Genetic Variants of VEGFA and FLT4 Are Determinants of Survival in Renal Cell Carcinoma Patients Treated with Sorafenib

Molecular markers of sorafenib efficacy in patients with metastatic renal cell carcinoma (mRCC) are not available. The purpose of this study was to discover genetic markers of survival in patients with mRCC treated with sorafenib. Germline variants from 56 genes were genotyped in 295 patients with mRCC. Variant-overall survival (OS) associations were tested in multivariate regression models. Mechanistic studies were conducted to validate clinical associations. VEGFA rs1885657, ITGAV rs3816375, and WWOX rs8047917 (sorafenib arm), and FLT4 rs307826 and VEGFA rs3024987 (sorafenib and placebo arms combined) were associated with shorter OS. FLT4 rs307826 increased VEGFR-3 phosphorylation, membrane trafficking, and receptor activation. VEGFA rs1885657 and rs58159269 increased transcriptional activity of the constructs containing these variants in endothelial and RCC cell lines, and VEGFA rs58159269 increased endothelial cell proliferation and tube formation. FLT4 rs307826 and VEGFA rs58159269 led to reduced sorafenib cytotoxicity. Genetic variation in VEGFA and FLT4 could affect survival in sorafenib-treated patients with mRCC. These markers should be examined in additional malignancies treated with sorafenib and in other angiogenesis inhibitors used in mRCC. Significance: Clinical and mechanistic data identify germline genetic variants in VEGFA and FLT4 as markers of survival in patients with metastatic renal cell carcinoma.Peer reviewe

The vitamin D receptor gene as a determinant of survival in pancreatic cancer patients: Genomic analysis and experimental validation

Purpose: Advanced pancreatic cancer is a highly refractory disease almost always associated with survival of little more than a year. New interventions based on novel targets are needed. We aim to identify new genetic determinants of overall survival (OS) in patients after treatment with gemcitabine using genome-wide screens of germline DNA. We aim also to support these findings with in vitro functional analysis. Patients and methods: Genome-wide screens of germline DNA in two independent cohorts of pancreatic cancer patients (from the Cancer and Leukemia Group B (CALGB) 80303 and the Mayo Clinic) were used to select new genes associated with OS. The vitamin D receptor gene (VDR) was selected, and the interactions of genetic variation in VDR with circulating vitamin D levels and gemcitabine treatment were evaluated. Functional effects of common VDR variants were also evaluated in experimental assays in human cell lines. Results: The rs2853564 variant in VDR was associated with OS in patients from both the Mayo Clinic (HR 0.81, 95% CI 0.70–0.94, p = 0.0059) and CALGB 80303 (HR 0.74, 0.63–0.87, p = 0.0002). rs2853564 interacted with high pre-treatment levels of 25-hydroxyvitamin D (25(OH)D, a measure of endogenous vitamin D) (p = 0.0079 for interaction) and with gemcitabine treatment (p = 0.024 for interaction) to confer increased OS. rs2853564 increased transcriptional activity in luciferase assays and reduced the binding of the IRF4 transcription factor. Conclusion: Our findings propose VDR as a novel determinant of survival in advanced pancreatic cancer patients. Common functional variation in this gene might interact with endogenous vitamin D and gemcitabine treatment to determine improved patient survival. These results support evidence for a modulatory role of the vitamin D pathway for the survival of advanced pancreatic cancer patients.</p

Discovery and Functional Assessment of Gene Variants in the Vascular Endothelial Growth Factor Pathway

Angiogenesis is a host-mediated mechanism in disease pathophysiology. The vascular endothelial growth factor (VEGF) pathway is a major determinant of angiogenesis, and a comprehensive annotation of the functional variation in this pathway is essential to understand the genetic basis of angiogenesis-related diseases. We assessed the allelic heterogeneity of gene expression, population specificity of cis expression quantitative trait loci (eQTLs), and eQTL function in luciferase assays in CEU and YRI HapMap lymphoblastoid cell lines (LCLs) in 23 resequenced genes. Among 356 cis-eQTLs, 155 and 174 were unique to CEU and YRI, respectively, and 27 were shared between CEU and YRI. Two cis-eQTLs provided mechanistic evidence for two GWAS findings. Five eQTLs were tested for function in luciferase assays and the effect of two KRAS variants was concordant with the eQTL effect. Two eQTLs found in each of PRKCE, PIK3C2A, and MAP2K6, could predict 44, 37 and 45% of the variance in gene expression, respectively. This is the first analysis focusing on the pattern of functional genetic variation of the VEGF pathway genes in CEU and YRI populations and providing mechanistic evidence for genetic association studies of diseases for which angiogenesis plays a pathophysiologic role

A Common Allele in FGF21 Associated with Sugar Intake Is Associated with Body Shape, Lower Total Body-Fat Percentage, and Higher Blood Pressure

Summary: Fibroblast growth factor 21 (FGF21) is a hormone that has insulin-sensitizing properties. Some trials of FGF21 analogs show weight loss and lipid-lowering effects. Recent studies have shown that a common allele in the FGF21 gene alters the balance of macronutrients consumed, but there was little evidence of an effect on metabolic traits. We studied a common FGF21 allele (A:rs838133) in 451,099 people from the UK Biobank study, aiming to use the human allele to inform potential adverse and beneficial effects of targeting FGF21. We replicated the association between the A allele and higher percentage carbohydrate intake. We then showed that this allele is more strongly associated with higher blood pressure and waist-hip ratio, despite an association with lower total body-fat percentage, than it is with BMI or type 2 diabetes. These human phenotypes of variation in the FGF21 gene will inform research into FGF21’s mechanisms and therapeutic potential. : Drugs targeting the hormone FGF21 may have beneficial health effects. Variations in human DNA in the FGF21 gene provide an indication of what those effects may be. Here, we show that variation in the FGF21 gene is associated with higher blood pressure and altered body shape, despite lower total body-fat percentage. Keywords: FGF21, BMI, waist-hip ratio, blood pressure, body fat, allele, genetic variant, UK Bioban

Identification and single-base gene-editing functional validation of a cis-EPO variant as a genetic predictor for EPO-increasing therapies

Hypoxia-inducible factor prolyl hydroxylase inhibitors (HIF-PHIs) are currently under clinical development for treating anemia in chronic kidney disease (CKD), but it is important to monitor their cardiovascular safety. Genetic variants can be used as predictors to help inform the potential risk of adverse effects associated with drug treatments. We therefore aimed to use human genetics to help assess the risk of adverse cardiovascular events associated with therapeutically altered EPO levels to help inform clinical trials studying the safety of HIF-PHIs. By performing a genome-wide association meta-analysis of EPO (n = 6,127), we identified a cis-EPO variant (rs1617640) lying in the EPO promoter region. We validated this variant as most likely causal in controlling EPO levels by using genetic and functional approaches, including single-base gene editing. Using this variant as a partial predictor for therapeutic modulation of EPO and large genome-wide association data in Mendelian randomization tests, we found no evidence (at p < 0.05) that genetically predicted long-term rises in endogenous EPO, equivalent to a 2.2-unit increase, increased risk of coronary artery disease (CAD, OR [95% CI] = 1.01 [0.93, 1.07]), myocardial infarction (MI, OR [95% CI] = 0.99 [0.87, 1.15]), or stroke (OR [95% CI] = 0.97 [0.87, 1.07]). We could exclude increased odds of 1.15 for cardiovascular disease for a 2.2-unit EPO increase. A combination of genetic and functional studies provides a powerful approach to investigate the potential therapeutic profile of EPO-increasing therapies for treating anemia in CKD

Genome-wide meta-analysis identifies genetic variants associated with glycemic response to sulfonylureas

OBJECTIVE: Sulfonylureas, the first available drugs for the management of type 2 diabetes, remain widely prescribed today. However, there exists significant variability in glycemic response to treatment. We aimed to establish heritability of sulfonylurea response and identify genetic variants and interacting treatments associated with HbA(1c) reduction. RESEARCH DESIGN AND METHODS: As an initiative of the Metformin Genetics Plus Consortium (MetGen Plus) and the DIabetes REsearCh on patient straTification (DIRECT) consortium, 5,485 White Europeans with type 2 diabetes treated with sulfonylureas were recruited from six referral centers in Europe and North America. We first estimated heritability using the generalized restricted maximum likelihood approach and then undertook genome-wide association studies of glycemic response to sulfonylureas measured as HbA(1c) reduction after 12 months of therapy followed by meta-analysis. These results were supported by acute glipizide challenge in humans who were naïve to type 2 diabetes medications, cis expression quantitative trait loci (eQTL), and functional validation in cellular models. Finally, we examined for possible drug-drug-gene interactions. RESULTS: After establishing that sulfonylurea response is heritable (mean ± SEM 37 ± 11%), we identified two independent loci near the GXYLT1 and SLCO1B1 genes associated with HbA(1c) reduction at a genome-wide scale (P < 5 × 10(−8)). The C allele at rs1234032, near GXYLT1, was associated with 0.14% (1.5 mmol/mol), P = 2.39 × 10(−8)), lower reduction in HbA(1c). Similarly, the C allele was associated with higher glucose trough levels (β = 1.61, P = 0.005) in healthy volunteers in the SUGAR-MGH given glipizide (N = 857). In 3,029 human whole blood samples, the C allele is a cis eQTL for increased expression of GXYLT1 (β = 0.21, P = 2.04 × 10(−58)). The C allele of rs10770791, in an intronic region of SLCO1B1, was associated with 0.11% (1.2 mmol/mol) greater reduction in HbA(1c) (P = 4.80 × 10(−8)). In 1,183 human liver samples, the C allele at rs10770791 is a cis eQTL for reduced SLCO1B1 expression (P = 1.61 × 10(−7)), which, together with functional studies in cells expressing SLCO1B1, supports a key role for hepatic SLCO1B1 (encoding OATP1B1) in regulation of sulfonylurea transport. Further, a significant interaction between statin use and SLCO1B1 genotype was observed (P = 0.001). In statin nonusers, C allele homozygotes at rs10770791 had a large absolute reduction in HbA(1c) (0.48 ± 0.12% [5.2 ± 1.26 mmol/mol]), equivalent to that associated with initiation of a dipeptidyl peptidase 4 inhibitor. CONCLUSIONS: We have identified clinically important genetic effects at genome-wide levels of significance, and important drug-drug-gene interactions, which include commonly prescribed statins. With increasing availability of genetic data embedded in clinical records these findings will be important in prescribing glucose-lowering drugs

Integration of DNA sequencing with population pharmacokinetics to improve the prediction of irinotecan exposure in cancer patients

Background: Irinotecan (CPT-11) is an anticancer agent widely used to treat adult solid tumours. Large interindividual variability in the clearance of irinotecan and SN-38, its active and toxic metabolite, results in highly unpredictable toxicity. Methods: In 217 cancer patients treated with intravenous irinotecan single agent or in combination, germline DNA was used to interrogate the variation in 84 genes by next-generation sequencing. A stepwise analytical framework including a population pharmacokinetic model with SNP- and gene-based testing was used to identify demographic/clinical/genetic factors that influence the clearance of irinotecan and SN-38. Results: Irinotecan clearance was influenced by rs4149057 in SLCO1B1, body surface area, and co-administration of 5-fluorouracil/leucovorin/bevacizumab. SN-38 clearance was influenced by rs887829 in UGT1A1, pre-treatment total bilirubin, and EGFR rare variant burden. Within each UGT1A1 genotype group, elevated pre-treatment total bilirubin and/or presence of at least one rare variant in EGFR resulted in significantly lower SN-38 clearance. The model reduced the interindividual variability in irinotecan clearance from 38 to 34% and SN-38 clearance from 49 to 32%. Conclusions: This new model significantly reduced the interindividual variability in the clearance of irinotecan and SN-38. New genetic factors of variability in clearance have been identified

Genomic Characterization of Metformin Hepatic Response.

Metformin is used as a first-line therapy for type 2 diabetes (T2D) and prescribed for numerous other diseases. However, its mechanism of action in the liver has yet to be characterized in a systematic manner. To comprehensively identify genes and regulatory elements associated with metformin treatment, we carried out RNA-seq and ChIP-seq (H3K27ac, H3K27me3) on primary human hepatocytes from the same donor treated with vehicle control, metformin or metformin and compound C, an AMP-activated protein kinase (AMPK) inhibitor (allowing to identify AMPK-independent pathways). We identified thousands of metformin responsive AMPK-dependent and AMPK-independent differentially expressed genes and regulatory elements. We functionally validated several elements for metformin-induced promoter and enhancer activity. These include an enhancer in an ataxia telangiectasia mutated (ATM) intron that has SNPs in linkage disequilibrium with a metformin treatment response GWAS lead SNP (rs11212617) that showed increased enhancer activity for the associated haplotype. Expression quantitative trait locus (eQTL) liver analysis and CRISPR activation suggest that this enhancer could be regulating ATM, which has a known role in AMPK activation, and potentially also EXPH5 and DDX10, its neighboring genes. Using ChIP-seq and siRNA knockdown, we further show that activating transcription factor 3 (ATF3), our top metformin upregulated AMPK-dependent gene, could have an important role in gluconeogenesis repression. Our findings provide a genome-wide representation of metformin hepatic response, highlight important sequences that could be associated with interindividual variability in glycemic response to metformin and identify novel T2D treatment candidates