Novel genomic therapeutic for desmoplastic small round cell tumor based on RNA sensor targeting of EWSR1::WT1 gene fusion

Ethan Giannotta

Document Type

Article

Publication Date

2025

Keywords

JGM

Abstract

The initiating tumorigenic event for many cancers is a chromosomal translocation that produces an oncogenic gene fusion. The development of RNA sensing technology can identify cancerous cells harboring these gene fusion transcripts and initiate the delivery of a downstream therapeutic payload. Here, we aim to build a novel therapeutic with ADAR-RNA sensors to target and kill cells containing the EWSR1::WT1 gene fusion found in desmoplastic small round cell tumor (DSRCT). We designed EWSR1::WT1 gene fusion transcripts containing the presence or absence of KTS isoform (+/-KTS) in exon 9 of WT1 and eight different therapeutic ADAR-RNA sensors with combinatorial alterations of base pair spacing between engineered stop codons (27bp or 42bp), linker type (E2A or XTEN80), and payloads (EGFP, TKSR39, or NTR1.1) and transfected these into HEK293T cells to test editing efficiency and cell viability with the presence or absence of a prodrug. One sensor in particular was identified to demonstrate the most promising and consistent patterns of editing efficiency and cell death when co-transfected with either the EWSR1::WT1 (+KTS) or EWSR1::WT1 (-KTS) fusion transcripts. New sensors should be designed to increase cell death before testing on the patient-derived JN-DSRCT-1 cell line that expresses the EWSR1::WT1 fusion transcripts.

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