Abstract
Simple SummaryAqueous humor (AH) liquid biopsy is an enriched source of cell-free circulating tumor-derived DNA for retinoblastoma (RB). The use of this AH liquid biopsy allows for genomic analysis of eyes in the absence of tumor tissue. Development of this platform was critical because direct tumor biopsy is prohibited in RB due to risk of extraocular tumor spread. In this retrospective study, we provide comprehensive, whole-genome analysis of the somatic copy number alterations (SCNAs) in 68 eyes of 64 RB patients. We show that the prevalence of specific SCNAs differ between eyes that required immediate enucleation (surgical removal) and eyes that were attempted to be saved but subsequently failed treatment, requiring secondary enucleation. Increases in chromosomal instability, or higher number of broad genomic alterations, predict higher risk clinical and biomarker features in these eyes. Prospective analyses are needed to further determine the clinical relevance and application of these findings.Aqueous humor (AH) liquid biopsy has been established as a surrogate tumor biopsy for retinoblastoma (RB). Previous AH studies have focused on highly recurrent RB somatic copy number alterations (SCNAs) including gain of 1q, 2p, 6p, and loss of 13q and 16q. In this retrospective study, we provide a comprehensive, whole-genome analysis of RB SCNAs and evaluate associated clinical features for 68 eyes of 64 RB patients from whom AH was obtained between December 2014 and October 2020. Shallow whole-genome sequencing of AH cell-free DNA was performed to assess for SCNAs. The prevalence of specific non-highly recurrent SCNAs, such as 20q gain and 8p loss, differed between primarily and secondarily enucleated eyes. Increases in chromosomal instability predict more advanced seeding morphology (p = 0.015); later age of diagnosis (p < 0.0001); greater odds of an endophytic tumor growth pattern (without retinal detachment; p = 0.047); tumor heights >10 mm (p = 0.09); and containing 6p gain, a biomarker of poor ocular prognosis (p = 0.004). The AH liquid biopsy platform is a high-yield method of whole-genome RB SCNA analysis, and SCNAs are associated with numerous clinical findings in RB eyes. Prospective analyses are encouraged to further elucidate the clinical relevance of specific SCNAs in RB.
Highlights
Retinoblastoma (RB), a cancer of the developing retina in infants and toddlers, is the most common pediatric intraocular malignancy [1] and accounts for 1% of childhood cancer mortality [2]
Demographics, clinical features, and somatic copy number alterations (SCNAs) findings of all participants are summarized in Figures S1 and S2
SCNAs in all eyes—including those that are actively undergoing therapy or even treatment-naïve [19,20,21,22,23,24,25,26]. The implementation of this organundergoing therapy or allows even treatment-naïve. Theinformation implementation this organspecific liquid biopsy for evaluation of tumoral from of less advanced specific liquid biopsy allows for evaluation of tumoral information from less advanced eyes, and importantly, with longitudinal sampling of the sameeyes eye, importantly, with longitudinal of under the same eye, the the potential detect and impactful new genomic alterationssampling that arise therapeutic potential to detect impactful new genomic alterations that arise under therapeutic pressure
Summary
Retinoblastoma (RB), a cancer of the developing retina in infants and toddlers, is the most common pediatric intraocular malignancy [1] and accounts for 1% of childhood cancer mortality [2]. In the vast majority of cases analyzed, biallelic inactivation of the tumor suppressor gene RB1 results in development of a premalignant retinoma [3,4,5,6,7], and additional mutational or epigenetic events may promote progression of a retinoma to fully malignant RB [3,8]. By activating oncogenes and inactivating tumor suppressor genes, somatic copy number alterations (SCNAs) are thought to contribute to subsequent RB progression [3,9,10,11,12,13]. Some tumors are negative for SCNAs or these genomic alterations are sub-clonal, suggesting a complicated series of heterogenous events for tumorigenesis in RB. As most studies have been done on tissue from enucleated eyes, we know very little about these events in less advanced eyes and what new alterations may form under selective therapeutic pressure during attempts to salvage the eye
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