Germline pathogenic variants in neuroblastoma patients are enriched in BARD1 and predict worse survival

Abstract

Background: Neuroblastoma is an embryonal cancer of the developing sympathetic nervous system. The genetic contribution of rare pathogenic or likely pathogenic germline variants in patients without a family history remains unclear.

Methods: Germline DNA sequencing was performed on 786 neuroblastoma patients. The frequency of rare cancer predisposition gene pathogenic or likely pathogenic variants in patients was compared with 2 cancer-free control cohorts. Matched tumor DNA sequencing was evaluated for second hits, and germline DNA array data from 5585 neuroblastoma patients and 23 505 cancer-free control children were analyzed to identify rare germline copy number variants. Patients with germline pathogenic or likely pathogenic variants were compared with those without to test for association with clinical characteristics, tumor features, and survival.

Results: We observed 116 pathogenic or likely pathogenic variants involving 13.9% (109 of 786) of neuroblastoma patients, representing a statistically significant excess burden compared with cancer-free participants (odds ratio [OR] = 1.60, 95% confidence interval [CI] = 1.27 to 2.00). BARD1 harbored the most statistically significant enrichment of pathogenic or likely pathogenic variants (OR = 32.30, 95% CI = 6.44 to 310.35). Rare germline copy number variants disrupting BARD1 were identified in patients but absent in cancer-free participants (OR = 29.47, 95% CI = 1.52 to 570.70). Patients harboring a germline pathogenic or likely pathogenic variant had a worse overall survival compared with those without (P = 8.6 x 10-3).

Conclusions: BARD1 is an important neuroblastoma predisposition gene harboring both common and rare germline pathogenic or likely pathogenic variations. The presence of any germline pathogenic or likely pathogenic variant in a cancer predisposition gene was independently predictive of worse overall survival. As centers move toward paired tumor-normal sequencing at diagnosis, efforts should be made to centralize data and provide an infrastructure to support cooperative longitudinal prospective studies of germline pathogenic variation.

Figure 1.

Germline pathogenic or likely pathogenic variants in cancer predisposition genes across 786 neuroblastoma patients. Oncoprint of known cancer predisposition genes harboring rare germline variants classified as pathogenic or likely pathogenic. Patients without pathogenic or likely pathogenic variants in these genes are not shown. Patients are ordered by Children’s Oncology Group risk group and annotated with clinical and tumor biologic features. Genes are color-coded according to mode of inheritance, when known. Bar chart to the right indicates the number of variants detected for each gene and whether pathogenicity was determined based on ClinVar or our modified InterVar automated assessment. All variants were manually reviewed for quality and evidence of pathogenicity. INSS = International Neuroblastoma Staging System; UTR = Untranslated Region.

Figure 2.

Neuroblastoma patients harbor an excess burden of rare pathogenic or likely pathogenic (P-LP) variants in cancer predisposition genes. A) Overall excess burden of P-LP variants (single nucleotide variations and indels) in neuroblastoma (NBL) vs Penn Medicine BioBank (PMBB) and the Genome Aggregation Database (gnomAD) v2.1 controls is shown for cancer predisposition genes and the subset of genes studied involved in DNA repair. B) Gene-based rare variant burden test results comparing the number of neuroblastoma subjects with P-LP variants to those detected in gnomAD v2.1 and PMBB. C) Lollipop figure depicting 8 germline P-LP variants in BARD1. D) Rare P-LP variants in BARD1 are observed predominantly in patients diagnosed with high-risk neuroblastoma. CI = confidence interval; COG = Children’s Oncology Group; CPG = cancer predisposition gene.

Figure 3.

Rare germline copy number variants disrupting BARD1 in neuroblastoma patients. A) BARD1 deletions were identified in 3 of 5585 neuroblastoma patients through copy number analysis of a large germline single nucleotide polymorphism (SNP) array dataset (medium blue, top track). No deletions were observed in 23 505 array-matched cancer-free control participants. The thick and thin bars represent minimum and maximum deletion coordinates, respectively. One deletion was validated and fine-mapped by whole genome sequencing (WGS; light blue, middle track). No BARD1 protein coding deletions were observed in 10 847 individuals in the gnomAD v2.1 structural variant dataset (dark blue, bottom track). B-D) The 3 array-based copy number variant calls are shown in log R ratio (LRR) and B allele frequency (BAF) plots. Darker shading indicates the minimum deleted region, whereas lighter shading indicates the maximum region. E) WGS validation for patient PALXTB is shown as relative sequencing coverage for matched blood and tumor samples. F) Rare BARD1 deletion copy number variants are enriched in neuroblastoma compared to cancer-free participants. G) Deletions disrupting BARD1 were observed exclusively in patients diagnosed with high-risk subset of neuroblastoma. gnomAD = Genome Aggregation Database.

Figure 4.

Neuroblastoma patients harboring germline pathogenic or likely pathogenic (P-LP) variants in cancer predisposition genes have worse overall survival. Kaplan–Meier plots of overall survival probability in neuroblastoma patients with and without P-LP variants cancer predisposition genes. A) All patients. B) Restricted to low- and intermediate-risk (non-high-risk) groups. C) Restricted to high-risk group. Statistical significance in panels A-C assessed by log-rank test (P < .05). D) Forest plot of hazard ratios from Cox proportional hazards model. amp = amplified; Dx = diagnosis.