Charbonneau, Bridget

Abstract Abstract 2702 Background: CXCR5 [chemokine (C-X-C motif) receptor 5; also known as Burkitt lymphoma receptor 1 (BCR1)] is expressed on mature B-cells, subsets of CD4+ and CD8+ T-cells, and skin-derived migratory dendritic cells. CXCL13 is the ligand for CXCR5, and together they are involved in guiding B-cells into the B-cell zones of secondary lymphoid organs as well as T-cell migration. CXCR5 is also expressed in malignant cells in NHL patients. Single nucleotide polymorphisms (SNPs) in CXCR5 have not been extensively studied, although two studies reported that SNPs from this chromosome 11 gene were associated with risk of primary biliary cirrhosis in a European population and with risk of NHL in an Asian population. This study evaluated the role of common germline genetic variation in the risk and prognosis of the common NHL subtypes of diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), mantle cell lymphoma (MCL), marginal zone lymphoma (MZL) and peripheral T-cell lymphoma (PTCL). Methods: Ten SNPs in CXCR5 were genotyped using a clinic-based study of 2694 NHL cases (including 586 DLBCL, 588 FL, 137 MCL, 230 MZL, and 158 PTCL) and 1521 controls enrolled from 2002–2009 in the Molecular Epidemiology Resource of the University of Iowa/Mayo Clinic Lymphoma SPORE. Clinical data were abstracted for all NHL patients and all were prospectively followed for disease progression, retreatment, transformation and death. SNPs were selected from HapMap using a standard tagging approach (r2≥0.80 and minor allele frequency (MAF) ≥0.05) and included SNPs 5kb upstream and downstream from the gene. Genotyping was conducted using a custom Illumina iSelect panel. All SNPs were in Hardy-Weinberg equilibrium in the control group. The most prevalent homozygous genotype was used as the reference group, and each SNP was modeled as having a log-additive effect in the regression model. For risk of NHL, odds ratios (ORs) and 95% confidence intervals were calculated using unconditional logistic regression, adjusted for age and gender. For prognosis, hazard ratios (HR) were calculated for both event-free (EFS) and overall (OS) survival using Cox proportional hazards regression, adjusting for age, treatment type, and IPI (DLBCL, PTCL), FLIPI (FL), or MIPI (MCL). To determine risk of FL transformation to DLBCL, a competing risk Cox model was used. A p<0.05 was declared as statistically significant. Results: Of the 10 CXCR5 tag SNPs in our study, 5 were associated with risk of NHL, with rs1790192 (MAF=0.39 among controls) having the strongest association (OR=1.19, 95%CI 1.08–1.30; p=0.0003). This SNP was strongly associated with FL (OR=1.44, 95%CI 1.25–1.66; p=3.1×10−7), and was also associated with DLBCL (OR=1.16, 95%CI 1.01–1.33; p=0.038) and PTCL (OR=1.29, 95%CI 1.02–1.64; p=0.037) but not MCL or MZL. For FL, none of the SNPs were associated with EFS or OS, although rs1790192 was associated with better outcome among FL patients that were observed (HR=0.64; 95%CI 0.47–0.87; p=0.0039). For DLBCL, none of the SNPs were associated with EFS, while three SNPs were associated with OS: rs523604 (HR=0.79; 95%CI 0.64–0.97; p=0.028), rs12363277 (HR=0.57; 95%CI 0.33–0.99; p=0.046), and rs3922 (HR=0.79; 95%CI 0.65–0.97; p=0.027). The latter SNP was also associated with risk of transformation from FL to DLBCL (HR=0.61; 95%CI 0.40–0.91; p=0.016). For MCL, MZL and PTCL, none of the SNPs were associated with either EFS or OS. Conclusions: Several germline CXCR5 polymorphisms were associated with NHL risk, particularly rs1790192 with risk of FL as well as DLBCL and PTCL. CXCR5 SNPs did not appear to be important in prognosis for most NHL subtypes, although rs1790192 was associated with a better outcome in untreated FL and rs3922 was associated with OS in DLBCL patients and with risk of FL transformation. These results provide additional evidence for a role of host genetic variation in CXCR5 in lymphomagenesis, mainly in risk of disease development, although there may also be a role for CXCR5 in FL and DLBCL prognosis. Replications of these findings in additional studies are needed. Disclosures: Ansell: Seattle Genetics, Inc.: Research Funding.

Abstract Abstract 149 Background. There are few established factors that predict risk of follicular lymphoma (FL) transformation, and only two studies have evaluated the role of germline genetic variation and risk of transformation. One study reported an association of a BCL6 SNP (397C>G) with risk of transformation, and a second study reported an association with rs6457327, which is in the HLA region of 6p21. This study evaluated the risk of FL transformation in the modern treatment era with germline SNPs that were previously reported to be associated with FL transformation (BCL6SNPs and rs6457327), along with SNPs (N=45) previously associated with either risk of FL or FL prognosis. Methods. Newly diagnosed FL patients were prospectively enrolled from 2002–2009 into the Molecular Epidemiology Resource of the University of Iowa/Mayo Clinic Lymphoma SPORE. Peripheral blood for genotyping was collected at enrollment, and clinical and treatment data were abstracted from medical records. All patients were actively followed for disease progression, retreatment, transformation and death. For this analysis, only FL patients with grade I-IIIa were included. Patients with composite DLBCL, FL grade IIIb, or other evidence of transformation at diagnosis were excluded. Transformation was based on either 1) biopsy confirmation of FL grade IIIb, DLBCL, or higher grade B-cell lymphoma; or 2) clinical indicators of transformation based on the published literature, including sudden rise in LDH, rapid discordant localized nodal growth, new involvement of unusual extranodal sites, new B-symptoms, or hypercalcemia. Genotyping was conducted using a custom Illumina iSelect panel. Cox regression was used to estimate Hazard Ratios (HRs) and 95% confidence intervals (CI) for individual SNPs with time to transformation, with death modeled as a competing risk. Each SNP was modeled with the most prevalent homozygous genotype used as the reference group, and each SNP was modeled as having a log-additive (per minor allele) effect in the regression model. An ordinal test was used to assess the trend, and p<0.05 was considered statistically significant. Results. There were 631 newly diagnosed grade I-IIIa FL patients, of whom 532 had DNA and were successfully genotyped. The median age at diagnosis was 60 years (range 23–93) and 53% were male. FLIPI distribution was 42% 0–1, 34% 2 and 24% 3–5. Initial therapies included observation (32%), rituximab (R) monotherapy (13%), alkylator-based chemotherapy +/− R (20%), and anthracycline-based chemotherapy +/− R (19%). At a median of 59 months of follow-up (range, 11–110), 58 patients died, 254 had an event (death, progression or retreatment) and 54 patients (10%) transformed. Transformation was biopsy proven in 43 of the 54 patients (80%). The SNP rs6457327 on chromosome 6p that was previously reported to be associated with transformation was positively associated with risk of transformation in our data (HR=1.40; 95%CI 0.97–2.04; p=0.07), although this association was weaker but still consistent with the prior report. Further, we evaluated additional typed SNPs from the HLA region of 6p, and found additional associations (p<0.05) with HLA-DRA (3 SNPs), HLA-DQA2 (2 SNPs), HLA-DOA (2 SNPs) and HLA-DPB1/HLA-DPA2 (2 SNPs), of which rs10484569 from HLA-DPB1/HLA-DPA2 was the most strongly associated with risk of transformation (HR=3.01; 95%CI 1.48–6.09; p=0.002). None of the BCL6 or other previously reported risk SNPs were associated with transformation. Of the SNPs previously associated with FL prognosis, only the FCGR2A rs1801274 (HR=1.41; 95%CI 0.94–2.10; p=0.09) and CD46 (CD46 molecule, complement regulatory protein) rs2466571 (HR=1.72; 95%CI 1.16–2.55; p=0.007) were associated with risk of transformation. For CD46, one additional SNP (rs2796269) was also associated with transformation (HR=0.61; 95%CI 0.40–0.95; p=0.03). Further adjustment for FLIPI score and treatment (observation versus all other) did not alter these associations. Conclusions. The previously reported association of rs6457327 with risk of transformation in the modern treatment era was replicated. In addition, several HLA loci and two FL prognosis SNPs were linked to transformation risk. The remaining published FL risk or prognosis SNPs were not associated with transformation. Our results support a role for germline genetic variation in the HLA region, FCRGR2A and CD46 in transformation of FL. Disclosures: No relevant conflicts of interest to declare.

Abstract Tumor infiltrating regulatory T cells (Tregs) appear to play a role in survival in a number of solid tumors, including ovarian cancer. We assessed genetic variation via tag SNPs (N=749) in 25 Treg associated genes (CD28, CTLA4, FOXP3, IDO1, IL10, IL10RA, IL15, 1L17RA, IL23A, IL23R, IL2RA, IL6, IL6R, IL8, LGALS1, LGALS9, MAP3K8, STAT5A, STAT5B, TGFB1, TGFB2, TGFB3, TGFBR1, TGRBR2, and TGFBR3) in relation to ovarian cancer survival. SNPs were selected based on r2 ≥ 0.8, MAF ≥ 0.05, 5 kb upstream and downstream from the gene. We had SNP and outcome data for ovarian cancer from 10,084 combined ovarian cancer cases, including 5,248 high grade serous, 1,452 endometrioid, 661 mucinous, and 795 clear cell cases of European descent across multiple studies from the Ovarian Cancer Association Consortium (OCAC). Cox proportional hazards regression modeling was used to estimate per-allele hazard ratios (HRs) and 95% confidence intervals (CIs) for associations with overall survival, adjusting for age at diagnosis, population substructure PCs, study site, histology (for the all case analysis), tumor stage, tumor grade and oral contraceptive use. The strongest associations were found for endometrioid histologic subtype and IL2RA SNPs (maximum r2=0.67) rs11256497 [HR=1.42, 95% CI: 1.22-1.64; p=5.7 x 10−6], rs791587 [HR=1.36, 95% CI: 1.17-1.57; p=6.2 x 10−5], rs2476491 [HR=1.40, 95% CI: 1.19-1.64; p=5.6 x 10−5], and rs10795763 [HR=1.35, 95% CI: 1.17-1.57; p=7.9 x 10−5], as well as clear cell ovarian cancer and CTLA4 SNP rs231775 [HR=0.67, 95% CI: 0.54-0.82; p=9.3 x 10−5]. No other associations were observed at p&lt;1 x 10−4. Overall, this study provided evidence that variation in genes related to Tregs, particularly IL2RA and CTLA4, influence ovarian cancer survival. Citation Format: Bridget Charbonneau, Keith Knutson, Robert A. Vierkant, Julie M. Cunningham, Zachary C. Fogarty, Kimberly R. Kalli, Matthew J. Maurer, Ann L. Oberg, Brooke L. Fridley, Claudia C. Preston, Paul D.P. Pharoah, Susan J. Ramus, Catherine M. Phelan, Kunle Odunsi, Kirsten Moysich, Ellen L. Goode. Polymorphisms in regulatory T cell related genes and ovarian cancer survival. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4844. doi:10.1158/1538-7445.AM2013-4844

Abstract Survival in epithelial ovarian cancer (EOC) is influenced by the host immune response, yet the key genetic determinants of inflammation and immunity that affect prognosis are not known. The nuclear factor-κB (NF-κB) transcription factor family plays an important role in many immune and inflammatory responses, including the response to cancer. We studied common inherited variation in 210 genes in the NF-κB family in 10,084 patients with invasive EOC (5,248 high-grade serous, 1,452 endometrioid, 795 clear cell, and 661 mucinous) from the Ovarian Cancer Association Consortium. Associations between genotype and overall survival were assessed using Cox regression for all patients and by major histology, adjusting for known prognostic factors and correcting for multiple testing (threshold for statistical significance, P &lt; 2.5 × 10−5). Results were statistically significant when assessed for patients of a single histology. Key associations were with caspase recruitment domain family, member 11 (CARD11) rs41324349 in patients with mucinous EOC [HR, 1.82; 95% confidence interval (CI), 1.41–2.35; P = 4.13 × 10−6] and tumor necrosis factor receptor superfamily, member 13B (TNFRSF13B) rs7501462 in patients with endometrioid EOC (HR, 0.68; 95% CI, 0.56–0.82; P = 2.33 × 10−5). Other associations of note included TNF receptor–associated factor 2 (TRAF2) rs17250239 in patients with high-grade serous EOC (HR, 0.84; 95% CI, 0.77–0.92; P = 6.49 × 10−5) and phospholipase C, gamma 1 (PLCG1) rs11696662 in patients with clear cell EOC (HR, 0.43; 95% CI, 0.26–0.73; P = 4.56 × 10−4). These associations highlight the potential importance of genes associated with host inflammation and immunity in modulating clinical outcomes in distinct EOC histologies. Cancer Epidemiol Biomarkers Prev; 23(7); 1421–7. ©2014 AACR.

Abstract To reveal biologic mechanisms underlying clinical outcome of high-grade serous (HGS) epithelial ovarian carcinomas (EOC), we evaluated the association between tumor epigenetic changes and time to recurrence (TTR). We assessed methylation at approximately 450,000 genome-wide CpGs in tumors of 337 Mayo Clinic (Rochester, MN) patients. Semi-supervised clustering of discovery (n = 168) and validation (n = 169) sets was used to determine clinically relevant methylation classes. Clustering identified two methylation classes based on 60 informative CpGs, which differed in TTR in the validation set [R vs. L class, P = 2.9 × 10−3, HR = 0.52; 95% confidence interval (CI), 0.34–0.80]. Follow-up analyses considered genome-wide tumor mRNA expression (n = 104) and CD8 T-cell infiltration (n = 89) in patient subsets. Hypomethylation of CpGs located in 6p21.3 in the R class associated with cis upregulation of genes enriched in immune response processes (TAP1, PSMB8, PSMB9, HLA-DQB1, HLA-DQB2, HLA-DMA, and HLA-DOA), increased CD8 T-cell tumor infiltration (P = 7.6 × 10−5), and trans-regulation of genes in immune-related pathways (P = 1.6 × 10−32). This is the most comprehensive assessment of clinical outcomes with regard to epithelial ovarian carcinoma tumor methylation to date. Collectively, these results suggest that an epigenetically mediated immune response is a predictor of recurrence and, possibly, treatment response for HGS EOC. Cancer Res; 74(11); 3084–91. ©2014 AACR.

Abstract The presence of regulatory T cells (Treg) in solid tumors is known to play a role in patient survival in ovarian cancer and other malignancies. We assessed inherited genetic variations via 749 tag single-nucleotide polymorphisms (SNP) in 25 Treg-associated genes (CD28, CTLA4, FOXP3, IDO1, IL10, IL10RA, IL15, 1L17RA, IL23A, IL23R, IL2RA, IL6, IL6R, IL8, LGALS1, LGALS9, MAP3K8, STAT5A, STAT5B, TGFB1, TGFB2, TGFB3, TGFBR1, TGRBR2, and TGFBR3) in relation to ovarian cancer survival. We analyzed genotype and overall survival in 10,084 women with invasive epithelial ovarian cancer, including 5,248 high-grade serous, 1,452 endometrioid, 795 clear cell, and 661 mucinous carcinoma cases of European descent across 28 studies from the Ovarian Cancer Association Consortium (OCAC). The strongest associations were found for endometrioid carcinoma and IL2RA SNPs rs11256497 [HR, 1.42; 95% confidence interval (CI), 1.22–1.64; P = 5.7 × 10−6], rs791587 (HR, 1.36; 95% CI, 1.17–1.57; P = 6.2 × 10−5), rs2476491 (HR, = 1.40; 95% CI, 1.19–1.64; P = 5.6 × 10−5), and rs10795763 (HR, 1.35; 95% CI, 1.17–1.57; P = 7.9 × 10−5), and for clear cell carcinoma and CTLA4 SNP rs231775 (HR, 0.67; 95% CI, 0.54–0.82; P = 9.3 × 10−5) after adjustment for age, study site, population stratification, stage, grade, and oral contraceptive use. The rs231775 allele associated with improved survival in our study also results in an amino acid change in CTLA4 and previously has been reported to be associated with autoimmune conditions. Thus, we found evidence that SNPs in genes related to Tregs seem to play a role in ovarian cancer survival, particularly in patients with clear cell and endometrioid epithelial ovarian cancer. Cancer Immunol Res; 2(4); 332–40. ©2014 AACR.

Abstract Background: Ovarian cancer is a leading cause of cancer-related death among women. In an effort to understand contributors to disease outcome, we evaluated single-nucleotide polymorphisms (SNP) previously associated with ovarian cancer recurrence or survival, specifically in angiogenesis, inflammation, mitosis, and drug disposition genes. Methods: Twenty-seven SNPs in VHL, HGF, IL18, PRKACB, ABCB1, CYP2C8, ERCC2, and ERCC1 previously associated with ovarian cancer outcome were genotyped in 10,084 invasive cases from 28 studies from the Ovarian Cancer Association Consortium with over 37,000-observed person-years and 4,478 deaths. Cox proportional hazards models were used to examine the association between candidate SNPs and ovarian cancer recurrence or survival with and without adjustment for key covariates. Results: We observed no association between genotype and ovarian cancer recurrence or survival for any of the SNPs examined. Conclusions: These results refute prior associations between these SNPs and ovarian cancer outcome and underscore the importance of maximally powered genetic association studies. Impact: These variants should not be used in prognostic models. Alternate approaches to uncovering inherited prognostic factors, if they exist, are needed. Cancer Epidemiol Biomarkers Prev; 22(5); 987–. ©2013 AACR.

Abstract As pathways of the immune system play an important role in multiple malignancies, we assessed the association between single-nucleotide polymorphisms (SNPs) in inflammation-related genes and risk of ovarian cancer in a multi-site case-control study. Using a custom array, we genotyped 930 epithelial ovarian cancer cases and 1,037 controls for SNPs in ALOX12, ALOX15, ALOX5, CCL11, CCL2, CCL3, CCR3, CRP, CXCL16, IL10, IL15RA, IL18, IL1A, IL1B, IL1RN, IL4R, IL6, IL6R, IL7R, IL8RA, IL8RB, IL9, NOS3, PTGS1, PTGS2, TLR2, and TNF. After analysis by logistic regression, SNPs with p&lt;0.10 were evaluated in an independent data set of 3,143 cases and 2,102 controls from the Follow-up of Ovarian Cancer Genetic Association and Interaction Studies (FOCI) collaboration. Combined analysis revealed association with interleukin 1, alpha (IL1A) SNPs (rs17561 and rs4848300, r2=0.990, p=0.005) which varied by histological subtype (heterogeneity p=0.03). For example, IL1A rs17561, which results in a missense change in amino acid at position 114 and correlates with multiple inflammatory phenotypes, was inversely associated with risk of clear cell, mucinous, and endometrioid subtypes, but not with the serous subtype (Table 1). Additional data on IL1A rs17561 in an even larger collection of ovarian cancer cases and controls are pending and will be analyzed soon. We also found that genotype at rs1864414 in ALOX5 (arachidonate 5-lipoxygenase) was associated with decreased risk combined across subtypes (OR 0.86, 95% CI 0.78-0.95; p=0.002). Thus, inherited variation in IL1A and ALOX5 appears to have a role in ovarian cancer risk, though it is limited to rarer subtypes for IL1A. These data add to existing literature on the importance of inflammation in tumorigenesis and growing evidence of subtype-specific traits in ovarian cancer and may provide important clues on prevention. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2603. doi:1538-7445.AM2012-2603

Abstract The importance of inflammation pathways to the development of many human cancers prompted us to examine the associations between single-nucleotide polymorphisms (SNP) in inflammation-related genes and risk of ovarian cancer. In a multisite case–control study, we genotyped SNPs in a large panel of inflammatory genes in 930 epithelial ovarian cancer cases and 1,037 controls using a custom array and analyzed by logistic regression. SNPs with P &lt; 0.10 were evaluated among 3,143 cases and 2,102 controls from the Follow-up of Ovarian Cancer Genetic Association and Interaction Studies (FOCI) post-GWAS collaboration. Combined analysis revealed association with SNPs rs17561 and rs4848300 in the interleukin gene IL1A which varied by histologic subtype (Pheterogeneity = 0.03). For example, IL1A rs17561, which correlates with numerous inflammatory phenotypes, was associated with decreased risk of clear cell, mucinous, and endometrioid subtype, but not with the most common serous subtype. Genotype at rs1864414 in the arachidonate 5-lipoxygenase ALOX5 was also associated with decreased risk. Thus, inherited variation in IL1A and ALOX5 seems to affect ovarian cancer risk which, for IL1A, is limited to rarer subtypes. Given the importance of inflammation in tumorigenesis and growing evidence of subtype-specific features in ovarian cancer, functional investigations will be important to help clarify the importance of inherited variation related to inflammation in ovarian carcinogenesis. Cancer Res; 72(5); 1064–9. ©2012 AACR.