Unraveling the Genetic Architecture of Hepatoblastoma Risk: Birth Defects and Increased Burden of Germline Damaging Variants in Gastrointestinal/Renal Cancer Predisposition and DNA Repair Genes

Abstract

The ultrarare hepatoblastoma (HB) is the most common pediatric liver cancer. HB risk is related to a few rare syndromes, and the molecular bases remain elusive for most cases. We investigated the burden of rare damaging germline variants in 30 Brazilian patients with HB and the presence of additional clinical signs. A high frequency of prematurity (20%) and birth defects (37%), especially craniofacial (17%, including craniosynostosis) and kidney (7%) anomalies, was observed. Putative pathogenic or likely pathogenic monoallelic germline variants mapped to 10 cancer predisposition genes (CPGs: APC, CHEK2, DROSHA, ERCC5, FAH, MSH2, MUTYH, RPS19, TGFBR2 and VHL) were detected in 33% of the patients, only 40% of them with a family history of cancer. These findings showed a predominance of CPGs with a known link to gastrointestinal/colorectal and renal cancer risk. A remarkable feature was an enrichment of rare damaging variants affecting different classes of DNA repair genes, particularly those known as Fanconi anemia genes. Moreover, several potentially deleterious variants mapped to genes impacting liver functions were disclosed. To our knowledge, this is the largest assessment of rare germline variants in HB patients to date, contributing to elucidate the genetic architecture of HB risk.

Keywords: CYP1A1; DNA repair; ERCC5; MSH2; MUTYH; VHL; cancer predisposition; hepatoblastoma.

FIGURE 1

Workflow of the exome sequencing analysis. Variants were filtered according to quality (Phred score >20, read depth >10, variant allele frequency >35%); coding and noncoding variants were separately analyzed; frequency (GnomAD, ABraOM, 1K Genomes); the effect on coding variants (frameshift, stop loss/gain, missense, splice site, nonsense); for missense variants, the prediction of pathogenicity in at least five out of six algorithms; HPO annotation (hepatoblastoma, abnormalities of the liver, and cancer). The filtered variants were visually examined using Integrative Genomics Viewer (IGV) software (http//www.broadinstitute.org/igv) to further filter out possible strand bias and homopolymeric region artifacts. All the filtered variants mapped to cancer predisposition genes were classified using the ACMG guidelines. One- Intronic variants, 3′UTR, 5′UTR; two- MAF: GnomAD, ABraOM, 1K Genomes; three- Frameshift, stop loss, stop gain, missense, splice site, nonsense variants; four- Missense variants with dbNSFP Functional Prediction of pathogenicity in at least five out of six algorithms; five- Terms used for Varelect and HPO annotation: Hepatoblastoma, abnormalities of the liver, and cancer. CNV—copy number variation, ROH - region of homozygosity, MAF—maximum allele frequency, CPG—cancer predisposition gene, VUS—variant of uncertain significance, HPO—Human Phenotype Ontology, HB—hepatoblastoma.

FIGURE 2

Distribution of the detected high-quality rare coding and noncoding variants detected in 30 HB patients (A). Sequence ontology of the rare coding variants detected after selection by the read depth (>10), Phred score (>20), alternative allele frequency (>0.35), and population frequency (<1%). A total of 2,107 variants were classified into 1,671 missense mutations and 436 LoF variants (B). Sequence ontology of the rare noncoding variants detected after selection by the read depth (>10), Phred score (>20), alternative allele frequency (>0.35), and population frequency (<0.1%). A total of 2,070 noncoding variants were distributed in intronic, intergenic, and 3′ and 5′ UTRs.

FIGURE 3

APC pathogenic variant segregates with the cancer phenotype in the family of patient P27. The patient P27 is indicated with the black arrow. P27, her mother, sister, and niece are carriers of the pathogenic p. Cys1249* variant in the APC gene, which was identified by exome sequencing in the patient, and validated by Sanger sequencing in their indicated relatives. Her mother and sister had colon polyps and stomach polyps, respectively. The maternal grandmother and uncle had colon cancer (both are dead).

FIGURE 4

Frequency of high-quality rare germline coding variants mapped to DNA repair genes in HB patients and a control group. A list of 220 DNA repair genes distributed in 16 categories was analyzed; the 12 categories with variants detected in either patients or controls are represented. PARP—poly (ADP-ribose) polymerase. *p value 0.0338; Fisher’s test.