Abstract A003 A zebrafish model of high-risk neuroblastoma exhibits multifocal primary disease
Authors: Woodward Kyle , Hamdad Farah , Ryu Borum , Tran Brian , Luna Perla , Anderson Nicole 
Published: September 2024

Abstract
Neuroblastoma (NB) is a highly metastatic tumor of the peripheral sympathetic nervous system (PSNS) and is responsible for 10% of childhood cancer deaths. Patients are designated as high-risk (HR) when MYCN-amplified or metastatic disease is present at greater than 18-months-of-age. MYCN-amplification remains one of the strongest prognostic factors of poor patient outcome and is detected in 20% of all NB patients. A GWAS study identified a SNP within LIM-domain-only 1 (LMO1) that is a permissive allele for neuroblastoma formation that results in increased LMO1 expression and metastasis. The most common site for primary disease in NB patients is the adrenal gland, but tumors can form along the paraspinal ganglion, thorax, pelvis, or cervical areas. Zebrafish models of NB use the dopamine-beta-hydroxylase (dbh) promoter to drive the expression of MYCN or c-MYC within the PSNS and result in primary tumor formation exclusively from the interrenal gland (adrenal gland). Our study utilizes MYCN_TT zebrafish which are an aggressive zebrafish model of HR-NB that highly expresses MYCN, have brightly EGFP-labeled tumor cells, and spontaneously metastasizes. To determine the impact of LMO1 overexpression on tumor formation in MYCN_TT zebrafish, we crossed MYCN_TT with dbh:LMO1 (LMO1) fish and we examined their offspring for tumorigenesis. At 9dpf hyperplasia was observed in of neuroblasts in arch-associated catecholaminergic (AAC) neurons and sympathetic cervical ganglion. The AAC is the carotid body in humans that is a bilateral sensory organ of the PSNS located at the common carotid artery and is neural crest in origin. Expansion of the AAC persisted in 60% of MYCN_TT;LMO1 fish at 14dpf and leads to the formation of secondary tumors. While primary tumors derived from the interrenal gland persisted in all MYCN_TT;LMO1 zebrafish, the secondary tumors from the AAC began to regress as early as 5wpf and continued to regress over time. At 15wpf only 20% MYCN_TT;LMO1 had a persisting AAC-derived secondary tumor. Strikingly, 10-15% of AAC-derived tumors go on to form massive tumors that invade nearby muscle and gill structures. Histological analysis of AAC-derived and IRG-derived tumors in MYCN_TT;LMO1 zebrafish demonstrated that both were composed of small, round, and undifferentiated neuroblasts. In addition, AAC-derive and IRG-derived tumors both stained positive for tyrosine hydroxylase a marker catecholamine synthesis in human NB. This study indicates that LMO1 overexpression is permissive of tumor formation outside of the IRG in the MYCN_TT zebrafish model. Despite identical genetic alterations in the MYCN_TT;LMO1 model, AAC-derived tumors have a wide range of fates from unstable tumors that regress to giant tumors, whereas all IRG-derived tumors persist overtime underscoring the influence of microenvironment. The MYCN_TT;LMO1 zebrafish are a novel model to study multifocal disease and the mechanisms that drive tumor stability in HR-NB.
Citation Format: Kyle Woodward, Farah Hamdad, Borum Ryu, Brian Tran, Perla Luna, Nicole Anderson. A zebrafish model of high-risk neuroblastoma exhibits multifocal primary disease [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pediatric Cancer Research; 2024 Sep 5-8; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl):Abstract nr A003.