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Genomic studies of pediatric cancer have primarily focused on specific tumor types or high-risk disease. Here, we used a three-platform sequencing approach, including whole-genome sequencing (WGS), whole-exome sequencing (WES), and RNA sequencing (RNA-seq), to examine tumor and germline genomes from 309 prospectively identified children with newly diagnosed (85%) or relapsed/refractory (15%) cancers, unselected for tumor type. Eighty-six percent of patients harbored diagnostic (53%), prognostic (57%), therapeutically relevant (25%), and/or cancer-predisposing (18%) variants. Inclusion of WGS enabled detection of activating gene fusions and enhancer hijacks (36% and 8% of tumors, respectively), small intragenic deletions (15% of tumors), and mutational signatures revealing of pathogenic variant effects. Evaluation of paired tumor–normal data revealed relevance to tumor development for 55% of pathogenic germline variants. This study demonstrates the power of a three-platform approach that incorporates WGS to interrogate and interpret the full range of genomic variants across newly diagnosed as well as relapsed/refractory pediatric cancers.Significance:Pediatric cancers are driven by diverse genomic lesions, and sequencing has proven useful in evaluating high-risk and relapsed/refractory cases. We show that combined WGS, WES, and RNA-seq of tumor and paired normal tissues enables identification and characterization of genetic drivers across the full spectrum of pediatric cancers.This article is highlighted in the In This Issue feature, p. 2945

Figure S1. Population Admixture Revealed by STRUCTURE Analysis Figure S2. Distribution of Polygenic Risk Score among Female Survivors of European Ancestry Figure S3. Cumulative Incidence of Subsequent Breast Cancer Differs by Polygenic Risk Score after Excluding Breast Cancer Predisposing Mutation Carriers Table S1. Previously Reported Loci Associated with Overall Breast Cancer Table S2. Polygenic Risk Score for 1133 Female Childhood Cancer Survivors of European Ancestry Table S3. Quintile Cutoffs for Polygenic Risk Score Based on 1133 Female Survivors or Estimated with Minor Allele Frequencies within OncoArray Control Population Table S4. Pathogenic or Likely Pathogenic Mutations in 11 Breast Cancer Predisposition Genes for 1133 Female Survivors of European Ancestry Table S5. Baseline Risk Model for Subsequent Breast Cancer Table S6. Association of Polygenic Risk Score with Rate of Subsequent Breast Cancer in Survivors of Childhood Cancer after Excluding Predisposing Mutation Carriers Table S7. Association of Polygenic Risk Score with Rate of Subsequent Breast Cancer in Survivors of Childhood Cancer after Excluding Predisposing Mutation Carriers (top 10% of PRS)

Purpose:The risk of subsequent breast cancer among female childhood cancer survivors is markedly elevated. We aimed to determine genetic contributions to this risk, focusing on polygenic determinants implicated in breast cancer susceptibility in the general population.Experimental Design:Whole-genome sequencing (30×) was performed on survivors in the St Jude Lifetime Cohort, and germline mutations in breast cancer predisposition genes were classified for pathogenicity. A polygenic risk score (PRS) was constructed for each survivor using 170 established common risk variants. Relative rate (RR) and 95% confidence interval (95% CI) of subsequent breast cancer incidence were estimated using multivariable piecewise exponential regression.Results:The analysis included 1,133 female survivors of European ancestry (median age at last follow-up = 35.4 years; range, 8.4–67.4), of whom 47 were diagnosed with one or more subsequent breast cancers (median age at subsequent breast cancer = 40.3 years; range, 24.5–53.0). Adjusting for attained age, age at primary diagnosis, chest irradiation, doses of alkylating agents and anthracyclines, and genotype eigenvectors, RRs for survivors with PRS in the highest versus lowest quintiles were 2.7 (95% CI, 1.0–7.3), 3.0 (95% CI, 1.1–8.1), and 2.4 (95% CI, 0.1–81.1) for all survivors and survivors with and without chest irradiation, respectively. Similar associations were observed after excluding carriers of pathogenic/likely pathogenic mutations in breast cancer predisposition genes. Notably, the PRS was associated with the subsequent breast cancer rate under the age of 45 years (RR = 3.2; 95% CI, 1.2–8.3).Conclusions:Genetic profiles comprised of small-effect common variants and large-effect predisposing mutations can inform personalized breast cancer risk and surveillance/intervention in female childhood cancer survivors.

Patients harboring germline pathogenic biallelic variants in genes involved in the recognition and repair of DNA damage are known to have a substantially increased cancer risk. Emerging evidence suggests that individuals harboring heterozygous variants in these same genes may also be at heightened, albeit lesser, risk for cancer. Herein, we sought to determine whether heterozygous variants in RECQL4, the gene encoding an essential DNA helicase that is defective in children with the autosomal recessive cancer-predisposing condition Rothmund–Thomson syndrome (RTS), are associated with increased risk for childhood cancer. To address this question, we interrogated germline sequence data from 4435 pediatric cancer patients at St. Jude Children's Research Hospital and 1127 from the National Cancer Institute Therapeutically Applicable Research to Generate Effective Treatment (TARGET) database and identified 24 (0.43%) who harbored loss-of-function (LOF) RECQL4 variants, including five of 249 (2.0%) with osteosarcoma (OS). These RECQL4 variants were significantly overrepresented in children with OS, the cancer most frequently observed in patients with RTS, as compared to 134,187 noncancer controls in the Genome Aggregation Database (gnomAD v2.1; P = 0.00087, odds ratio [OR] = 7.1, 95% CI, 2.9–17). Nine of the 24 (38%) individuals possessed the same c.1573delT (p.Cys525Alafs) variant located in the highly conserved DNA helicase domain, suggesting that disruption of this domain is central to oncogenesis. Altogether these data expand our understanding of the genetic factors predisposing to childhood cancer and reveal a novel association between heterozygous RECQL4 LOF variants and development of pediatric OS.

Pathogenic or likely pathogenic (P/LP) monoallelic germline mutations in BRCA2 increase risk of developing breast, ovarian, prostate, and pancreatic cancers. In a prior report from the SJLIFE study, BRCA2 was the third most frequently mutated gene (14 occurrences) among 3006 survivors of childhood cancer with the highest number observed among lymphoma survivors (7/586). To further investigate BRCA2 as a potential predisposition gene for pediatric/adolescent lymphoma, we analyzed 781 additional lymphoma survivors in the SJLIFE and CCSS cohorts with whole-genome sequencing (30X coverage). In the combined set of 1367 survivors (808 Hodgkin’s lymphoma [HL], 559 non-Hodgkin’s lymphoma [NHL]; 54% male; median age at diagnosis 12.6 [range 1.1-22.7] years), 13 P/LP mutations in BRCA2 were identified, with 7 mapped to the breast or ovarian cancer cluster regions defined by the Consortium of Investigators of Modifiers of BRCA1/2. Compared to reference controls in the Genome Aggregation Database (gnomAD) (Table 1), a significant association was observed between lymphoma and mutations in BRCA2 (odds ratio [OR], 3.1; 95% CI, 1.7-5.5) but not BRCA1. When stratified by diagnosis, the association was significant for NHL (OR, 4.8; 95% CI, 2.0-9.6) but not for HL. BRCA2 mutation carriers included a broad spectrum of NHL histological subtypes. Our findings support inclusion of pediatric/adolescent NHL in the spectrum of cancers associated with germline BRCA2 mutations. Approximately 1.4% of survivors of pediatric/adolescent NHL are carriers of a P/LP mutation in BRCA2, which may be the underlying etiology of their primary diagnosis. Clinically, counselling regarding BRCA2 mutation status should be considered for pediatric/adolescent NHL patients. Large scale genetic studies of newly diagnosed pediatric/adolescent lymphoma patients are warranted to replicate and refine diagnosis-specific risk estimates. Table 1.Comparisons of Mutation Carriers for BRCA1/2 Genes Between Lymphoma Survivors and gnomAD ControlsLymphoma SurvivorsgnomAD Controls (Hu et al. JAMA 2018)Cancer Risk (Fisher''s Exact Test)GeneCancer DiagnosisCarriersNon-CarriersCarriersNon-CarriersOdds Ratio (95% CI)P ValueBRCA2HL+NHL1313543131024263.1 (1.7-5.5)0.00045HL58033131024262.0 (0.7-4.8)0.11NHL85513131024264.8 (2.0-9.6)0.00041BRCA1HL+NHL313642081039141.1 (0.2-3.3)0.76HL18072081039140.6 (0.02-3.5)1.0NHL25572081039141.8 (0.2-6.6)0.31 Citation Format: Zhaoming Wang, Ti-Cheng Chang, Carmen L. Wilson, Chimene A. Kesserwan, Todd M. Gibson, Nan Li, John Easton, Heather L. Mulder, Gang Wu, Michael N. Edmonson, Michael C. Rusch, James R. Downing, Kim E. Nichols, Smita Bhatia, Gregory T. Armstrong, Melissa M. Hudson, Jinghui Zhang, Yutaka Yasui, Leslie L. Robison. Germline mutations in BRCA2 and pediatric/adolescent non-Hodgkin's lymphoma: A report from the St. Jude Lifetime (SJLIFE) and Childhood Cancer Survivor Study (CCSS) cohorts [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 4178.