Abstract 3404: Exploring the role of Twist1 in the pathogenesis of malignant pleural mesothelioma (MPM)

Abstract

Abstract Background: MPM is caused by lethal neoplastic growth of the pleura surrounding lungs. It is resistance to most standard anti-cancer treatment regimens and needs discovery of newer therapeutic approaches. MPM is characterized by massive loco-regional invasion of the malignant pleural cells into the lung parenchyma. Twist1 is a transcription factor, which promotes invasion and metastasis of tumor cells, increases chemotherapeutic resistance and is involved in the pathobiology of many cancers. Also recent studies have highlighted the potential of twist1 as a therapeutic target in cancer. But there is no report investigating its function in mesothelioma. Methods: We extracted total RNA from 53 frozen resected tumor tissue specimens, comprised of 39 epitheloid, 7 sarcomatoid and 7 biphasic histotypes, along with paired normal tissue. The RNA was labeled and hybridized to Affymetrix U133 plus 2.0 microarray to obtain transcriptomic profiles. Bioinformatic analysis of the microarray data using a 2 sample t-test was applied, on a probe-by-probe basis followed by Beta-uniform Mixture for multiple comparisons, to determine the differences between tumor vs normal specimens. The microarray results were validated by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) using Taqman assays on the ABI 7300 platform. For all qRT-PCR experiments the twist1 transcript levels were determined relative to endogenous GAPDH as control using ΔΔCT calculation. We performed Western blot analysis on a panel of 16 mesothelioma cell lines including Met-5A (SV-40 immortalized) and HCT-4012 (telomerase immortalized) pleural mesothelial control cell lines. Results: The bioinformatic analysis of microarray data showed that twist1 transcript level was 8.7 fold higher in tumors (p = 1.1E-16) compared to paired normal specimens. Using qRT-PCR, we compared twist1 transcript levels in 12 pairs of tumor vs paired normal tissue specimens and found that twist1 was upregulated to more than 10 fold in MPM tumors (p < E-4). Western blot showed that 10 MPM cancer cell lines had higher expression of twist1 protein compared to Met-5A and HCT-4012 cell lines. The highest expression was seen in 2 of the sarcomatoid cell lines - RS5 and DM3, suggesting a correlation with metastatic phenotype since sarcomatoid tumors are highly metastatic in nature. Conclusion: Our preliminary findings suggest that twist1 is upregulated in MPM tumors and cell lines and may play a role in the development of MPM. Further studies are needed to investigate its role in the process of tumorigenesis and metastasis. Supported by Grants: DoD W81XWH-07-1-0306 (I.I.W and AST), Fleming Foundation, IASLC Young Investigator Award 2011-2013 (MS). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3404. doi:1538-7445.AM2012-3404