Retinoblastoma (RB) is a childhood eye tumor, caused by RB1 mutation. Though diagnosing RB is easier, prognosticating RB is limited to examining the patient under anesthesia and imaging technique. The aim of the study is to find exosomal miRNA biomarkers to prognosticate RB. Exosomes were isolated from one control - MIO-M1 and two RB cell lines - WERI-Rb-1 and NCC-RbC-51. Small RNA sequencing was performed on exosomal miRNA isolated from the three cell lines. miRNAs specific to each cell line were shortlisted. A total of 243, 606 and 400 miRNAs were identified in MIO-M1, WERI-Rb-1 and NCC-RbC-51 cell lines respectively. Nine miRNAs were shortlisted based on adjusted p value and literature, MIO-M1 specific (n = 1), WERI-RB-1 specific (n = 2), NCC-RbC-51 specific (n = 2) and miRNAs common to both RB cell lines (n = 4) were chosen. Validation was done using specific Taqman miRNA assays.miRNA validation was carried out on cell lines, cell line derived exosomes, primary RB tissues and exosomes isolated from serum of the RB patients. Validation of the miRNAs in cell lines and exosomes derived from the cell lines, confirmed the sequencing data. However, only 2 miRNAs - hsa-miR-301b-3p and hsa-miR-216b-5p were upregulated in the primary RB tissues. None of the miRNAs had significant expression in the serum exosomes of RB patients. Therefore, serum exosomal miRNA may not be ideal for prognosticating RB.Further research on other body fluids like CSF and vitreous could serve as potential source for biomarkers for prognosticating RB.
Keywords: Exosomal miRNA; Patient serum; Retinoblastoma; Small RNA sequencing; Taqman probes.
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