Abstract 1482: Tissue specific splicing program of hMENA: impact on tumor immune microenvironment in node-negative NSCLC
Authors: Modugno Francesca Di , Spada Sheila , Carlo Anna Di , Trono Paola , Sperduti Isabella , Antoniani Barbara , Gallo Enzo , Campo Giulia , Facciolo Francesco , Visca Paolo , Nisticò Paola 
Published: August 2019

Deciphering the complexity of the tumor microenvironment (TME)is essential to unveil mechanisms of therapy resistance and develop novel microenvironment-related anti-tumor treatment. Actin cytoskeleton dynamics act as platforms for gene regulation and key signaling transduction pathways involved in the cross-talk among tumor cells and cellular and non-cellular components of TME.The actin regulatory protein hMENA undergoes tissue specific splicing, generating two alternatively expressed isoforms hMENA11a and hMENAΔv6 with a crucial role in EMT. We have previously demonstrated that hMENA11a and hMENAΔv6, respectively inhibit or increase cell invasiveness, TGFβ and β1 integrin signaling and the secretion of several key extracellular matrix (ECM) proteins. Early node-negative NSCLC patients show a prolonged disease-free survival (DFS) when expressing high tumor hMENA11a/low stromal FN1. Tertiary Lymphoid Structures (TLS), sites of transient lymphoid neo-genesis and determinants of antitumor immunity, have been associated with a favorable clinical outcome in NSCLC patientsandfound in responding lesions of ICB-treated melanoma patients.

The aim of the present study was to analyzethe pattern ofhMENA isoforms as biomarker of EMT signature in the context of ECM composition and TLS presence and localization.

We evaluated by gain and loss of function experiments the role of hMENA isoforms in TLS neogenesis. hMENA isoforms expression, TLS presence and stromal fibronectin were evaluated in 110primary tumors of node negative NSCLC patients by immunohistochemical analysis using pan-hMENA, hMENA11a, CD3, CD20 and fibronectin (FN) antibodies. The Chi-Square or Fisher Exact tests were used to estimate associations among categorical variables.

We found, by RNA-SEQ analysisand subsequent validation by QRT-PCR, in NSCLC cell lines depleted for the expression of ‘epithelial’ hMENA11a isoform,that hMENA11asustains the expression of lymphotoxin beta receptor (LTBR), a regulator of TLS formation.

The evaluation of TLS presence and spatial distribution in the primary tumors indicatedthat the presence of TLS within the tumor core is significantly correlatedwith hMENA11a expression in tumor cells, whereas the presence of TLS at the margin oftumor nests correlates with the absence of hMENA11a. When we evaluated also the fibronectin we found a trend of association between low stromal fibronectin and intratumoral TLS, however a low level of stromal FN in concomitance with the expression of hMENA11ain tumor cells,strongly associated with intra-tumoralTLS presence.

Our findings indicate that the alternative splicing of hMENA is crucial in the reciprocal signaling between tumor cells and their immune microenvironment, by participating in tertiary lymphoid structure neo genesis and spatial distribution.

Funded by Airc

Citation Format: Francesca Di Modugno, Sheila Spada, Anna Di Carlo, Paola Trono, Isabella Sperduti, Barbara Antoniani, Enzo Gallo, Giulia Campo, Francesco Facciolo, Paolo Visca, Paola Nisticò. Tissue specific splicing program of hMENA: impact on tumor immune microenvironment in node-negative NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1482.