Pikielny, Claudio W.

Background: Rare, functionally deleterious genetic variants have been implicated as having substantially larger effect sizes than common variants, potentially accounting for much of the missing heritability in lung cancer (LC). Over 25 years, the Genetic Epidemiology of LC Consortium (GELCC) has curated an invaluable resource of specimens and data from individuals with a strong family history of LC. Methods: Whole-genome and exome sequencing data were analyzed in 129 high-risk familial LC (FLC) families, defined as having two first-degree relatives or three or more second-degree relatives affected by LC (177 affected FLC and 309 unaffected relatives). The discovery focused on rare (allele frequency < 1% in gnomAD global population) and deleterious variations (missense, stop-gained, frameshift) enriched in FLC. Promising candidates were further external validated in 2,408 sporadic lung cancer (SLC) cases and 885 population controls from the International Lung Cancer Consortium. Results: We identified 168 variants with moderate-to-large effects on FLC risk, 100 of which were validated for SLC risk (Table 1 Table 1. Top 20 candidates Gene Rare (MAF < 1%), deleterious variants No. of variant carrier counts Odds Ratio, LC vs Control FLC | Relative | SLC | Control FLC+SLC SLC Oncogene ERBB3 / HER3 p.A1131T 3 | 1 | 2 | 0 35 15 JAK p.V651M 2 | 3 | 7 | 0 62 51 NOTCH3 p.R1560P 3 | 1 | 4 | 0 48 29 ROS1 p.N785X 6 | 4 | 10 | 0 110 74 USP6 p.R522 fs del 3 | 6 | 6 | 0 62 45 Tumor suppressor gene ATM p.L2332P 3 | 2 | 8 | 0 76 59 LZTS2 p.K458R 4 | 4 | 18 | 0 152 133 NPIPB13 p.V270 fs del 13 | 14 | 10 | 0 159 74 PARK2 / PRKN p.P153R 3 | 3 | 8 | 0 76 59 RB1 p.A525G 3 | 0 | 6 | 0 62 45 SLX4 p.S1716T 3 | 2 | 9 | 0 83 66 SYNE1 p.M6566I 2 | 0 | 7 | 0 62 52 TGFBRAP1 p.T301R 5 | 0 | 11 | 0 110 82 WNK1 p.K583 fs del 4 | 0 | 23 | 0 187 171 Other cancer-associated gene CFTR p.R74W 4 | 2 | 4 | 0 55 30 FLG p.R3404 fs del 7 | 2 | 4 | 0 69 30 LTN1 p.R349H 6 | 1 | 10 | 0 110 74 MLNR p.Q334 fs del 3 | 2 | 19 | 0 152 134 SLC39A11 p.R38Q 4 | 2 | 13 | 0 117 96 TTYH2 p.R493C 5 | 4 | 14 | 0 131 104 Dose effect of the top candidates No. candidates 0 variant allele 67 | 194 | 1911 | 834 reference reference 1 variant allele 50 | 66 | 363 | 50 3.5 (2.6 - 4.7) 3.2 (2.3 - 4.3) 2 variant alleles 15 | 18 | 62 | 1 32 (4.5 - 230) 27 (3.8 - 195) 3+ variant alleles 45 | 31 | 72 | 0 98 (6 - 1506) 62 (3.9 -1012) lists 20 top hits). Among these, 15 variants mapped to our previously identified lung cancer linkage locus at 6q23-25 (ROS1, PARK2, SYNE1). Others were enriched in oncogenes (ERBB3, JAK1, MET), tumor suppressor genes (ATM, BRCA2, RB1), and genes involved in the extracellular matrix (COL6A3, FLG, MUC5B). Importantly, individuals carrying more than two combinations of these variants exhibited exceptionally strong dose effects, with odds ratios (OR) exceeding 27. Conclusion: Our findings underscore the significant role of rare, high-penetrance variants in FLC etiology and highlight promising targets for early detection and personalized treatment. Future in-depth mechanistic studies are planned to evaluate the pathogenic effects of these specific alleles. Citation Format: Yanhong Liu, Claudio Pikielny, Xiangjun Xiao, Bo Peng, Yafang Li, Jinyoung Byun, Chao Cheng, Dakai Zhu, Spiridon Tsavachidis, Colette Gaba, Elena Kupert, Ellen L. Goode, Erin L. Crawford, Kristen Purrington, Marshall Anderson, Michael Cole, Paul Brennan, Geoffrey Liu, James McKay, John K. Field, Rayjean J. Hung, David C. Christiani, Diptasri Mandal, James C. Willey, Ann Schwartz, Joan Bailey-Wilson, Susan M. Pinney, Christopher I. Amos. High-penetrance rare variants underlying familial lung cancer risk: Insights from the Genetic Epidemiology of Lung Cancer Consortium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3815.

Abstract Although lung cancer is known to be caused by environmental factors, it has also been shown to have genetic components, and the genetic etiology of lung cancer remains understudied. We previously identified a lung cancer risk locus on 6q23-25 using microsatellite data in families with a history of lung cancer. To further elucidate that signal, we performed targeted sequencing on nine of our most strongly linked families. Two-point linkage analysis of the sequencing data revealed that the signal was heterogeneous and that different families likely had different risk variants. Three specific haplotypes were shared by some of the families: 6q25.3-26 in families 42 and 44, 6q25.2-25.3 in families 47 and 59, and 6q24.2-25.1 in families 30, 33, and 35. Region-based logarithm of the odds scores and expression data identified the likely candidate genes for each haplotype overlap: ARID1B at 6q25.3, MAP3K4 at 6q26, and UTRN (6q24.1) and PHACTR2 (6q24.2). Further annotation was used to zero in on potential risk variants in those genes. All four genes are good candidate genes for lung cancer risk, having been linked to either lung cancer specifically or other cancers. However, this is the first time any of these genes has been implicated in germline risk. Functional analysis of these four genes is planned for future work. Significance: This study identifies four genes associated with lung cancer risk, which could help guide future lung cancer prevention and treatment approaches.

Abstract Background: Lung cancer kills more people than any other cancer in the United States. In addition to environmental factors, lung cancer has genetic risk factors as well, though the genetic etiology is still not well understood. We have performed whole exome sequencing on 262 individuals from 28 extended families with a family history of lung cancer. Methods: Parametric genetic linkage analysis was performed on these samples using two distinct analyses—the lung cancer only (LCO) analysis, where only patients with lung cancer were coded as affected, and the all aggregated cancers (AAC) analysis, where other cancers seen in the pedigree were coded as affected. Results: The AAC analysis yielded a genome-wide significant result at rs61943670 in POLR3B at 12q23.3. POLR3B has been implicated somatically in lung cancer, but this germline finding is novel and is a significant expression quantitative trait locus in lung tissue. Interesting genome-wide suggestive haplotypes were also found within individual families, particularly near SSPO at 7p36.1 in one family and a large linked haplotype spanning 4q21.3-28.3 in a different family. The 4q haplotype contains potential causal rare variants in DSPP at 4q22.1 and PTPN13 at 4q21.3. Conclusions: Regions on 12q, 7p, and 4q are linked to increased cancer risk in highly aggregated lung cancer families, 12q across families and 7p and 4q within a single family. POLR3B, SSPO, DSPP, and PTPN13 are currently the best candidate genes. Impact: Functional work on these genes is planned for future studies and if confirmed would lead to potential biomarkers for risk in cancer.

Abstract Lung cancer (LC) is the primary cause of cancer-related deaths in the United States. Whereas smoking and other environmental factors strongly increase LC risk, multiple genetic variants also contribute to risk in smokers. Furthermore, among smokers, some families have been identified with an abnormally high prevalence of LC, suggesting that unknown genetic factors can greatly increase LC risk in smokers. The typically short survival after LC diagnosis impedes collection of detailed genotypic information on any single large family pedigree, impairing the identification of putative high-risk factors. Therefore, the Genetic Epidemiology of LC Consortium has collected epidemiological and genetic data from a number of families with high numbers of LC cases from eight different sites across the US. In this study, we have obtained whole exome sequences (WES) from 290 members of 28 families, including 66 LC cases. We used a gene-based approach to allow for the possibility that different families may contain different variants of the same gene. Variants were filtered for i) allele frequency, ii) functional effect using combined annotation-dependent depletion (CADD), and, iii) affecting a gene with either a known or suspected role in cancer. We further selected variants based on their segregation in family pedigrees in a pattern consistent with a large effect on LC risk. Candidate LC risk genes were then identified as those represented in at least two families by the same or different variants. We further culled the list of genes by requiring the presence of at least one rare, functional variant enriched in the WES of 1060 cases relative to 899 controls from the Transdisciplinary Research on Cancer of the Lung consortium. This analysis narrowed our results to two genes, one being E2A, a member of the E family of bHLH transcription factors. Whereas loss-of-function mutations in E2A drive lymphoid cancers, the E2A protein also participates in an oncogenic heterodimer with TWIST1 that promotes the epithelial-mesenchymal transition and is implicated in multiple cancer types. Furthermore, the E2A/TWIST1 heterodimer is the primary TWIST1-containing complex implicated in oncogenesis, and silencing of E2A in KRAS-mutant non-small cell lung cancer (NSCLC) cell lines results in oncogene-stimulated senescence and apoptosis. Our data identified three distinct E2A variants present in all 10 sequenced LC cases in the 5 families in which those variants are found. Finally, two of these E2A variants are located only 57 nucleotides from each other, immediately adjacent to sequences encoding a transcription activation domain, suggesting that both variants alter the same specific protein function. These data identify E2A variants as likely high risk factors for LC in smokers and validate our general approach for identifying genetic factors with a large impact on LC risk. Citation Format: Claudio W. Pikielny, Anthony M. Musolf, Mariza de Andrade, Diptasri Mandal, Colette Gaba, Ping Yang, yafang Li, Ming You, Richard Wilson, Elena Y. Kupert, Marshall W. Anderson, Ann G. Schwartz, Susan M. Pinney, Ambrose I. Granizo-Mackenzie, Yanhong Liu, Ramaswamy Govindan, James McKay, Rayjean Hung, John K. Field, David C. Christiani, Joan E. Bailey-Wilson, Christopher I. Amos. Familial studies identify variants in the E2A transcription factor as putative risk factors for lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-053.

Abstract Lung cancer (LC) kills more people in the United States each year than any other cancer. While it is well known that a variety of environmental factors (particularly tobacco smoke) strongly increase the risk of LC, there are multiple associated genetic variants with small contributions to risk. High aggregation of LC within rare individual families suggest that there are high-risk genetic variants as well. However, these genetic risk factors for LC are under studied due to the rapid fatality of LC. We studied 28 highly aggregated extended high-risk familial lung cancer (HRFLC) families collected from eight different sites across the US. Whole exome sequencing was performed on 290 individuals from these families to identify potential risk variants for HRFLC using genetic linkage analysis. Quality control was performed on the sequence data, filtering on parameters such as read depth, genotype quality, missingness, and Mendelian inconsistencies. Identity-by-descent (IBD) values were also calculated to verify correct familial relationships. Quality control procedures left approximately 400,000 SNVs and indels for analysis.Parametric two-point linkage analysis was performed assuming an autosomal dominant mode of inheritance. Disease allele frequency was set to 1% with a penetrance of 80% for carriers and 1% phenocopy rate. While we did not identify any genome-wide significant variants across the 28 families, multiple suggestive variants were identified. The largest cluster of suggestive variants was located at 14q32 in the CATSPERB gene. Given the likely locus heterogeneity in LC (combined with the lack of power in some families), it is not surprising that none of the variants were significant across the families; looking at the individual family results proved more informative. Long haplotypes linked to LC risk were identified in multiple families. These long runs of positive linkages, which have little to no negative linkage evidence across them, are characteristic of true linkage signals in these types of analysis. Two of the most interesting linked regions were at 7p36.1 and 4q21.23-28.23. The 7p signal (observed in a single family) was genome-wide suggestive and located within the SSPO gene. SSPO has been implicated in breast and skin cancer (melanoma); it is a novel lung cancer signal. The 4q linkage (again observed in a single family) covers a large chunk of 4q and contains multiple potential candidate genes, however, the best candidate gene is PTPN13, a gene implicated in lung cancer but never in familial lung cancer. We are currently evaluating the individual family results of several other pedigrees and plan to perform additional analyses to confirm these linkages. Citation Format: Anthony Musolf, Haiming Sun, Bilal A. Moiz, Claudio W. Pikielny, Mariza de Andrade, Diptasri Mandal, Colette Gaba, Ping Yang, Yafang Li, Ming You, Richard K. Wilson, Elena Y. Kupert, Marshall W. Anderson, Ann G. Schwartz, Susan M. Pinney, Christopher I. Amos, Ramaswamy Govindan, Joan E. Bailey-Wilson. Familial lung cancer exhibits multiple novel linked haplotypes within pedigrees [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4176.

Abstract Background: Recent genome-wide association studies (GWAS) have identified 28 susceptibility loci for Lung cancer (LC), 32 loci for smoking behavior, 10 loci for chronic obstructive pulmonary disease (COPD), 63 loci for pulmonary function and related phenotypes, totaling 107 GWAS susceptibility loci (as of November 2014). Given the common variants that have been found to be associated with LC in GWAS, targeted-region exome sequencing provides a cost-effective approach for further investigation of high-priority regions of the genome and has the great potential to identify rare causal variants in GWAS loci. Patients and Methods: Using an extreme phenotype approach, we selected 48 sporadic LC patients reporting heavy smoking histories with the presence of severe COPD in whom environmental factors are considered overwhelming, and 54 familial LC cases from families with at least three first-degree relatives with LC who are likely enriched for genomic signal, to search for the disease-causing rare mutations within the target 107 GWAS loci using exome sequencing data. Results: By focusing on exome profiles of these target 107 loci, we identified two disease-causing rare mutations on 6p21.32 TNXB p. Arg504His and 10q25.1 CCDC147 p.Arg696Cys. The homozygous Arg504His mutation presented in four familial and one sporadic LC cases. Notably, the minor allele frequency (MAF) of this variant in Caucasians from 1000 Genomes is very rare (<0.001). The heterozygous CCDC147 Arg696Cys variant were identified in two familial cases and one sporadic case. The MAF for this variant in the Caucasians from 1000 Genomes is 0.004. Both deleterious changes lead to truncation of the putative proteins, which disrupts the predicted domain and alters the cellular localization. We also observed several other suggestive rare mutations on G7C, TNS1, DBH1, and CHRNA5. Conclusion: Our target exome sequencing results demonstrate novel highly disruptive LC risk-conferring TNXB and CCDC147 mutations. Since the rare variants have appreciable effects on disease risk prediction, families and patients may warrant screening for mutations in TNXB and CCDC147 (if validated) to assess their potential LC risk. Citation Format: Yanhong Liu, Farrah Kheradmand, Michael Scheurer, Caleb Davis, David Wheeler, Edwin Silverman, Shete Sanjay, Spiridon Tsavachidis, Georgina Armstrong, Elena Kupert, Marshall Anderson, Yafang Li, Claudio Pikielny, Joan E. Bailey-Wilson, Ming You, Colette Gaba, Mariza DeAndrade, Diptasri Mandal, Claire Simpson, Susan Pinney, Christopher Amos, Margaret Spitz. Target exome sequencing for disease-causing rare mutations in familial and sporadic lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4600. doi:10.1158/1538-7445.AM2015-4600