Molecular analysis distinguishes metastatic disease from second cancers in patients with retinoblastoma

The pediatric ocular tumor retinoblastoma readily metastasizes, but these lesions can masquerade as histologically similar pediatric small round blue cell tumors. Since 98% of retinoblastomas have RB1 mutations and a characteristic genomic copy number “signature”, genetic analysis is an appealing adjunct to histopathology to distinguish retinoblastoma metastasis from second primary cancer in retinoblastoma patients. Here, we describe such an approach in two retinoblastoma cases. In patient one, allele-specific (AS)-PCR for a somatic nonsense mutation confirmed that a temple mass was metastatic retinoblastoma. In a second patient, a rib mass shared somatic copy number gains and losses with the primary tumor. For definitive diagnosis, however, an RB1 mutation was needed, but heterozygous promoter→exon 11 deletion was the only RB1 mutation detected in the primary tumor. We used a novel application of inverse PCR to identify the deletion breakpoint. Subsequently, AS-PCR designed for the breakpoint confirmed that the rib mass was metastatic retinoblastoma. These cases demonstrate that personalized molecular testing can confirm retinoblastoma metastases and rule out a second primary cancer, thereby helping to direct the clinical management.