Abstract 5189: TGFβ induces profound changes in the contribution and interaction of H157 NSCLC with its microenvironment
Authors: Rouzaut Ana , Salvo Elizabeth , Garasa Saray , Teijeira Alvaro 
Published: February 2011

Introduction:

Transforming growth factor TGFβ plays a dual role in tumorigenesis, acting as a tumor suppressor early in the process, but turning into tumor promoter in late-stage disease. Cancer metastasis is linked to the ability of the tumor cell to degrade its pre-existing extracellular millieu while assembling a tumor specific niche.

Objective:

To test the involvement of TGFβ in these processes, we analyzed the expression of 84 genes related to extracellular matrix proteins and adhesion molecules in H157 NSCLC cells exposed to TGFβ for 24 hours, with or without being exposed to specific inhibitor of TGFβII-receptor activation. We accompanied the expression data with functional assays that sustained what was observed at the mRNA level.

Methodology:

H157 NSCLC cells were exposed to TGFβ (2 ng/ml) for 24 hours. After that time, gene expression was profiled by means of the “ECM and adhesion molecules SABiosciences RT2 Profiler™ PCR Array”. Anchorage independent growth, cell migration towards growth factor gradients, adhesion to and transmigration through endothelial monolayers were performed in order to measure alterations in their adhesive properties. Integrin expression was measured by flow cytometry and their activation was measured by detecting FAK phosphorylation by western blot. To quantify if TGFβ altered the ECM degrading capability of NSCLC cells, gelatin zymography experiments were performed.

Results:

Our findings showed profound changes in H157 gene expression after TGFβ exposure: there are significant alterations in the expression of ECM molecules such as collagen, fibronectin, laminins and a prominent increase in the production of versican a protein that has been related to metastasis in several cancers. Besides, several MMPs (MMP 2, 3, 10, 11 and 14) and ADAMTS (1 and 13) increase their expression in TGFβ exposed H157 cells. We could also observe important increases in the expression of cell adhesion molecules such as the integrins α2, αv, β1 and β3. All of these results confirmed the different phenotype of NSCLC after TGFβ exposure. To provide experimental support for the transcriptional findings, we measured integrin expression and FAK activation in TGFβ exposed cells. Then, we used specific anti-αvβ3 integrin blocking antibodies to abolish the increases observed in H157 NSCLC cells migration, adhesion and transmigration through endothelial cells after TGFβ exposure. Finally, we measured matrix degrading activity and substrate independent growth of TGFβ treated cells in the presence or in the absence of TGFβ inhibitors.

Conclusions:

All the results aforementioned provide a detailed molecular profile of the pro-metastatic activity of TGFβ and support the use of TGFβ inhibitors to block it in lung cancer patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5189.