Copy Number Alterations in Hepatoblastoma: Literature Review and a Brazilian Cohort Analysis Highlight New Biological Pathways

Abstract

Hepatoblastoma (HB) is a rare embryonal tumor, although it is the most common pediatric liver cancer. The aim of this study was to provide an accurate cytogenomic profile of this type of cancer, for which information in cancer databases is lacking. We performed an extensive literature review of cytogenetic studies on HBs disclosing that the most frequent copy number alterations (CNAs) are gains of 1q, 2/2q, 8/8q, and 20; and losses at 1p and 4q. Furthermore, the CNA profile of a Brazilian cohort of 26 HBs was obtained by array-CGH; the most recurrent CNAs were the same as shown in the literature review. Importantly, HBs from female patients, high-risk stratification tumors, tumors who developed in older patients (> 3 years at diagnosis) or from patients with metastasis and/or deceased carried a higher diversity of chromosomal alterations, specifically chromosomal losses at 1p, 4, 11q and 18q. In addition, we distinguished three major CNA profiles: no detectable CNA, few CNAs and tumors with complex genomes. Tumors with simpler genomes exhibited a significant association with the epithelial fetal subtype of HBs; in contrast, the complex genome group included three cases with epithelial embryonal histology, as well as the only HB with HCC features. A significant association of complex HB genomes was observed with older patients who developed high-risk tumors, metastasis, and deceased. Moreover, two patients with HBs exhibiting complex genomes were born with congenital anomalies. Together, these findings suggest that a high load of CNAs, mainly chromosomal losses, particularly losses at 1p and 18, increases the tendency to HB aggressiveness. Additionally, we identified six hot-spot chromosome regions most frequently affected in the entire group: 1q31.3q42.3, 2q23.3q37.3, and 20p13p11.1 gains, besides a 5,3 Mb amplification at 2q24.2q24.3, and losses at 1p36.33p35.1, 4p14 and 4q21.22q25. An in-silico analysis using the genes mapped to these six regions revealed several enriched biological pathways such as ERK Signaling, MicroRNAs in Cancer, and the PI3K-Akt Signaling, in addition to the WNT Signaling pathway; further investigation is required to evaluate if disturbances of these pathways can contribute to HB tumorigenesis. The analyzed gene set was found to be associated with neoplasms, abnormalities of metabolism/homeostasis and liver morphology, as well as abnormal embryonic development and cytokine secretion. In conclusion, we have provided a comprehensive characterization of the spectrum of chromosomal alterations reported in HBs and identified specific genomic regions recurrently altered in a Brazilian HB group, pointing to new biological pathways, and relevant clinical associations.

Keywords: array-CGH; copy number alteration; cytogenomics; hepatoblastoma; pediatric cancer.

Figure 1

Copy number alterations profile of the Brazilian hepatoblastoma cohort investigated by chromosomal microarray analysis using aCGH. Ideograms of chromosomes are depicted with copy number gains represented in blue bars at right, and losses in red at left; the width of the bars indicates the frequency of the alteration in the entire group. Chromosome Y was not evaluated. Image generated using the software Nexus Copy Number 9.0 (BioDiscovery).

Figure 2

Illustrative CNA profiles representing the three categories of tumors identified by aCGH in the Brazilian HB cohort. No detectable CNA (HB42T), few alterations (HB43T) and complex genomes (HB70T). Microarray probes are plotted according to their genomic coordinates, each color representing a chromosome, from 1 to 22, X, from the short arm to the long arm. The Y axis represents the log₂ scale of the copy number ratio tumor/control (values close to 0 indicate regions with similar copy number between tumor and control samples; positive values represent gains; negative values represent losses).

Figure 3

Comparison of CNA profiles of specific clinical features. The X axis displays chromosome profiles from 1 to 22, and X; chromosome Y was not evaluated. Copy number gains are represented in blue bars and losses in red; the width of the bars indicates the frequency of the alteration in the group. (A) Tumors with complex genomes versus samples with few CNAs. (B) Tumors from females versus tumors from males. (C) Tumors from patients who deceased versus tumors from alive patients. (D) High-risk versus low-risk tumors. (E) Tumors from patients diagnosed > 3 years-old versus samples from patients diagnosed < 3 years-old.

Figure 4

2q24.2q24.3 amplification detected in two HBs. Microarray probes are plotted according to their genomic coordinates of the affected segment on chromosome 2, from the short arm to the long arm. The Y axis represents the log₂ scale of the copy number ratio tumor/control (values close to 0 indicate regions with similar copy number between tumor and control samples, and positive values represent gains). Note the amplification regions as the highest peaks in the graphs.

Figure 5

Literature review of the cytogenomic data on hepatoblastomas. (A) Summary of the reported hepatoblastoma copy number chromosomal alterations in the literature from 1985 to 2017: chromosomes 1 to 12. (B) Summary of the reported hepatoblastoma copy number chromosomal alterations in the literature from 1985 to 2017: chromosomes 13 to 22, X, Y, and dmin. All references were these data were based can be found in the Supplementary Table 5.

Figure 6

Copy number gains and losses distributed per chromosome in 365 hepatoblastomas, based on our review of literature and the present study. Columns above the X axis represent the total number of gains per chromosome; below the X axis, the total number of losses. The frequencies of the most commonly chromosomal alterations detected in HBs in the literature along with our data are presented as percentages.