Parent inheritance of RB1 hypomorphic mutations and somatic mosaicism can explain low penetrance in retinoblastoma

Retinoblastoma is the most common eye cancer in children. Numerous families have been described displaying reduced penetrance and expressivity. An extensive molecular characterization of seven families led us to identify the two main mechanisms underlying these phenomena: i) mosaicism of amorphic mutations; and ii) parent-of-origin-effect of hypomorphic mutations. Somatic mosaicism for RB1 splicing mutations (c.1960+5G>C and c.2106+2T>C) leading to a complete loss of function was demonstrated by high-depth NGS in two families. In both cases, the healthy carrier parent (one with retinoma) showed a mutation frequency lower than that expected for a heterozygous individual, indicating a 56-60% mosaicism level. Parent-of-origin-effect is a mechanism by which the phenotypic impact of an allele depends on the parental origin. An imprinting mechanism justifying a ~3 fold excess of the RB1 maternal canonical transcript has been previously described. As a consequence, hypomorphic mutations, if maternally inherited, may retain sufficient suppressor activity to prevent tumor onset. In five low-penetrant families we identified paternally inherited hypomorphic mutations in the affected members, namely a deletion resulting in the loss of 37 aminoacids at the N-terminus (c.608-16_608del), an exonic substitution with a “leaky” splicing effect (c.1331A>G), a partially deleterious substitution (p.Arg661Trp) and a truncating C-terminal mutation (c.2663+2T>C). The identification of these mechanisms changes the genetic/prenatal counseling and the clinical management of families, raising to a 50% the recurrence risk for unaffected carriers, offspring of RB1-mutated mothers, and imposing the need for second tumor surveillance in unaffected carriers at risk of developing adult-onset cancer such as osteosarcoma or leiomyosarcoma.