Abstract 3666: Co-occurring mutations of tumors suppressor genes, NF2 and LATS2, in malignant pleural mesothelioma
Authors: Tranchant Robin , Tallet Anne , Quetel Lisa , Renier Annie , Le Pimpec-Barthes Françoise , Zucman-Rossi Jessica , Jaurand Marie-Claude , Jean Didier 
Published: July 2016

Abstract
Background: Malignant Pleural Mesothelioma (MPM), mainly due to asbestos exposure, is an aggressive thoracic tumor resistant to conventional anti-cancer therapies. MPM patients have poor prognosis and a median survival around 12 months. These tumors are heterogeneous at the clinical and molecular level. To better characterize MPM heterogeneity, we recently identified a robust MPM transcriptomic classification defining two subgroups (C1 and C2). Epithelioid MPM, the most frequent histologic subtype, was found in both tumor subgroups, with a worse survival prognosis in the C2 subgroup. MPM heterogeneity is also found at the genetic level. Literature data showed alteration of Hippo signaling pathway in MPM linked to a frequent inactivation of NF2 tumor suppressor gene (30-40%). LATS2 tumor suppressor gene, another member of the Hippo pathway, was also shown to be altered but the frequency is not well determined.
Methods: Sixty-one primary cultures of MPM established in our laboratory were screened by Sanger sequencing for genes involved in mesothelial carcinogenesis, including CDKN2A, CDKN2B, BAP1, TP53, NF2 and LATS2. Transcriptomic data were obtained by Affymetrix microarray. Functional studies were performed using siRNAs to knockdown gene expression in three different MPM in culture. Gene expression was analyzed by quantitative RT-PCR and protein expression by Western blot. Cell proliferation was determined by MTS assay.
Results: Alteration frequencies of NF2 and LATS2 genes were 39% (24 cases) and 11% (7 cases), respectively. Five of seven LATS2 mutations were found associated with NF2 mutations in MPM of the C2 tumor subgroup. Patients with a co-occurring mutations in NF2 and LATS2 in tumor cells showed a worse prognosis (Logrank p-value of 0.02). These double mutants shared a similar transcriptomic profile. We identified MOK gene, a member of MAP kinase family, which is specifically deregulated in these double mutants (P&lt;0.01). Modeling NF2 and LATS2 co-inactivation by RNA interference resulted in a specific up-regulation of MOK expression (P&lt;0.01). Knockdown also induced a significant increase of MPM proliferation when the cells reach confluence (P&lt;0.01). In the MPM double mutants, MOK knockdown showed that MOK may be involved in the loss of contact inhibition.
Conclusions: In summary, we identified and characterized cell cultures of MPM tumors with co-occurring mutations of two members of the Hippo signaling pathway, NF2 and LATS2. Our findings demonstrate that NF2 and LATS2 inactivation leads to MOK overexpression and loss of contact inhibition. Novel target therapies are needed for MPM patients and MOK kinase could be a new potential therapeutic target.
Citation Format: Robin Tranchant, Anne Tallet, Lisa Quetel, Annie Renier, Françoise Le Pimpec-Barthes, Jessica Zucman-Rossi, Marie-Claude Jaurand, Didier Jean. Co-occurring mutations of tumors suppressor genes, NF2 and LATS2, in malignant pleural mesothelioma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3666.