Ewing Sarcoma (ES) is the second most common bone cancer in children and adolescents with a high metastatic potential. Tumor development is driven by the specific t(11;22)(q24;q12) chromosomal translocation resulting in generation of the chimeric transcription factor EWS-FLI1.
Recently, ES has been reported to be exquisitely sensitive to inhibitors of poly(ADP-ribose) polymerase 1 (PARP1). This enzyme uses NAD+ as substrate and was demonstrated to regulate EWS-FLI1 in a feed-back mechanism. Another major mammalian NAD+ consumer is the deacetylase SIRT1 which we observed to be highly expressed in ES metastases, validated with immunohistochemistry of 250 primary tumors and 30 metastases. PARP1 and SIRT1 play pivotal roles in coupling cellular metabolism to transcriptional gene regulation as well as to stress response. Both of them regulate pro-apoptotic transcription factors that are suppressed in ES including p53, FOXO1 and 3, and NFκB. It has been demonstrated that PARP1 activity is at least partially dependent on acetylation which is counteracted by SIRT1 activity. Severe cell stress such as DNA damage leads to massive activation of PARP1, thus resulting in depletion of the cellular NAD+ pool and finally cell death. Usually, NAD+ is regenerated from nicotinamide via NAMPT or from the reduction of pyruvate via LDHA (Warburg effect), but can also be synthesized de novo from tryptophan. Interestingly, the knockdown of EWS-FLI1 in ES cells comes along with increased TDO and suppressed KMO expression, both being critical enzymes in tryptophan metabolism and NAD+ de novo synthesis. As a consequence, cellular tryptophan consumption drastically increases, which might indicate a regulatory function of EWS-FLI1 in maintaining the balance of the cellular NAD+ metabolism. In addition, we observed a striking sensitivity of ES cells to interferon gamma treatment at very low doses (5U/ml) resulting in the induction of the tryptophan metabolizing enzyme IDO and the production of the immune-suppressive metabolite kynurenine. We are currently studying how these changes in tryptophan metabolism affect cellular NAD+ pools and SIRT1 and PARP1 activities in ES. This study serves for a better understanding of EWS-FLI1 and of the microenvironment in the post-translational control of SIRT1 and PARP1 regulated gene transcription, and the consequences for PARP1 and/or SIRT1 directed therapies of ES.
Supported by the Austrian Science fund, grant I1225-B19.
Citation Format: Cornelia N. Mutz, Jozef Ban, Stephan Niedan, Maximilian O. Kauer, Dave N.T. Aryee, Dietmar Fuchs, Andreas Heitger, Heinrich Kovar. Investigating the NAD metabolome in Ewing sarcoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1436. doi:10.1158/1538-7445.AM2014-1436