Francis, Stephen

Abstract Background The CCDC26 germline variant rs55705857 is causal for development of IDH mutant (IDHmut) adult glioma. However, ~60% of IDHmut patients do not carry the rs55705857 risk allele. We aimed to identify variants associated with developing IDHmut glioma among patients that do not have the rs55705857 risk allele and to further understand development of IDHwt glioma. Methods We used three datasets that included 1216 IDHmut and 1442 IDHwt glioma patients and a case–case design to perform genome-wide association (GWAS) analyses comparing IDHmut versus IDHwt glioma. Analyses were performed overall and stratified by rs55705857 genotype and sex. Multivariable logistic regression and regression trees were used to develop models to predict IDH status using germline variants, age, and contrast enhancement on MRI. Results Three regions were identified comparing IDHmut versus IDHwt: rs55705857 (meta P-value [P] = 1.35 × 10−43), PHLDB1 (rs7125115, P = 3.46 × 10−17), and D2HGDH (rs71430382, P = 2.43 × 10−12). When analyzing only patients that do not have the rs55705857 risk allele, PHLDB1 (rs7125115, P = 1.73 × 10−13) and D2HGDH (rs71430382, P = 8.86 × 10−10) were identified. Among patients who have the rs55705857 risk allele, four variants between ROBO1 and ROBO2 (rs4680975, P = 4.65 × 10−7) increased the likelihood of having an IDHwt tumor. Tumor expression of ROBO1 was associated with rs4680975 genotype in IDHwt patients that have the rs55705857 risk allele (P = 0.034). Multivariable logistic analysis demonstrated that rs55705857, rs71430382 (D2HGDH), and age predicted IDH mutation status. Conclusions To understand the development of adult glioma, we demonstrate that D2HGDH and PHLDB1 should be prioritized for functional studies in IDHmut tumors. The ROBO1 region warrants further investigation in IDHwt tumors.

Abstract Background Temozolomide (TMZ) treatment has demonstrated a variable impact on glioma prognosis. This study examines associations of survival with DNA repair gene germline polymorphisms among glioma patients who did and did not have TMZ treatment. Identifying genetic markers which sensitize tumor cells to TMZ could personalize therapy and improve outcomes. Methods We evaluated TMZ-related survival associations of pathogenic germline SNPs and genetically predicted transcript levels within 34 DNA repair genes among 1504 glioma patients from the University of California San Francisco Adult Glioma Study (UCSF AGS) and Mayo Clinic, whose diagnoses spanned pre- and post-TMZ eras within the major known glioma prognostic molecular subtypes. Results Among those who received TMZ, 5 SNPs were associated with overall survival, but not in those who did not receive TMZ. Only rs2308321-G, in MGMT, was associated with decreased survival (hazard ratio = 1.21, P = .019) for all glioma subtypes. Rs73191162-T (near UNG), rs13076508-C (near PARP3), rs7840433-A (near NEIL2), and rs3130618-A (near MSH5) were only associated with survival and TMZ treatment for certain subtypes, suggesting subtype-specific germline chemosensitization. Genetically predicted elevated expression of PNKP, compared to normal brain expression, was associated with markedly poor survival in TMZ-treated patients with isocitrate dehydrogenase (IDH)-mutant and 1p/19q non-codeleted gliomas (P = .015), with a median difference of over 70 months in overall survival times. Similarly, NEIL2 and TDG expressions were associated with altered TMZ-related survival only among certain subtypes. Conclusions Functional germline alterations within DNA repair genes were associated with TMZ sensitivity, measured by overall survival among adults with glioma. These variants should be evaluated in prospective analyses and functional studies.

Abstract Acquired tumor alterations are part of the WHO diagnosis of glioma. However, germline predisposition to these alterations is unknown. GWAS in European and Asian populations identified germline variants in 30 regions, many in or near genes that also acquire alterations, suggesting that interactions between inherited and acquired genetics are involved in glioma development. To evaluate such interactions, we used 3686 glioma cases and 1889 controls. A case-control design and multinomial logistic regression were used to identify germline variants associated with development of TERT-promoter mutant gliomas, generating two interesting observations. First, TERT germline variants were associated with predisposition to IDHwt glioma that have a TERT promoter mutation, but not with IDH-mutant codeleted gliomas that have a TERT promoter mutation. Second, two novel regions were observed in this more homogeneous subgroup of TERT mutated tumors: variants in TXNDC11 (OR=2.66, p=5.7x10-8) and RUVBL2 (OR=3.1, p=5.8x10-8). High tumor expression of TXNDC11 is associated with poor prognosis in glioma and RUVBL2 interacts with TERT to affect telomerase activity. Similar analyses are being performed on additional clinically relevant acquired alterations. In parallel, case-case GWAS analyses were performed to evaluate germline variants associated with other tumor alterations. Comparing IDH-mutant versus IDHwt stratified by rs55705857 genotype, the most significant variants were PHLDB1 (p=1.2x10-13) and D2HGDH (p=2.1x10-10), highlighting the importance of these two variants independent of CCDC26 for the development of IDH-mutant glioma. As a parallel approach, we utilized recursive partitioning and regression trees (rpart) to classify IDH mutation status using the 30 known variants. The top four branches of the regression tree included CCDC26, D2HGDH, PHLDB1, and MAML2, as well as two Asian variants (RAB27A, CYP4F12). This highlights the importance of utilizing variants discovered in non-European populations in analyses. Overall, understanding these interactions are necessary to develop therapeutics, stratify clinical trials, and to develop relevant tumor models.

Abstract Gliomas are heterogeneous brain tumors with prognostically-significant subtypes and few risk factors. IDH-wildtype glioblastoma, the most common subtype, carries a dismal prognosis. Using the largest GWAS dataset with histological (11,304 cases, 304,523 controls) and molecular (3,418 cases, 8,156 controls) subtypes, we analyzed the plasma and brain proteome to uncover proteins associated with glioma susceptibility. Using protein quantitative trait loci (pQTL) for 1,776 proteins in brain tissue, we identified 21 candidate proteins, including 9 associations with high probability (>70%) of colocalization (shared causal genetic variants for glioma risk and protein abundance). We confirmed known risk genes such as EGFR (p=1.9×10-35) and D2HGDH (p=1.1×10-5). We also identified novel risk proteins, including ENPP6, a choline phosphodiesterase overexpressed in glioma, for IDH-wildtype glioblastoma (OR=0.62, p=7.6×10-5), and galectin-3, a lectin encoded by a tumor fitness gene in CRISPR screens of glioma proliferation and a clinical drug target, for glioma overall (OR=1.37, p=1.3×10-5). Analyses of the plasma proteome using pQTLs for 1,362 proteins from 3 cohorts (N=88,838) identified 9 colocalized associations in whole blood. Most risk signals were subtype-specific, such as CRELD1 (p=2.5×10-5) for triple-negative glioblastoma. We found three candidate risk proteins for IDH-mutant 1p19q-intact glioma: immunoglobulin glycoprotein BCAM (p=9.6×10-8), neurotrophin receptor SorCS2 (p=3.6×10-5) and immune checkpoint PD-1 (p=1.1×10-5). Interestingly, genetically-predicted PD-1 levels had a larger effect on IDH-mutant 1p19q-intact (OR=2.75, 95% CI=1.61-4.69) than IDH-wildtype (OR=0.73, 95% CI=0.51-1.05) tumors, suggesting germline differences in immune checkpoint mechanisms among subtypes. For all colocalized candidates, genetically-predicted protein abundance was correlated with cis-regulated transcriptional activity in brain tissue. For example, genetically-predicted SORCS2 gene expression in brain tissue was associated with plasma SorCS2 levels (Z=16.1, p=1.1×10-68). Our analysis identified candidate proteins involved in immune response and neuronal proliferation, which may enable prioritization of drug targets and provide insight into disease mechanisms for glioma.

Abstract Background Polygenic risk scores (PRS) aggregate the contribution of many risk variants to provide a personalized genetic susceptibility profile. Since sample sizes of glioma genome-wide association studies (GWAS) remain modest, there is a need to efficiently capture genetic risk using available data. Methods We applied a method based on continuous shrinkage priors (PRS-CS) to model the joint effects of over 1 million common variants on disease risk and compared this to an approach (PRS-CT) that only selects a limited set of independent variants that reach genome-wide significance (P < 5 × 10–8). PRS models were trained using GWAS stratified by histological (10 346 cases and 14 687 controls) and molecular subtype (2632 cases and 2445 controls), and validated in 2 independent cohorts. Results PRS-CS was generally more predictive than PRS-CT with a median increase in explained variance (R2) of 24% (interquartile range = 11–30%) across glioma subtypes. Improvements were pronounced for glioblastoma (GBM), with PRS-CS yielding larger odds ratios (OR) per standard deviation (SD) (OR = 1.93, P = 2.0 × 10–54 vs. OR = 1.83, P = 9.4 × 10–50) and higher explained variance (R2 = 2.82% vs. R2 = 2.56%). Individuals in the 80th percentile of the PRS-CS distribution had a significantly higher risk of GBM (0.107%) at age 60 compared to those with average PRS (0.046%, P = 2.4 × 10–12). Lifetime absolute risk reached 1.18% for glioma and 0.76% for IDH wildtype tumors for individuals in the 95th PRS percentile. PRS-CS augmented the classification of IDH mutation status in cases when added to demographic factors (AUC = 0.839 vs. AUC = 0.895, PΔAUC = 6.8 × 10–9). Conclusions Genome-wide PRS has the potential to enhance the detection of high-risk individuals and help distinguish between prognostic glioma subtypes.

Abstract Background: Genome-wide association studies (GWAS) revealed that 25 regions in 24 genes are associated with adult diffuse glioma development. These regions were identified by performing GWAS of glioma overall and by pathology (GBM and nonGBM). The 2016 WHO Classification of Tumors of the Central Nervous System utilizes two somatic alterations to molecularly-classify adult diffuse glioma: IDH mutation and 1p/19q codeletion. We hypothesized that germline variants may increase susceptibility to, or interact with, these somatic alterations to accelerate the development of specific molecular subtypes of glioma. We further hypothesize that germline variants associated with IDH-mutated glioma might be associated with other IDH-mutated tumors, namely, cholangiocarcinoma, acute myeloid leukemia (AML) and melanoma. Methods: We performed a GWAS by glioma molecular subtype - as defined by presence or absence of IDH somatic mutation and 1p/19q codeletion. A total of 1320 glioma cases and 1889 controls were used in the discovery set, and 799 glioma cases and 808 controls in the validation set. A meta-analysis was performed with a genome-wide p-value threshold of 5 × 10−8. GTEx data were used to perform an expression quantitative trait loci (eQTL) analysis. For germline variants that were significantly associated with IDH-mutated glioma, we evaluated pleiotropy with cholangiocarcinoma, AML and melanoma using TCGA and Mayo Biobank controls. Results: Variants in or near D2HGDH on chromosome 2 were genome-wide significant in IDH-mutated glioma (meta p-value = 2.82 × 10−10). TCGA reported that the D2HGDH region was commonly deleted in IDH-mutated gliomas that do not have 1p/19q codeletion. In TCGA data for IDH-mutated, non-codeleted glioma, we observed that the D2HGDH variant was inversely associated with tumor deletions of D2HGDH (odds ratio=0.57, p-value=0.015). The eQTL analyses demonstrated significant associations between D2HGDH germline variant and expression of D2HGDH (p=2.2 × 10−11). Further stratifying IDH-mutated glioma by 1p/19q codeletion status, one variant near FAM20C on chromosome 7 was genome-wide significant in gliomas that have IDH mutation and 1p/19q codeletion (meta p-value=9.56 × 10−9). Analyses are currently underway to evaluate pleiotropy of these IDH-mutated glioma germline variants with other IDH-mutated tumors including cholangiocarcinoma, AML and melanoma. Variants in or near GMEB2 on chromosome 20 were genome-wide significant in IDH wild-type glioma (meta p-value=2.60 × 10−10). The most significant variant in the GMEB2 region remained significant after adjustment for the known RTEL1 glioma risk variant nearby on chromosome 20 (p=0.029). Conclusions: We identified and validated novel germline variants in two genes that are associated with etiology of IDH-mutated and one gene that is associated with IDH wild-type adult diffuse glioma. Citation Format: Jeanette E. Eckel-Passow, Kristen L. Drucker, Thomas M. Kollmeyer, Matthew L. Kosel, Paul A. Decker, Annette M. Molinaro, Terri Rice, Corrine E. Praska, Lauren E. Clark, Alissa A. Caron, Alexej Abyzov, Anthony Batzler, Jun S. Song, Melike Pekmezci, Helen M. Hansen, Lucie S. McCoy, Paige M. Bracci, Joseph Wiemels, John K. Wiencke, Stephen Francis, Terence C. Burns, Caterina Giannini, Daniel H. Lachance, Margaret Wrensch, Robert B. Jenkins. Adult diffuse glioma GWAS by molecular subtype identifies variants in D2HGDH, FAM20C and GMEB2 [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1193.

Abstract Background Twenty-five germline variants are associated with adult diffuse glioma, and some of these variants have been shown to be associated with particular subtypes of glioma. We hypothesized that additional germline variants could be identified if a genome-wide association study (GWAS) were performed by molecular subtype. Methods A total of 1320 glioma cases and 1889 controls were used in the discovery set and 799 glioma cases and 808 controls in the validation set. Glioma cases were classified into molecular subtypes based on combinations of isocitrate dehydrogenase (IDH) mutation, telomerase reverse transcriptase (TERT) promoter mutation, and 1p/19q codeletion. Logistic regression was applied to the discovery and validation sets to test for associations of variants with each of the subtypes. A meta-analysis was subsequently performed using a genome-wide P-value threshold of 5 × 10−8. Results Nine variants in or near D-2-hydroxyglutarate dehydrogenase (D2HGDH) on chromosome 2 were genome-wide significant in IDH-mutated glioma (most significant was rs5839764, meta P = 2.82 × 10−10). Further stratifying by 1p/19q codeletion status, one variant in D2HGDH was genome-wide significant in IDH-mutated non-codeleted glioma (rs1106639, meta P = 4.96 × 10−8). Further stratifying by TERT mutation, one variant near FAM20C (family with sequence similarity 20, member C) on chromosome 7 was genome-wide significant in gliomas that have IDH mutation, TERT mutation, and 1p/19q codeletion (rs111976262, meta P = 9.56 × 10−9). Thirty-six variants in or near GMEB2 on chromosome 20 near regulator of telomere elongation helicase 1 (RTEL1) were genome-wide significant in IDH wild-type glioma (most significant was rs4809313, meta P = 2.60 × 10−10). Conclusions Performing a GWAS by molecular subtype identified 2 new regions and a candidate independent region near RTEL1, which were associated with specific glioma molecular subtypes.

Abstract During the past six years, researchers have made major progress identifying common inherited genetic variation that increases risk for primary adult glioma. This paper summarizes knowledge about rare familial cancer syndromes that include adult glioma and reviews the available literature on the more recently discovered common inherited variation. Ten independent inherited variants in eight chromosomal regions have been convincingly associated with increased risk for adult glioma. Most of these variants increase relative risk of primary adult glioma by 20% to 40%, but the TP53 variant rs78378222 confers a two-fold relative risk (ie, 200%), and rs557505857 on chromosome 8 confers a six-fold relative risk of IDH-mutated astrocytomas and oligodendroglial tumors (ie, 600%). Even with a six-fold relative risk, the overall risk of developing adult glioma is too low for screening for the high-risk variant on chromosome 8. Future studies will help clarify which inherited adult glioma risk variants are associated with subtypes defined by histology and/or acquired tumor mutations. This review also provides an information sheet for primary adult glioma patients and their families.