Abstract 6734: Identification of the novel TENM3-ALK fusion in an AYA case with ALK rearranged neuroblastoma
Authors: Hiwatari Mitsuteru , Seki Masafumi , Matsuno Ryosuke , Yoshida Kenichi , Nagasawa Takeshi , Sato-Otsubo Aiko , Yamamoto Shohei , Kato Motohiro , Watanabe Kentaro , Sekiguchi Masahiro , Miyano Satoru , Ogawa Seishi , Takita Junko 
Published: April 2023

Abstract
Neuroblastoma is the most common extracranial solid tumor that occurs in childhood and the most common cancer in infancy. Fewer than 5% of neuroblastomas occur in adolescents and young adults (AYAs), for whom the disease has an indolent and fatal course. Chen et al. previously reported that missense mutations in the protein- tyrosine kinase domain of anaplastic lymphoma kinase (ALK) occur in approximately 10% of cases of sporadic neuroblastoma, and these missense mutations result in the activation of the ALK protein kinase domain, which plays a key role in the tumorigenic process as seen in ALK fusion proteins. Since the initial discovery of the NPM-ALK fusion protein in human anaplastic lymphoma cell lines, more than 24 different ALK fusion proteins have been discovered in various malignancies. Till date, ALK-containing chimeric proteins have not been reported in neuroblastoma. In this study, a translocation within chromosome 2p and 4q was found to bring about the formation of an in-frame fusion gene that was composed of portions of the teneurin transmembrane protein 3 (TENM3) gene and ALK gene in tumor cells from an AYA patient with neuroblastoma. The patient was 19-year-old female diagnosed with stage 4 neuroblastoma (unfavorable histology, poorly differentiated subtype, MYCN DNA not amplified). Computed tomography revealed a mass in the left adrenal gland with bone, spine, liver, and lung metastasis. ALK rearrangement was confirmed via FISH analysis. Crizotinib resulted in no disease progression for 5 months, except for metastatic spinal disease, but it did not exhibit high efficacy compared with its effects against lymphoma and lung cancer. This 5′ in-frame fusion partner gene in neuroblastoma has not yet been identified, and its role is not well-known in oncogenesis. We generated the cell line with the expression of the full length TENM3-ALK cDNA in NIH-3T3 cells led to the formation of a fusion protein. A cell-based evaluation of this predicted ALK protein variant revealed activation of downstream targets-STAT3, AKT and ERK- and ALK inhibitors, crizotinib or lorlatinib, treatment inhibited the growth of xenograft tumors with stable TENM3-ALK expression. We determined whether TENM3-ALK affects transformation and proliferation using a soft agar colony formation assay. As expected, the cells that expressed an empty vector and wild type ALK line did not produce a considerable number of colonies, whereas TENM3-ALK transfected cells showed anchorage-independent growth, resulting in the production of a higher number of colonies. The evidence for the role of ALK receptor signaling in stimulating neuroblastoma tumor cell growth and the demonstrated in vitro and in vivo efficacy of the ALK selective inhibitors in this study provide biological and clinical justification for the further exploration of this combination and testing in patients with recurrent or refractory neuroblastoma.
Citation Format: Mitsuteru Hiwatari, Masafumi Seki, Ryosuke Matsuno, Kenichi Yoshida, Takeshi Nagasawa, Aiko Sato-Otsubo, Shohei Yamamoto, Motohiro Kato, Kentaro Watanabe, Masahiro Sekiguchi, Satoru Miyano, Seishi Ogawa, Junko Takita. Identification of the novel TENM3-ALK fusion in an AYA case with ALK rearranged neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6734.