship has been slow to emerge. Here, we examine the associations of three HLA loci with medically refractory pediatric acute lymphoblastic leukemia (pALL) patients in a case-control study involving 2,438 cases and 41,750 controls. An analysis of alleles from the class I loci, HLA-A and HLA-B, and the class II locus DRB1 illuminates a spectrum of extremely significant allelic associations conferring both predisposition and protection. Genotypes constructed from predisposing, protective, and neutral allelic categories point to an additive mode of disease causation. For all three loci, genotypes homozygous for predisposing alleles are at highest disease risk while the favorable effect of homozygous protective genotypes is less striking. Analysis of A-B and B-DRB1 haplotypes reveals locus-specific differences in disease effects, while that all three loci influence pALL; the influence of HLA-B is greater than that of HLA-A, and the predisposing effect of DRB1 exceeds that of HLA-B. We propose that the continuum in disease susceptibility suggests a system in which many alleles take part in disease predisposition based on differences in binding affinity to one or a few peptides of exogenous origin. This work provides evidence that an immune response mediated by alleles from several HLA loci plays a critical role in the pathogenesis of pALL, adding to the numerous studies pointing to a role for an infectious origin in pALL.     

Development of microsatellite markers for the endangered semi‐shrub Chimaphila umbellata (Ericaceae)

Based on the DNA sequencing reads obtained using 454 pyrosequencing, primers amplifying 16 microsatellite loci were developed for the endangered semi‐shrub Chimaphila umbellata, which occurs sporadically in the Japanese Archipelago. These 16 loci were polymorphic in the populations sampled from the Hokkaido and Tohoku Districts; the mean number of alleles was 3.31 and 3.44, and the mean expected heterozygosity was 0.42 and 0.44, respectively. These loci were not linked to each other and contained no null alleles. Amplification using these primers was also tested in the congeneric species C. japonica, but only three of them successfully amplified DNA of the species. These markers will be used to examine genetic diversity and genetic differentiation in populations of C. umbellata.     

New polymorphic microsatellite markers for bluefin leatherjacket (<Emphasis Type="Italic">Navodon septentrionalis</Emphasis> Gunther, 1877)

A microsatellite-enriched genomic DNA library of Navodon septentrionalis was generated and screened by sequencing. Ten dinucleotide microsatellite loci were characterized by genotyping 24 samples. The observed number of alleles ranged from two to seven with an average of 4.40, while the effective number of alleles ranged from 1.49 to 5.70 with an average of 3.31. The observed and expected heterozygosities ranged from 0.2917 to 0.9167 and from 0.3369 to 0.8422, respectively. No significant linkage disequilibrium between pairs of loci was found, but one loci significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction. These polymorphic microsatellite loci would be useful for investigating genetic population structure and molecule-assisted breeding for N. septentrionalis.     

Genetic control of polyamine-dependent susceptibility to skin tumorigenesis

Overexpression of an ornithine decarboxylase (ODC) transgene greatly increases the susceptibility of mouse skin to carcinogen-induced tumor development. Like many phenotypes in transgenic models, this enhanced susceptibility phenotype is strongly influenced by genetic background. We have mapped tumor-modifier genes in intraspecific crosses between transgenic K6/ODC mice on a susceptible strain background (C57Bl/6J), a moderately resistant background (FVB), or a highly resistant background (C3H/HeJ). We identified several quantitative trait loci that influenced either tumor multiplicity or predisposition to the development of squamous cell carcinoma, but not both phenotypes. Because we did not use a tumor-promotion protocol to induce tumors, most of the quantitative trait loci mapped in this study are distinct from skin tumor-susceptibility loci identified previously. The use of a combined transgenic-standard strain approach to genetic analysis has resulted in detection of previously unknown genetic loci affecting skin tumor susceptibility.     

GWASs and the age of human as the model organism for autoimmune genetic research

Genetic studies have identified more than 150 autoimmune loci, and next-generation sequencing will identify more. Is it time to make human the model organism for autoimmune research?Human genetics - linking inherited variation in DNA sequence with traits such as susceptibility to disease - provides prima facie evidence that a gene and a pathway are associated with a disease. The most recent wave of genomic technology has allowed human genomes to be scanned for variant DNA sequences (or alleles) in many people to determine which alleles are associated with a particular disease or phenotype of interest. Termed genome-wide association studies, or GWASs, this approach has identified hundreds of alleles that are associated with a variety of human traits [1,2]. By most accounts, the GWAS approach has been very successful at identifying new regions of the genome (or loci) that are important in disease, even though the effect sizes of most alleles are modest.The GWAS approach has been particularly successful at uncovering risk alleles for autoimmune diseases. Collectively, autoimmune diseases are common, affecting more than 5% of the adult population [3]. These diseases include rheumatoid arthritis (RA), type 1 diabetes (T1D), inflammatory bowel disease (IBD), systemic lupus erythematosus (SLE), multiple sclerosis (MS), psoriasis and celiac disease (among others). RA is a chronic inflammatory disease that destroys free moving joints. T1D is a form of diabetes that results from the destruction of insulin-producing beta cells of the pancreas. IBD is a group of inflammatory conditions of the colon and small intestine; the two major types are Crohn''s disease and ulcerative colitis. In SLE, the immune system attacks a wide variety of organs, including the heart, joints, skin, lungs, blood vessels, liver, kidneys and nervous system. MS is an autoimmune disease in which the fatty myelin sheaths around the axons of the brain and spinal cord are damaged, leading to a broad spectrum of signs and symptoms. Psoriasis is a chronic disease in which the skin develops red, scaly patches, which is the result of areas of inflammation and excessive skin production. Celiac disease is an autoimmune disorder of the small intestine caused by a reaction to storage proteins (called glutens) found in cereal grains; the ensuing excessive immune reaction leads to an attack on the intestinal villi and tissue damage, resulting in malabsorption of nutrients.So far, approximately 150 loci have been identified that increase risk of these autoimmune diseases [4-14]. For each disease, the strongest genetic risk factors reside within the major histocompatibility complex (MHC) region on chromosome 6 [15]. Most associated alleles in other regions are common in the general population, but increase the disease risk by only 10 to 20% (corresponding to an odds ratio (OR) of 1.10 to 1.20 per copy of the risk allele). (The OR is a measure of the strength of association; it refers to the ratio of the odds of an event occurring in one group (such as cases) to the odds of it occurring in another group (such as controls).) For any given autoimmune disease, the known genetic risk alleles explain between 10 and 20% of variance in disease risk, whereas more than 50% of disease risk is estimated to be heritable. The remaining 30% or so of unexplained genetic disease risk is termed the missing heritability.The challenges now are, first, to find the causal mutation responsible for the signal of association; second, to understand which gene is disrupted by the causal mutation and how it is disrupted (that is, whether the mutation results in gain of function, loss of function, or a new function altogether); third, to understand which cell type and biological pathways are altered by these mutations; and finally to find additional mutations that explain the missing heritability [16]. The next wave of genomic technology - next-generation sequencing - will be a powerful ally in this effort. In particular, next-generation sequencing will help localize the causal mutation, as well as help identify rare alleles that confer risk of autoimmune disease.Thus, an important question remains: what is the most appropriate scientific approach to understand function of risk alleles discovered in human genetics research? Is the mouse the most appropriate model organism, or do these genetic discoveries provide new resources to enable functional studies directly in human immune cells?Here, I discuss the confluence of events that create a unique opportunity to use human subjects as the ''model organism'' for the study of autoimmune disease pathogenesis. In addition to GWASs and next-generation sequencing, registries of blood draws from healthy, consenting human volunteers enable functional studies of genetic variants in a wide range of primary human immune cells, and human stem cell technology has advanced to the point at which induced pluripotent stem (iPS) cells can be derived from patients with specific mutations and differentiated into diverse immune lineages. These resources should allow investigators to understand the altered cellular state in diseases that are uniquely human, which should ultimately lead to new therapeutics to treat or prevent the devastating consequences of autoimmune disease.     

Genetic determinants of plasma triglycerides

Plasma triglyceride (TG) concentration is reemerging as an important cardiovascular disease risk factor. More complete understanding of the genes and variants that modulate plasma TG should enable development of markers for risk prediction, diagnosis, prognosis, and response to therapies and might help specify new directions for therapeutic interventions. Recent genome-wide association studies (GWAS) have identified both known and novel loci associated with plasma TG concentration. However, genetic variation at these loci explains only ～10% of overall TG variation within the population. As the GWAS approach may be reaching its limit for discovering genetic determinants of TG, alternative genetic strategies, such as rare variant sequencing studies and evaluation of animal models, may provide complementary information to flesh out knowledge of clinically and biologically important pathways in TG metabolism. Herein, we review genes recently implicated in TG metabolism and describe how some of these genes likely modulate plasma TG concentration. We also discuss lessons regarding plasma TG metabolism learned from various genomic and genetic experimental approaches. Treatment of patients with moderate to severe hypertriglyceridemia with existing therapies is often challenging; thus, gene products and pathways found in recent genetic research studies provide hope for development of more effective clinical strategies.     

Genetic Analysis of <Emphasis Type="Italic">Porphyra yezoensis</Emphasis> Using Microsatellite Markers

Microsatellites are repetitive genomic elements that show high levels of variation and therefore are useful tools for studying genetic polymorphism and constructing genetic linkage maps of eukaryotic organisms. Porphyra yezoensis Ueda is an economically important seaweed that is being targeted for genetic improvement using marker-assisted breeding. Hence, in an attempt to develop microsatellite markers for P. yezoensis, a microsatellite (or simple sequence repeat)-enriched library was constructed to identify (GA)n and (CA)n motifs. A total of 71 perfect microsatellite clones were identified, of which 30 simple sequence repeat primer pairs were developed. Of these, 24 (80%) amplified polymerase chain reaction products of expected sizes. Twelve primer pairs amplified two to four bands, whereas another 12 primer pairs produced monomorphic banding patterns. Data for 12 loci were analyzed using POPGENE software version 1.32. A total of 29 alleles were produced at 12 loci, with an average of 2.42 alleles (Na) and 1.81 effective alleles (Ne) per locus. These markers were used to analyze the genetic diversity within 11 geographically different lines of P. yezoensis. Overall, these lines were clustered into two divisions with those from close geographic locations clustering together. Further cloning and sequencing of size variant alleles at two microsatellite loci revealed that the variable numbers of motif repeats in different alleles were major sources of polymorphisms.     

Bone tumorigenesis induced by alpha-particle radiation: mapping of genetic loci influencing predisposition in mice

The present study was carried out to determine the extent to which genetic factors modify the incidence of radiation-induced bone tumorigenesis in mice, and to map putative susceptibility genes. We conducted a genome-wide linkage analysis in a cohort of 47 interstrain backcrossed mice. After the mice were injected with the bone-seeking alpha-particle-emitting radionuclide (227)Th, 21 of the mice developed osteosarcomas. Two loci, one on chromosome 7 close to D7Mit145 and a second on chromosome 14 (D14Mit125), exhibited suggestive linkage to osteosarcoma predisposition, with LOD scores of 1.37 and 1.05, respectively. The LOD score increased considerably when interaction between these two loci was taken into account (LOD = 3.48). Nine of 12 mice inheriting a susceptibility allele at both loci developed osteosarcomas after (227)Th injection, compared to only four osteosarcomas in 18 animals that did not inherit either of the susceptibility alleles. Variance component analysis revealed that these genetic factors determine approximately one-fifth of the total incidence of osteosarcomas. This study demonstrates the presence of a genetic component that modulates predisposition to radiation-induced osteosarcoma.     

Polymorphic microsatellite markers for studies of Aedes aegypti (Diptera: Culicidae), the vector of dengue and yellow fever

A significant challenge to population genetic studies of the dengue vector, Aedes aegypti, has been the lack of polymorphic microsatellite loci. In an effort to develop useful markers, we evaluated the genetic variation at 17 microsatellite loci identified in the A. aegypti genome. Nine loci with at least five alleles were identified in field‐collected specimens from Thailand. An additional two loci carried five alleles if samples from an A. aegypti laboratory colony were included. Our results greatly increase the number of highly variable markers available for the study of the genetics and the population structure of this medically important species.     

Genetics of familial melanoma: 20 years after CDKN2A

Twenty years ago, the first familial melanoma susceptibility gene, CDKN2A, was identified. Two years later, another high‐penetrance gene, CDK4, was found to be responsible for melanoma development in some families. Progress in identifying new familial melanoma genes was subsequently slow; however, with the advent of next‐generation sequencing, a small number of new high‐penetrance genes have recently been uncovered. This approach has identified the lineage‐specific oncogene MITF as a susceptibility gene both in melanoma families and in the general population, as well as the discovery of telomere maintenance as a key pathway underlying melanoma predisposition. Given these rapid recent advances, this approach seems likely to continue to pay dividends. Here, we review the currently known familial melanoma genes, providing evidence that most additionally confer risk to other cancers, indicating that they are likely general tumour suppressor genes or oncogenes, which has significant implications for surveillance and screening.     

Genetic predisposition for Type 2 diabetes, but not for overweight/obesity, is associated with a restricted adipogenesis

Background

Development of Type 2 diabetes, like obesity, is promoted by a genetic predisposition. Although several genetic variants have been identified they only account for a small proportion of risk. We have asked if genetic risk is associated with abnormalities in storing excess lipids in the abdominal subcutaneous adipose tissue.

Methodology/Principal Findings

We recruited 164 lean and 500 overweight/obese individuals with or without a genetic predisposition for Type 2 diabetes or obesity. Adipose cell size was measured in biopsies from the abdominal adipose tissue as well as insulin sensitivity (HOMA index), HDL-cholesterol and Apo AI and Apo B. 166 additional non-obese individuals with a genetic predisposition for Type 2 diabetes underwent a euglycemic hyperinsulinemic clamp to measure insulin sensitivity. Genetic predisposition for Type 2 diabetes, but not for overweight/obesity, was associated with inappropriate expansion of the adipose cells, reduced insulin sensitivity and a more proatherogenic lipid profile in non-obese individuals. However, obesity per se induced a similar expansion of adipose cells and dysmetabolic state irrespective of genetic predisposition.

Conclusions/Significance

Genetic predisposition for Type 2 diabetes, but not obesity, is associated with an impaired ability to recruit new adipose cells to store excess lipids in the subcutaneous adipose tissue, thereby promoting ectopic lipid deposition. This becomes particularly evident in non-obese individuals since obesity per se promotes a dysmetabolic state irrespective of genetic predisposition. These results identify a novel susceptibility factor making individuals with a genetic predisposition for Type 2 diabetes particularly sensitive to the environment and caloric excess.