Abstract B25: Anti-Mullerian hormone (AMH) supports epithelial identity and survival signaling in lung cancer

Abstract

Abstract The purpose of this study is to characterize resistance mechanisms of non-small cell lung cancer (NSCLC) to heat shock protein 90 (HSP90) inhibition. Unexpectedly, we identified anti-Mullerian hormone (AMH) and its type II receptor (AMHR2) as two resistance associated genes. AMH and AMHR2 are TGF-ß/BMP superfamily members that share common type I receptors with BMP, resulting in overlapping signaling outputs. AMH and AMHR2 have thus far predominantly been studied in the context of gonadal development, in regulation of the female reproductive cycle and in gynecological malignancies, and AMH is not known to be expressed outside of these tissues. Using a focused RNAi library designed to detect genes associated with resistance to the HSP90 inhibitor ganetespib, siRNAs against AMH and AMHR2 sensitized 4 out of 5 and 3 out of 5 NSCLC cell lines to ganetespib, respectively. We then for the first time confirmed expression of AMH and AMHR2 in this non-gonadal tumor environment. Furthermore, analysis of data from The Cancer Genome Atlas (TCGA) indicated that AMH and AMHR2 are significantly upregulated in a subpopulation of NSCLC. Strikingly, depletion of AMH/AMHR2 induced EMT-like features, including downregulation of cadherins, a mesenchymal morphology, increased invasion, and expression of mesenchymal markers: features that are generally associated with resistance to chemotherapy. In contrast, inhibition of HSP90 selected for a more epithelial-like population of cells, as evident by increased E-cadherin or P-cadherin, and downregulation of mesenchymal markers such as VIM and ZEB1. To confirm that mesenchymal-like cells are indeed more responsive to HSP90 inhibition, we depleted E-cadherin or P-cadherin and again observed sensitization to ganetespib. We further found that AMH and AMHR2 did not confer sensitization to cisplatin. Interestingly, depletion of cadherins, and the associated induction of EMT-like properties, also resulted in downregulation of AMH and AMHR2, further supporting the link between AMH/AMHR2 expression and epithelial identity. In vivo studies confirmed that AMH depletion sensitizes NSCLC cells to ganetespib and showed that it also significantly reduces overall tumor volume. Reduction of AMH or AMHR2 also decreased phosphorylation of direct effector SMAD proteins, and depressed activity of NFκB and AKT, both regulators of EMT and survival. These results for the first time indicate the presence of an AMH-AMHR2-NFκB-AKT signaling axis of therapeutic relevance in NSCLC. To further explore the therapeutic potential of this observation, we treated NSCLC with combinations of ganetespib and the proteasome inhibitor bortezomib- an inhibitor previously shown to disrupt NFκB signaling in some cancers - and observed significant synergy between the two drugs. In conclusion, our work for the first time characterizes AMH and AMHR2 in NSCLC. We go on to show that AMH and AMHR2 regulated canonical-SMAD signaling as well as non-canonical AKT-NFκB signaling, in addition to regulating the epithelial identify of cancer cells. Importantly, our results also suggest that AMH and AMHR2 may serve as biomarkers to predict resistance to HSP90 inhibitors, and that it may be beneficial to prime lung tumors with HSP90 inhibitors, to reverse EMT and decrease survival signaling, prior to treatment with chemotherapy. Note: This abstract was not presented at the conference. Citation Format: Tim N. Beck, Vladislav Korobeynikov, Alexander Kudinov, Rachel Georgeopoulos, Emmanuelle Nicolas, Margret B. Einarson, Yan Zhou, Yanis Boumber, David A. Proia, Ilya G. Serebriiskii, Erica A. Golemis. Anti-Mullerian hormone (AMH) supports epithelial identity and survival signaling in lung cancer. [abstract]. In: Proceedings of the Fourth AACR International Conference on Frontiers in Basic Cancer Research; 2015 Oct 23-26; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2016;76(3 Suppl):Abstract nr B25.