Abstract 4900: An ALK fusion gene regulates different signaling pathways in mortal versus immortalized normal human cells for cellular senescence and transformation

Abstract

Abstract Accumulating results of clinical trials lead targeted therapies to be the first choice for unresectable or recurrent lung cancer with driver mutations. Echinoderm Microtubule Associated Protein Like 4 (EML4) - Anaplastic lymphoma kinase (ALK) fusion is known as such a driver mutation. It presents in 3-6% of non-small cell lung carcinoma (NSCLC). EML4-ALK fusion protein generate the constitutive ALK kinase activity in NSCLC. The basic understanding of EML4-ALK remains insufficient due to the lack of functional studies using normal human cells. We investigated the role of EML4-ALK in mortal and immortalized normal human cells. The expression of EML4-ALK in normal, mortal human fibroblasts caused accumulated DNA damage, telomere shortening and the early induction of cellular senescence with senescence-associated beta-galactosidase activity and upregulation of p16INK4A and p21WAF1. In contrast, when EML4-ALK was expressed in telomerase reverse transcriptase (hTERT)-immortalized normal human fibroblasts and bronchial epithelial cells, the cells showed accelerated proliferation in vitro and anchorage-independent growth in soft agar, revealing its transformation activity. No chromosome aberrations, no mutations or loss of p53, nor impairment of the p16INK4A response was associated with this in vitro transformation, likely reflecting certain clinical features of EML4-ALK-positive NSCLC. In both mortal and immortalized cells, EML4-ALK induced the phosphorylation of STAT3, which is involved in both cellular senescence and transformation. Our data validate that EML4-ALK functions as an oncogene, although an additional oncogenic event(s) seems to be required for full tumorigenicity in vivo. This study also suggests that telomerase-mediated immortalization manifests the oncogenic activity of EML4-ALK, switching from its senescence-inducing activity. The RNA-seq analysis showed that the STAT3-induced cytokine/interferon signaling pathways were most significantly upregulated by EML4-ALK in mortal fibroblasts, consistent with accumulated DNA damage and senescence induction. The blood coagulation pathway activated by EML4-ALK in hTERT-immortalized cells may contribute to increased risk of disseminated intravenous coagulation in patients with EML4-ALK-positive cancer. These results suggest that EML4-ALK regulates the different signaling pathways in mortal versus immortalized normal human cells to induce the different cellular outcomes. Citation Format: Masaru Matsumoto, Akihiko Miyanaga, Jessica Beck, Izumi Horikawa, Mohammed Khan, Delphine Lissa, Masahiro Seike, Akihiko Gemma, Hiroyuki Mano, Curtis Harris. An ALK fusion gene regulates different signaling pathways in mortal versus immortalized normal human cells for cellular senescence and transformation [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4900.