Abstract
10591 Background: A large fraction of non-small cell lung cancers (NSCLC) have mutant KRAS, which predicts poor response to current cytotoxic therapy and portends a poor prognosis. Reactivation of senescence in KRAS mutant NSCLC may be an effective therapeutic strategy. We previously have shown that Twist1 cooperates with mutant KRAS to induce adenocarcinoma of the lung in mouse models and inhibition of Twist1 in these models leads to reactivation of KRAS induced senescence. In the current study, we explore whether TWIST1 plays a critical role in KRAS mutant human NSCLC. Methods: TWIST1 mRNA levels were examined through quantitative PCR. TWIST1, p53 and p21 were targeted with shRNAs. Reactivation of senescence was assessed through senescence associated beta-galactosidase staining (SA-β-Gal) and immunobloting (p21 and p27). Growth inhibition was determined through the crystal violet colony formation assay in vitro and xenografts experiments in vivo. Results: TWIST1 was overexpressed in 61% of the 164 human lung tumor samples examined. Silencing of TWIST1 in the KRAS mutant NSCLC cell line, H460, led to dramatic growth inhibition and induction of the senescence associated markers, p21, p27, and SA-β-Gal staining. Similar growth inhibition and induction of senescence were observed in six additional KRAS mutant NSCLC cell lines. Interestingly, growth inhibition by silencing of TWIST1 was independent of p53 or p16 mutational status. Silencing of p53 in two p53 wildtype NSCLC cell lines could not rescue loss of TWIST1 expression. Similar results were seen for inhibition of p21. To extend these observations in vivo, A549 cells containing a doxycycline-inducible shRNA against TWIST1 were implanted in NOD-SCID mice. Significant growth inhibition of established xenograft tumors was observed upon the addition of doxycycline. Conclusions: We have demonstrated TWIST1 is frequently overexpressed in human lung tumors. Furthermore, inhibition of TWIST1 in KRAS mutant lung cancer leads to reactivation of OIS and dramatic growth inhibition independent of the presence of p53 or p16. Together these findings suggest that inhibition of TWIST1 in KRAS mutant NSCLC represents a novel and exciting therapeutic strategy.
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