Mitchell, Braxton D.

Abstract Exonic variants present some of the strongest links between genotype and phenotype. However, these variants can have significant inter-individual pathogenicity differences, known as variable penetrance. In this study, we propose a model where genetically controlled mRNA splicing modulates the pathogenicity of exonic variants. By first cataloging exonic inclusion from RNA-sequencing data in GTEx V8, we find that pathogenic alleles are depleted on highly included exons. Using a large-scale phased whole genome sequencing data from the TOPMed consortium, we observe that this effect may be driven by common splice-regulatory genetic variants, and that natural selection acts on haplotype configurations that reduce the transcript inclusion of putatively pathogenic variants, especially when limiting to haploinsufficient genes. Finally, we test if this effect may be relevant for autism risk using families from the Simons Simplex Collection, but find that splicing of pathogenic alleles has a penetrance reducing effect here as well. Overall, our results indicate that common splice-regulatory variants may play a role in reducing the damaging effects of rare exonic variants.

Abstract With aging, somatic mutations in hematopoietic stem and progenitor cells (HSPC) can give rise to clonal hematopoiesis of indeterminate potential (CHIP), a premalignant state associated with diverse age-related diseases. Here we report the largest multi-ancestry genome-wide analysis of CHIP to date (N = 1,018,305), including individuals of African (N = 85,978), Admixed American (N = 191,371), East Asian (N = 13,532), European (N = 694,015), and South Asian (N = 13,193) ancestry. Multi-ancestry meta-analyses identified 72 genome-wide significant loci, including 44 novel associations implicating genes such as AFF1 , ATF7IP , ATP8B4 , BCL2 , CEBPA , CYRIA , DNM2 , ELF1 , NKX2-3 , PIK3CB , PRDM16 , RPN1 , TERC , and TRIM4 . Notably, variants at MECOM and PHF20L1 showed opposite allelic effects between DNMT3A - and non- DNMT3A -driven CHIP, highlighting driver-specific germline influences. Implicated loci converge on pathways regulating telomere maintenance, cell-cycle control, hematopoietic transcription, DNA damage response and immune signaling. These findings support a model in which germline variation both expands the HSPC pool and biases clonal selection in a driver-dependent manner, providing a mechanistic basis for inter-individual heterogeneity in CHIP. Phenome-wide association analyses further linked CHIP to hematologic, neoplastic, and circulatory traits, with enrichment across hematopoietic and non-hematopoietic cell types. Together, this work expands the genomic and phenomic landscape of CHIP and reveals germline–somatic interactions that shape clonal evolution during aging.

Hypertension is a common bevacizumab‐induced toxicity. No markers are available to predict patients at risk of developing hypertension. We hypothesized that genetic risk of essential hypertension, as measured by a blood pressure polygenic risk score (PRS), would be associated with risk of severe bevacizumab‐induced hypertension. PRSs were calculated for 1,027 bevacizumab‐treated patients of European descent with cancer from four clinical trials (Alliance for Clinical Trials in Oncology (Alliance) / Cancer and Leukemia Group B (CALGB) 80303, 40503, 90401, 40502) using summary systolic blood pressure (SBP) and diastolic blood pressure (DBP) genome‐wide association results obtained from 757,601 individuals of European descent. The association between PRS and grade 3 bevacizumab‐induced hypertension (Common Toxicity Criteria for Adverse Events version 3) in each trial was performed by multivariable logistic regression. Fixed‐effect meta‐analyses odds ratios (ORs) per standard deviation (SD) of the association of PRS (quantitative) and hypertension across trials were estimated by inverse‐variance weighting. PRSs were additionally stratified into quintiles, with the bottom quintile as the referent group. The OR of the association between hypertension and each quintile vs. the referent group was determined by logistic regression. The most significant PRS (quantitative)‐hypertension association included up to 67 single‐nucleotide variants (SNPs) associated with SBP (P = 0.0077, OR per SD = 1.31, 95% confidence interval (CI), 1.07–1.60), and up to 53 SNPs associated with DBP (P = 0.0209, OR per SD = 1.27, 95% CI, 1.04–1.56). Patients in the top quintile had a higher risk of developing bevacizumab‐induced hypertension compared with patients in the bottom quintile using SNPs associated with SBP (P = 4.75 × 10−4, OR = 3.72, 95% CI, 1.84–8.16) and DBP (P = 0.076, OR = 1.83, 95% CI, 0.95–3.64). Genetic variants associated with essential hypertension, mainly SBP, increase the risk of severe bevacizumab‐induced hypertension.

Human genetic studies support an inverse causal relationship between leukocyte telomere length (LTL) and coronary artery disease (CAD), but directionally mixed effects for LTL and diverse malignancies. Clonal hematopoiesis of indeterminate potential (CHIP), characterized by expansion of hematopoietic cells bearing leukemogenic mutations, predisposes both hematologic malignancy and CAD. TERT (which encodes telomerase reverse transcriptase) is the most significantly associated germline locus for CHIP in genome-wide association studies. Here, we investigated the relationship between CHIP, LTL, and CAD in the Trans-Omics for Precision Medicine (TOPMed) program ( n = 63,302) and UK Biobank ( n = 47,080). Bidirectional Mendelian randomization studies were consistent with longer genetically imputed LTL increasing propensity to develop CHIP, but CHIP then, in turn, hastens to shorten measured LTL (mLTL). We also demonstrated evidence of modest mediation between CHIP and CAD by mLTL. Our data promote an understanding of potential causal relationships across CHIP and LTL toward prevention of CAD.

BACKGROUND AND PURPOSE: Ischemic stroke is a heterogeneous complex disease that results from the interaction between numerous genetic and environmental risk factors. Stroke heritability is approximately 40% in the overall population, similar to that of cancer, although there may be important differences in the etiologies of young (<49 years old) versus older onset ischemic stroke. The goal of this study was to conduct a mechanistic genome-wide association (GWA) analysis contrasting the gene pathways involved in young- and old-onset stroke. METHODS: Our analysis included 946 young subjects (448 ischemic stroke subjects and 498 age and sex matched controls, mean age of stroke onset 41 years) and 2558 older subjects (1070 ischemic stroke subjects and 1488 controls, mean age of stroke onset 66.5 years). All subjects were of European ancestry. The young onset cases and controls were genotyped by Illumina Omni1-Quad array with imputation up to 1.8 M SNPs and the old onset cases and controls by a combination of 550 v1 and v3 as well as 610 Illumina arrays with imputation up to 5.0 M SNPs. Single SNP association testing was performed within each contributing study by logistic regression adjusting for age, sex and population structure. Numbers of genes assigned with significant SNPs (P<=0.0001) were 18 and 25 with young- and old-onset groups, respectively, which were used as an input to pathway analysis by the Pathway Studio (Ariadne Genomics, Co.) RESULTS: SNP-level analyses did not reveal any significant associations with each group that met Bonferroni adjusted thresholds for statistical significance. In analyzing the ‘suggestively associated SNPs’, protein-protein interaction pathway analysis in the young-onset group revealed a strong segregation of significant hub proteins (CAT, RUNX1, and EIF4E) with more than 100 connections, while in the old-onset group the interaction networks were dispersed without much segregation. We observed that protein-protein interaction networks have a power-law connectivity distribution in both young- and old-onset groups. We identified 3 significant and focused pathways, ROS metabolism, melanogenesis, and IGF1R->CdEBPA/FOXO1A signaling, enriched for young-onset stroke (P<0.04). In the old-onset group, 10 pathways were identified involving broader signaling pathways. CONCLUSIONS: Young-onset stroke may involve genetic predisposition characterized by a few distinct and finite pathways. Our data suggests that young onset stroke may involve a few pathways that relate to a small number of key hub proteins, including ROS metabolism, melanogenesis, and IGF1R->CEBPA/FOXO1A signaling. Our analysis is consistent with the hypothesis that different pathways may be involved in young versus old onset stroke.

Introduction: Estrogens have been associated with risk of stroke in both men and women. Estradiol and estrone are synthesized from testosterone via the aromatase enzyme in both men and postmenopausal women. Genetic variation affecting function of the aromatase gene (CYP19A1) has been linked to elevated estradiol levels. We tested the hypothesis that these variants, as well as a modifying gene, are associated with ischemic stroke in three cohorts: the Vitamin Intervention for Stroke Prevention trial (VISP), Women’s Health Initiative (WHI), and Genetic Susceptibility to Early-Onset Stroke study (GEOS). Methods: Genomic data for individuals of European descent with ischemic stroke was compared against an equal number of controls: VISP (1723 cases - 65% men), WHI (436 cases - all postmenopausal women), GEOS (557 cases - all men age <55). Genotyping was funded through the Genomics and Randomized Trials Network (GARNET) and Gene Environment Association Studies (GENEVA) consortia. All cohorts used the Illumina Omni1-Quad chip for genotyping with imputation as needed. Eight candidate single nucleotide polymorphisms (SNPs) in CYP19A1 on chromosome 15 and an additional regulatory SNP in a modifier locus on chromosome 8 were chosen for analysis due to coding status or previous association with estradiol levels. Analyses were adjusted for age, hormone therapy use (women), and the top three principal components. Association results were determined as odds ratios for individual SNPs (effect allele = minor), with significance level of p<0.05. Results: GWAS results are tabulated. There were no significant differences in allele frequency for four SNPs in CYP19A1 among stroke patients and controls. The additional four SNPs were monomorphic in all three populations. The modifier gene SNP (rs1864729) was less common among young men with prior stroke in GEOS, but not men in VISP or in women. Conclusions: Aromatase genetic variation was not associated with risk of stroke in these three cohorts. One SNP that modifies aromatase function was less common among young, but not older men with stroke. Replication in other studies is necessary.

Introduction: Genetic studies of early-onset disease have been an effective strategy to identify novel pathways and drug targets generalizable also to later-onset disease. Few studies have investigated the sex-specific genetic associations with early-onset ischemic stroke even though several features of ischemic stroke differ between males and females. We hypothesized that stratifying the GWAS by sex would reveal novel stroke loci. Methods: We performed a transethnic ischemic stroke GWAS of 3,056 female cases and 4,462 male cases < 60 years-old and 16,192 and 16,048 sex-matched controls, respectively, from the Early Onset Stroke Genetics Consortium. Results: We identified a significant association in women with a locus in close proximity to TMX1 , a transmembrane platelet protein that inhibits platelet function. Additionally, we identified 2 other suggestive (P < 5 x 10 -6 ) loci in females (see Table), i.e., at APOH , which encodes beta2-glycoprotein I, an established GWAS locus for lipoprotein(a), and LRFN2 which has been previously reported to associate with obesity-related measures and type II diabetes. We observed suggestive evidence for association in males with MMP3/MMP12 , a known stroke susceptibility locus. Conclusions: Despite a very modest sample size, sex-specific analyses identified suggestive associations at biologically important novel loci in females and a known stroke locus in males. Further studies of sex-specific associations in both early- and later-onset ischemic stroke are needed.

Background and Purpose: Family-based methods for estimating heritability cannot discriminate between shared genetic and shared environmental exposures. Recently, methods have been developed for estimating heritability in population samples using genome-wide SNPs. We used the approach developed by Visscher and colleagues to estimate the heritability of ischemic stroke in Caucasian subjects. In addition to evaluating the overall heritability of ischemic stroke, we assessed whether stroke heritability varies by age, gender, and stroke subtype. Methods: Using publicly available software (GCTA and PLINK), we estimated ischemic stroke heritability stratified by age and gender using genome-wide association (GWA) data from three Caucasian ischemic stroke studies: Ischemic Stroke Genetics Study (ISGS), Bio-Repository of DNA in Stroke (BRAINS), and Genetics of Early-Onset Stroke (GEOS). Weighted means of site-specific heritability point estimates were combined according to a standard fixed effects model. Results: Conclusions: A SNP-based approach may be useful in discerning differences in ischemic stroke heritability between different cohorts and subtypes. Overall, our analysis estimated ischemic stroke heritability to be 31% (SE = 7%), with a suggestion of higher heritability for younger cases. Small vessel stroke showed the highest heritability (58 ± 19%), with cardioembolic showing the lowest heritability (16 ± 14%). It should be emphasized that heritability estimates are population-specific and that the method used only reflects the heritability captured by common SNP variants measured in GWA studies, and not phenotypic variability explained by rare variants.