Caporaso, Neil E.

Abstract Background Lung adenocarcinoma in never-smokers is a major public health burden, especially among East Asian women. Polygenic risk scores (PRSs) are promising for risk stratification but are primarily developed in European-ancestry populations. We aimed to develop and validate single- and multi-ancestry PRSs for East Asian never-smokers to improve lung adenocarcinoma risk prediction. Methods PRSs were developed using genome-wide association study summary statistics from East Asian (8002 cases; 20 782 controls) and European (2058 cases; 5575 controls) populations. Single-ancestry models included a PRS with 25 single nucleotide variants (PRS-25), PRS clumping and thresholding (CT), and LDpred2; multi-ancestry models included LDpred2 plus European PRS-128, PRS-CSx, and clumping and thresholding, super learning, empirical Bayes (CT-SLEB). Performance was evaluated in independent East Asian data from the Female Lung Cancer Consortium and externally validated in the Nanjing Lung Cancer Cohort. We assessed predictive accuracy via area under the curve operating procedure (AUC), with 10-year and (ages 30-80 years) absolute risks estimates. Results The best multi-ancestry PRS, using East Asian and European data via CT-SLEB, outperformed the best East Asian-only (LDpred2; AUC = 0.629, 95% confidence interval [CI] = 0.618 to 0.641), achieving an AUC of 0.640 (95% CI = 0.629 to 0.653) and odds ratio of 1.71 (95% CI = 1.61 to 1.82) per standard deviation increase. Nanjing Lung Cancer Cohort validation confirmed robust performance (AUC = 0.649, 95% CI = 0.623 to 0.676). The top 20% PRS group had a 3.92-fold higher lung adenocarcinoma risk than the bottom 20%. Further, the top 5% PRS group reached a 6.69% lifetime absolute risk. Notably, this group reached the average population 10-year lung adenocarcinoma risk at age 50 years (0.42%) by age 41 years, 9 years earlier. Conclusions Multi-ancestry PRS approaches enhance lung adenocarcinoma risk stratification in East Asian never-smokers, with consistent external validation, suggesting future clinical utility.

ABSTRACT Understanding lung cancer evolution can identify tools for intercepting its growth. In a landscape analysis of 1024 lung adenocarcinomas (LUAD) with deep whole-genome sequencing integrated with multiomic data, we identified 542 LUAD that displayed diverse clonal architecture. In this group, we observed an interplay between mobile elements, endogenous and exogenous mutational processes, distinct driver genes, and epidemiological features. Our results revealed divergent evolutionary trajectories based on tobacco smoking exposure, ancestry, and sex. LUAD from smokers showed an abundance of tobacco-related C:G>A:T driver mutations in KRAS plus short subclonal diversification. LUAD in never smokers showed early occurrence of copy number alterations and EGFR mutations associated with SBS5 and SBS40a mutational signatures. Tumors harboring EGFR mutations exhibited long latency, particularly in females of European-ancestry (EU_N). In EU_N, EGFR mutations preceded the occurrence of other driver genes, including TP53 and RBM10. Tumors from Asian never smokers showed a short clonal evolution and presented with heterogeneous repetitive patterns for the inferred mutational order. Importantly, we found that the mutational signature ID2 is a marker of a previously unrecognized mechanism for LUAD evolution. Tumors with ID2 showed short latency and high L1 retrotransposon activity linked to L1 promoter demethylation. These tumors exhibited an aggressive phenotype, characterized by increased genomic instability, elevated hypoxia scores, low burden of neoantigens, propensity to develop metastasis, and poor overall survival. Re-activated L1 retrotransposition-induced mutagenesis can contribute to the origin of the mutational signature ID2, including through the regulation of the transcriptional factor ZNF695 , a member of the KZFP family. The complex nature of LUAD evolution creates both challenges and opportunities for screening and treatment plans.

Introduction: Over 80,000 new cases of Non-Hodgkin's Lymphoma (NHL) are diagnosed annually, but modifiable risk factors are largely unknown. Circadian disruption has been identified as a risk factor for several cancers, including breast and prostate cancer, and affects immune cells such as Natural Killer (NK) cells and T-helper cells. Disruptions in circadian rhythms can lead to aberrant immune cell trafficking and proliferation. Despite this biological connection, evidence linking circadian disruptions to NHL remains inconclusive. Only single Mendelian Randomization (MR) study (PMID: 32895918) has explored the causal relationship between sleep traits and overall NHL risk, without examining NHL subtypes. Studies have shown distinct risk factor profiles among NHL subtypes. Therefore, this study aims to investigate the causal relationship between sleep traits and NHL subtypes in European populations using MR. Methods: We conducted a two-sample MR analyses using genome-wide association studies (GWAS) summary data on sleep traits and NHL subtypes. The GWAS summary statistics on sleep traits were obtained from publicly available UKBiobank datasets. The sleep traits analyzed included chronotype (N=449,734), insomnia (N=237,627), sleep duration (N=446,118), daytime sleepiness (N=452,071), short sleep (N=411,934), and long sleep (N=339,926). Only independent, genome-wide significant SNPs (p < 5×10-8) were considered as valid instruments. The NHL subtype-specific summary statistics came from the InterLymph including follicular lymphoma (FL: N=6,508 cases / 64,183 controls), mantle cell lymphoma (MCL: N=1,169 cases / 61,603 controls), and Waldenström macroglobulinemia/lymphoplasmacytic lymphoma (WM/LPL: N=1,697 cases / 59,333 controls). Our primary analysis used the random-effects inverse-variance weighted (IVW) method. In addition, we employed MR-Egger to adjust for directional pleiotropy. Heterogeneity in the MR results was evaluated with Cochran's Q statistic. Results: The IVW method did not identify any statistically significant causal association between sleep traits and the risk of NHL subtypes (FL,MCL,WM/LPL), but did yield several intriguing associations. Chronotype (early rising) showed a tendency towards a marginally an increased NHL risk across subtypes [Odds ratio (OR) range: 1.07 - 2.56]. Conversely, a decreased risk was noted for long sleep (≥ 9 hours vs. 7-8 hours) [OR range: 0.19 - 0.62], insomnia [OR range: 0.72 - 0.88], and short sleep (excludes MCL) [OR range: 0.12 - 0.72]. Confidence limit for long sleep was wide due to small number of variants (N=8). The MR-Egger regression analysis did not provide evidence of significant directional horizontal pleiotropy. The F-statistics for all sleep traits were greater than 10 indicating absences of weak instrument bias. Conclusions: Our study found no statistically significant associations between sleep traits and three NHL subtypes (FL, MCL, WM/LPL) in European populations. However, we observed potential influences of chronotype, long and short sleep, and insomnia on NHL risk. These findings align with and extend previous MR study by providing subtype-specific insights. Future directions include expanding our analysis to additional NHL subtypes, generating polygenic risk scores, and conducting stratified analyses using the InterLymph population. These efforts aim to further elucidate the complex relationship between sleep characteristics and NHL risk across diverse subtypes and populations.

Abstract Background: Clinical, molecular, and genetic epidemiology studies displayed remarkable differences between ever- and never-smoking lung cancer. Methods: We conducted a stratified multi-population (European, East Asian, and African descent) association study on 44,823 ever-smokers and 20,074 never-smokers to identify novel variants that were missed in the non-stratified analysis. Functional analysis including expression quantitative trait loci (eQTL) colocalization and DNA damage assays, and annotation studies were conducted to evaluate the functional roles of the variants. We further evaluated the impact of smoking quantity on lung cancer risk for the variants associated with ever-smoking lung cancer. Results: Five novel independent loci, GABRA4, intergenic region 12q24.33, LRRC4C, LINC01088, and LCNL1 were identified with the association at two or three populations (P < 5 × 10−8). Further functional analysis provided multiple lines of evidence suggesting the variants affect lung cancer risk through excessive DNA damage (GABRA4) or cis-regulation of gene expression (LCNL1). The risk of variants from 12 independent regions, including the well-known CHRNA5, associated with ever-smoking lung cancer was evaluated for never-smokers, light-smokers (packyear ≤ 20), and moderate-to-heavy-smokers (packyear > 20). Different risk patterns were observed for the variants among the different groups by smoking behavior. Conclusions: We identified novel variants associated with lung cancer in only ever- or never-smoking groups that were missed by prior main-effect association studies. Impact: Our study highlights the genetic heterogeneity between ever- and never-smoking lung cancer and provides etiologic insights into the complicated genetic architecture of this deadly cancer.

Abstract Differences by sex in lung cancer incidence and mortality have been reported which cannot be fully explained by sex differences in smoking behavior, implying existence of genetic and molecular basis for sex disparity in lung cancer development. However, the information about sex dimorphism in lung cancer risk is quite limited despite the great success in lung cancer association studies. By adopting a stringent two-stage analysis strategy, we performed a genome-wide gene–sex interaction analysis using genotypes from a lung cancer cohort including ~ 47 000 individuals with European ancestry. Three low-frequency variants (minor allele frequency < 0.05), rs17662871 [odds ratio (OR) = 0.71, P = 4.29×10−8); rs79942605 (OR = 2.17, P = 2.81×10−8) and rs208908 (OR = 0.70, P = 4.54×10−8) were identified with different risk effect of lung cancer between men and women. Further expression quantitative trait loci and functional annotation analysis suggested rs208908 affects lung cancer risk through differential regulation of Coxsackie virus and adenovirus receptor gene expression in lung tissues between men and women. Our study is one of the first studies to provide novel insights about the genetic and molecular basis for sex disparity in lung cancer development.

Abstract Background: Somatic EGFR mutations define a subset of non–small cell lung cancers (NSCLC) that have clinical impact on NSCLC risk and outcome. However, EGFR-mutation-status is often missing in epidemiologic datasets. We developed and tested pragmatic approaches to account for EGFR-mutation-status based on variables commonly included in epidemiologic datasets and evaluated the clinical utility of these approaches. Methods: Through analysis of the International Lung Cancer Consortium (ILCCO) epidemiologic datasets, we developed a regression model for EGFR-status; we then applied a clinical-restriction approach using the optimal cut-point, and a second epidemiologic, multiple imputation approach to ILCCO survival analyses that did and did not account for EGFR-status. Results: Of 35,356 ILCCO patients with NSCLC, EGFR-mutation-status was available in 4,231 patients. A model regressing known EGFR-mutation-status on clinical and demographic variables achieved a concordance index of 0.75 (95% CI, 0.74–0.77) in the training and 0.77 (95% CI, 0.74–0.79) in the testing dataset. At an optimal cut-point of probability-score = 0.335, sensitivity = 69% and specificity = 72.5% for determining EGFR-wildtype status. In both restriction-based and imputation-based regression analyses of the individual roles of BMI on overall survival of patients with NSCLC, similar results were observed between overall and EGFR-mutation-negative cohort analyses of patients of all ancestries. However, our approach identified some differences: EGFR-mutated Asian patients did not incur a survival benefit from being obese, as observed in EGFR-wildtype Asian patients. Conclusions: We introduce a pragmatic method to evaluate the potential impact of EGFR-status on epidemiological analyses of NSCLC. Impact: The proposed method is generalizable in the common occurrence in which EGFR-status data are missing.

Abstract There is growing evidence showing that sex disparity in lung cancer risk cannot be fully explained by sex differences in smoking behavior, implying existence of genetic and molecular basis for sex disparity in lung cancer development. However, the information about sex dimorphism in lung cancer risk is quite limited despite the great success in lung cancer association studies. By adopting a stringent two-stage analysis strategy, we performed a genome-wide gene-sex interaction analysis using genotype from a lung cancer cohort including ~ 47,000 individuals with European ancestry. Three low-frequency variants (minor allele frequency < 0.05), rs17662871 (OR=0.71, p=4.29x10-8); rs79942605 (OR=2.17, p=2.81x10-8); and rs208908 (OR=0.70, p=4.54x10-8), were identified with different risk effect of lung cancer between men and women. Further eQTL and functional annotation analysis suggested rs208908 affect lung cancer risk through differential regulation of CXADR (Coxsackie Virus And Adenovirus Receptor) gene expression in lung tissues between men and women. Our study is one of the first studies to provide novel insights about the genetic and molecular basis for sex disparity in lung cancer development.

Abstract Background and Significance : Chronic lymphocytic leukemia (CLL) is the most heritable hematologic malignancy; however, no common CLL predisposition genes are known. Monoclonal B lymphocytosis (MBL) is a hematologic syndrome characterized by small accumulations of B lymphocytes in the peripheral blood. MBL has a CLL-like immunophenotype, may progress to overt CLL, and is over represented in CLL families. Therefore, MBL observed in the context of familial CLL may be a marker of inherited risk for development of CLL. Detailed characterization of family-associated MBL may also provide mechanistic insights into the pathogenesis of familial CLL. Our strategy was to detail the biologic characteristics of CLL in family-associated MBL. Methods : Persons with MBL were identified by flow cytometric screening of peripheral blood from unaffected members of CLL kindreds ascertained by Genetic Epidemiology of CLL Consortium (GEC) sites. Flow cytometry was used to determine the surface immunophenotype including CD38 and intracellular ZAP-70. We defined MBL as populations of CD19+, CD5+, CD20lo, CD23+ B cells that comprised at least 2% of the CD19+ peripheral B cell compartment and did not exceed 5.0 × 109 MBL cells/L. RNA and genomic DNA from single MBL cells isolated by flow cytometric sorting were analyzed using PCR to determine immunoglobulin heavy and light chain sequences. MBL cells were sorted in bulk for FISH for loci associated with clinical CLL. Results : Twenty-two out of 190 (12%) unaffected family members were found to have MBL. We observed significant variability in the size of the MBL clone as a percentage of the CD19+ B cell compartment (mean 32%; range 2%–97%). Nonetheless, the absolute size of the MBL clone was small, 15 of 17 individuals had &lt; 200 × 106 MBL cells/L. CD38 expression (defined as CD38 surface expression in ≥30% of MBL cells) was observed in 8 of 18 subjects tested. ZAP-70 (defined as intracellular expression in ≥ 20% of MBL cells) was expressed in 4 of 12 participants. Among 12 subjects tested, 5 MBL expressed both surface IgD and IgM, 3 expressed IgD only, 2 expressed IgM only, and 2 did not express IgD or IgM. Analysis of immunoglobulin heavy and light chains has been completed in 8 individuals. Both immunoglobulin heavy chain variable (IgVH) region mutated (n = 12) and unmutated (n = 4) sequences were observed. Four of 8 individuals had 2 or more unrelated MBL clones (range 2–5), including one individual with both unmutated and mutated clones. Among the 16 MBL clones identified in these 8 subjects, VH3 or VH4 rearrangements were observed in all MBL clones. The most commonly rearranged IgVH genes were 3–07 (3 MBL clones), 3–15 (3), and 4–34 (3). No VH1 family gene rearrangements were observed. In one individual, a VH3–07 MBL clone showed intraclonal diversification suggestive of antigen driven immunoglobulin sequence changes. Twenty productive light chain rearrangements were identified among the 16 MBL clones, with 11 Vλ and 9 Vκ genes used. We observed 6 productive rearrangements of Vλ1–51. MBL cells were bulk sorted for FISH from 9 subjects. Mono or biallelic deletion of 13q14.3 was observed in 5 subjects, the other 4 were normal. Conclusions : Our findings confirm that MBL is commonly observed among the unaffected family members from CLL kindreds. We found that some MBL clones express ZAP-70, CD38 or have unmutated IgVH genes and thus are similar to clinical CLL. The clinical outcome of these MBL clones in relation to our baseline prognostic characterization will be of interest. Small MBL clones are commonly oligoclonal. Importantly, although the immunoglobulin heavy and light chain genes rearranged in these MBL clones are all commonly used in CLL, the absence of VH1 and abundance of Vλ1–51 rearrangements do suggests differences in BCR usage between CLL and MBL. Further investigation of family associated MBL may clarify the genetics and immunobiology of familial CLL.

Abstract Background: Lipid traits have been inconsistently linked to risk of non-Hodgkin lymphoma (NHL). We examined the association of genetically predicted lipid traits with risk of diffuse large B-cell lymphoma (DLBCL), chronic lymphocytic leukemia (CLL), follicular lymphoma (FL), and marginal zone lymphoma (MZL) using Mendelian randomization (MR) analysis. Methods: Genome-wide association study data from the InterLymph Consortium were available for 2,661 DLBCLs, 2,179 CLLs, 2,142 FLs, 824 MZLs, and 6,221 controls. SNPs associated (P &lt; 5 × 10−8) with high-density lipoprotein (HDL, n = 164), low-density lipoprotein (LDL, n = 137), total cholesterol (TC, n = 161), and triglycerides (TG, n = 123) were used as instrumental variables (IV), explaining 14.6%, 27.7%, 16.8%, and 12.8% of phenotypic variation, respectively. Associations between each lipid trait and NHL subtype were calculated using the MR inverse variance–weighted method, estimating odds ratios (OR) per standard deviation and 95% confidence intervals (CI). Results: HDL was positively associated with DLBCL (OR = 1.14; 95% CI, 1.00–1.30) and MZL (OR = 1.09; 95% CI, 1.01–1.18), while TG was inversely associated with MZL risk (OR = 0.90; 95% CI, 0.83–0.99), all at nominal significance (P &lt; 0.05). A positive trend was observed for HDL with FL risk (OR = 1.08; 95% CI, 0.99–1.19; P = 0.087). No associations were noteworthy after adjusting for multiple testing. Conclusions: We did not find evidence of a clear or strong association of these lipid traits with the most common NHL subtypes. While these IVs have been previously linked to other cancers, our findings do not support any causal associations with these NHL subtypes. Impact: Our results suggest that prior reported inverse associations of lipid traits are not likely to be causal and could represent reverse causality or confounding.

Abstract Abstract 4876 Background Recent studies have found obesity to be associated with a 1.5- to 2-fold elevated risk of developing multiple myeloma. This is of particular interest given that elevated levels of the pro-inflammatory cytokine interleukin (IL)-6 have been found in obese persons, and, at the same time, IL-6 has well-known proliferative and anti-apoptotic effects on monoclonal plasma-cells. Also insulin-like growth factors (IGFs) have been proposed to play a role since obesity often causes insulin resistance, which in turn modulates the bioavailability of IGF-1 Similar to IL-6, prior studies have found IGF-1 to have both growth and survival effects on monoclonal plasma-cells. Based on these facts, we have speculated that obesity might increase the risk of the multiple myeloma precursor monoclonal gammopathy of undetermined significance (MGUS), or, alternatively, that obesity may increase the risk for transformation from MGUS to multiple myeloma. We conducted the first large screening study designed to assess the association between obesity and MGUS among almost 2,000 African-American and Caucasian women. Methods We included 1000 African-American and 996 Caucasian women (age 40-79, median 48 years) from the Southern Community Cohort Study to assess MGUS risk in relation to obesity. Per our sampling strategy, about 50% of the participants were obese. Medical record-abstracted weight and height (measured on the day of study enrollment) and self-reported values had very high concordance (Pearson correlation >0.95). Serum samples from all subjects were analyzed by electrophoresis performed on agarose gel; samples with a discrete or localized band were subjected to immunofixation. Using logistic regression models, we estimated odds ratios (ORs) as measures of risk. Results Among all study participants, 39 (3.9%) African-Americans and 21 (2.1%) Caucasians were found to have MGUS, yielding a 1.9-fold (95%CI 1.1-2.3; p=0.021) higher risk of MGUS among African-Americans (vs. Caucasians). On multivariate analysis, we found obesity (OR=1.8, 95%CI 1.03-3.1; p=0.039), African-American race (OR=1.8, 95%CI 1.04-3.1; p=0.037), and increasing age (quartiles: ≥55 vs. <43 years) (OR=2.5, 95%CI 1.1-5.7; p=0.028) to be independently associated with an excess risk of MGUS. Another interesting finding was that the distribution of the monoclonal immunoglobulin isotype usage among African-American and Caucasian women was significantly different (p=0.007). Their respective rates were: IgG in 79.5% and 71.3 %; IgA in 7.7% and 0%; IgM in 7.7% and 19%; biclonal in 5.1% and 4.7%; and triclonal in 0% and 4.7%. The distribution of serum light-chain types between the two races was also significantly different (P=0.003, chi-square test): kappa in 53.8% and 47.6%; lambda in 43.6% and 42.8%; biclonal 2.6% and 4.7%; and triclonal in 0% and 4.7%. Conclusions Our finding that MGUS is twice as common among obese (vs. non-obese) women, and independent of race, supports the hypothesis that obesity is etiologically linked to myelomagenesis and may have public health impact. The observed 2-fold excess of MGUS among African-Americans (vs. Caucasians) of similar socio-economic status, coupled with other recent studies supports a role for susceptibility genes as the cause for racial disparity in the prevalence of MGUS. Disclosures No relevant conflicts of interest to declare.