Spectrum of gross deletions and insertions in the RB1 gene in patients with retinoblastoma and association with phenotypic expression

Quantitative multiplex PCR and genomic real-time PCR were used to complete an RB1 mutation analysis in 57 of 433 and 72 of 262 patients with hereditary and isolated unilateral retinoblastoma, respectively. These patients were selected because in previous analyses, which focused mainly on the identification of point mutations, no RB1 mutation was found. We identified gross deletions and insertions in peripheral blood DNA from 26 of 57 patients (46%) with hereditary retinoblastoma, and in six of 72 patients (8.3%) with isolated unilateral disease. In addition, we identified 32 somatic mutations in tumor DNA from 31 of 72 patients (43%) with isolated unilateral retinoblastoma. Together with our previous results, we found that gross RB1 alterations were present in the peripheral blood DNA from 65 of 433 (15%) and 17 of 262 (6.5%) patients with bilateral or familial and isolated unilateral retinoblastoma, respectively. Including reported gross deletions, an analysis of the frequency of breakpoints per intron length shows higher densities in introns 13, 16, 23, and 24. Genotype-phenotype analyses showed that on the whole, carriers of gross deletions develop fewer retinoblastomas compared to patients who are heterozygous for other types of RB1 null mutations. Specifically, carriers of cytogenetic and submicroscopic whole gene deletions often have unilateral tumors only. By contrast, almost all patients with gross deletions with one breakpoint in RB1 have bilateral retinoblastoma.