High-Level Amplified Proficient Retinoblastoma Tumors Retain Distinct Molecular Signatures

Purpose: Retinoblastomas are malignant eye tumors diagnosed in young children. Most retinoblastomas are genetically characterized by biallelic inactivation of the gene. However, 1.5% of tumors demonstrate high-level amplification of the proto-oncogene . Patients with -amplified -proficient retinoblastoma receive a diagnosis at an earlier age and show a clinically and histologically more malignant phenotype. This study aimed to identify genome-wide molecular features that distinguish this subtype from other retinoblastomas.

Design: Cohort study.

Participants: Forty-seven retinoblastoma tumors, comprising 36 , 4 , and 7 tumors. In total, 5 retinoblastomas displayed high-level amplification, with 3 being , 1 being , and 1 being .

Methods: Integrated analysis, based on gene expression, methylation, and methylation-expression correlations, was performed to identify distinct molecular components of -amplified -proficient retinoblastomas compared with other retinoblastoma subtypes. The methylation and methylation-expression correlation analysis was initially conducted within a subset of samples (n = 15) for which methylation profiles were available. The significant findings were cross-validated in the entire cohort (n = 47) and in publicly available data.

Main Outcome Measures: Differentially expressed genes/pathways, differentially methylated genes, and methylation-driven differential gene expression.

Results: A large number of genes (n = 3155) were identified with distinct expression patterns in -amplified -proficient retinoblastomas. The upregulated and downregulated genes were associated with translation and cell-cycle processes, respectively. Methylation analysis revealed distinct methylated patterns in -amplified -proficient tumors, many of which showing significant impact on gene expression. Data integration identified a 40-gene expression signature with hypermethylated state resulting in a significant downregulation in -amplified -proficient retinoblastomas. Cross-validation using the entire cohort and the public domain expression data verified the overall lower expression of these genes not only in retinoblastomas with a -amplified -proficient background, but also in amplified neuroblastomas. These include the metabolism-associated gene and the cyclin-dependent kinase inhibitor gene .

Conclusions: -amplified -proficient retinoblastomas display significantly distinct molecular features compared with other retinoblastomas, including a set of 40 hypermethylation-driven downregulated genes. This gene set can give insight into the biology of -amplified retinoblastomas and may help us to understand the more aggressive clinical behavior.