Epidemiologic Approaches to Understanding Complex Diseases: Applications in Congenital Heart Disease and Cancer.

Epidemiology provides a means of investigating the architecture of complex diseases by combining advances in technology with our current understanding of the epidemiology of disease. In this dissertation, epidemiologic methods are applied to further understand the etiology of colorectal cancer, childhood cancers, and congenital heart disease (CHD). The low-penetrance genes that contribute to risk of familial colorectal cancers (CRCs) are mostly unknown. Two leading candidate regions resulting from the Molecular Epidemiology of Colorectal Cancer (MECC) study genome-wide association study included rs10210149 on chromosome 2q11.2-q12 and rs16931815 on chromosome 12p11.23. After excluding potential pathogenic mutations by Sanger sequencing, allele-specific expression analyses were performed for GPR45, STK38L, and TGFBRAP1. GPR45 was associated with a 27% increase in expression for each additional copy of the C allele of rs10210149 (p-trend = 0.01). Common genetic variation and risk of congenital heart disease has not previously been studied. We investigated variation in ISL1, a marker of cells that contribute to specific developmental fields of the embryonic human heart, and risk of CHD in a two-stage case-control study. Eight genic and flanking ISL1 SNPs were significantly associated with complex CHD. A replication study analyzed the three SNPs within ISL1 (rs3762977, IVS1+17C>T, rs1017) and confirmed that genetic variation in ISL1 is associated with risk of CHD. Our results demonstrate that two ISL1 haplotypes contribute to risk of CHD in whites (Summary Odds Ratio (OR) =1.27, 95% Confidence Interval (CI) 1.09 – 1.48, P = 0.0018) and black/African Americans(Summary OR=1.57, 95% CI 1.07 – 2.30, P = 0.0216). Linking epidemiologic approaches to cancer epidemiology and cardiovascular disease is stimulated by the well known association between forms of congenital heart disease and cancer. We conducted a retrospective cohort study of CHD at the Children’s Hospital of Philadelphia (CHOP), and children with CHD demonstrated a 3.7-fold increase in the rate of pediatric cancer compared to the US population (Standardized Incidence Ratio (SIR) = 3.72, 95% CI = 1.53 – 9.04, p = 0.0037). Rates were higher for children with both syndromic (SIR=12.49, 95% CI 1.28 – 121.74, p=0.03) and non-syndromic (SIR=2.41, 95% CI 0.88 – 6.60, p=0.086) heart disease.Ph.D.Epidemiological ScienceUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/78933/1/stkristn_1.pd

Allele-Specific Expression of APC in Adenomatous Polyposis Families

BACKGROUND & AIMS: Germline mutations in the APC gene cause of most cases of familial adenomatous polyposis (FAP) and a lesser proportion of attenuated FAP (AFAP). Systematic analysis of APC at the RNA level could provide insight into the pathogenicity of identified mutations and the molecular basis of FAP/AFAP in families without identifiable mutations. Here, we analyzed the prevalence of imbalances in the allelic expression of APC in polyposis families with germline mutations in the gene and without detectable mutations in APC and/or MUTYH. METHODS: Allele-specific expression (ASE) was determined by single nucleotide primer extension using an exon 11 polymorphism as an allele-specific marker. In total, 52 APC-mutation-positive (36 families) and 24 APC/MUTYH-mutation-negative (23 families) informative patients were analyzed. Seventy-six controls also were included. RESULTS: Of the APC-mutation-positive families, most of those in whom the mutation was located before the last exon of the gene (12 of 14) had ASE imbalance, which is consistent with a mechanism of nonsense-mediated decay. Of the APC/MUTYH-mutation-negative Families, 2 (9%) had ASE imbalance, which might cause the disease. Normal allele expression was restored shortly after lymphocytes were cultured with puromycin, supporting a 'nonsense-mediated' hypothesis. CONCLUSIONS: ASE analysis might be used to determine the pathogenesis of some cases of FAP and AFAP in which APC mutations are not found. ASE also might be used to prioritize the order in which different areas of APC are tested. RNA-level studies are important for the molecular diagnosis of FAP

In vivo characterization of glutamine metabolism identifies therapeutic targets in clear cell renal cell carcinoma

Targeting metabolic vulnerabilities has been proposed as a therapeutic strategy in renal cell carcinoma (RCC). Here, we analyzed the metabolism of patient-derived xenografts (tumorgrafts) from diverse subtypes of RCC. Tumorgrafts from VHL-mutant clear cell RCC (ccRCC) retained metabolic features of human ccRCC and engaged in oxidative and reductive glutamine metabolism. Genetic silencing of isocitrate dehydrogenase-1 or isocitrate dehydrogenase-2 impaired reductive labeling of tricarboxylic acid (TCA) cycle intermediates in vivo and suppressed growth of tumors generated from tumorgraft-derived cells. Glutaminase inhibition reduced the contribution of glutamine to the TCA cycle and resulted in modest suppression of tumorgraft growth. Infusions with [amide-15N]glutamine revealed persistent amidotransferase activity during glutaminase inhibition, and blocking these activities with the amidotransferase inhibitor JHU-083 also reduced tumor growth in both immunocompromised and immunocompetent mice. We conclude that ccRCC tumorgrafts catabolize glutamine via multiple pathways, perhaps explaining why it has been challenging to achieve therapeutic responses in patients by inhibiting glutaminase

Common Variation in ISL1 Confers Genetic Susceptibility for Human Congenital Heart Disease

Congenital heart disease (CHD) is the most common birth abnormality and the etiology is unknown in the overwhelming majority of cases. ISLET1 (ISL1) is a transcription factor that marks cardiac progenitor cells and generates diverse multipotent cardiovascular cell lineages. The fundamental role of ISL1 in cardiac morphogenesis makes this an exceptional candidate gene to consider as a cause of complex congenital heart disease. We evaluated whether genetic variation in ISL1 fits the common variant–common disease hypothesis. A 2-stage case-control study examined 27 polymorphisms mapping to the ISL1 locus in 300 patients with complex congenital heart disease and 2,201 healthy pediatric controls. Eight genic and flanking ISL1 SNPs were significantly associated with complex congenital heart disease. A replication study analyzed these candidate SNPs in 1,044 new cases and 3,934 independent controls and confirmed that genetic variation in ISL1 is associated with risk of non-syndromic congenital heart disease. Our results demonstrate that two different ISL1 haplotypes contribute to risk of CHD in white and black/African American populations

Reduced fire severity offers near-term buffer to climate-driven declines in conifer resilience across the western United States

Increasing fire severity and warmer, drier postfire conditions are making forests in the western United States (West) vulnerable to ecological transformation. Yet, the relative importance of and interactions between these drivers of forest change remain unresolved, particularly over upcoming decades. Here, we assess how the interactive impacts of changing climate and wildfire activity influenced conifer regeneration after 334 wildfires, using a dataset of postfire conifer regeneration from 10,230 field plots. Our findings highlight declining regeneration capacity across the West over the past four decades for the eight dominant conifer species studied. Postfire regeneration is sensitive to high-severity fire, which limits seed availability, and postfire climate, which influences seedling establishment. In the near-term, projected differences in recruitment probability between low- and high-severity fire scenarios were larger than projected climate change impacts for most species, suggesting that reductions in fire severity, and resultant impacts on seed availability, could partially offset expected climate-driven declines in postfire regeneration. Across 40 to 42% of the study area, we project postfire conifer regeneration to be likely following low-severity but not high-severity fire under future climate scenarios (2031 to 2050). However, increasingly warm, dry climate conditions are projected to eventually outweigh the influence of fire severity and seed availability. The percent of the study area considered unlikely to experience conifer regeneration, regardless of fire severity, increased from 5% in 1981 to 2000 to 26 to 31% by mid-century, highlighting a limited time window over which management actions that reduce fire severity may effectively support postfire conifer regeneration. © 2023 the Author(s)

Genome-Wide Association Study in BRCA1 Mutation Carriers Identifies Novel Loci Associated with Breast and Ovarian Cancer Risk

BRCA1-associated breast and ovarian cancer risks can be modified by common genetic variants. To identify further cancer risk-modifying loci, we performed a multi-stage GWAS of 11,705 BRCA1 carriers (of whom 5,920 were diagnosed with breast and 1,839 were diagnosed with ovarian cancer), with a further replication in an additional sample of 2,646 BRCA1 carriers. We identified a novel breast cancer risk modifier locus at 1q32 for BRCA1 carriers (rs2290854, P = 2.7×10-8, HR = 1.14, 95% CI: 1.09-1.20). In addition, we identified two novel ovarian cancer risk modifier loci: 17q21.31 (rs17631303, P = 1.4×10-8, HR = 1.27, 95% CI: 1.17-1.38) and 4q32.3 (rs4691139, P = 3.4×10-8, HR = 1.20, 95% CI: 1.17-1.38). The 4q32.3 locus was not associated with ovarian cancer risk in the general population or BRCA2 carriers, suggesting a BRCA1-specific associat