Ewing sarcoma (ES) is an aggressive bone and soft tissue tumor. We previously reported that the scaffolding protein menin is overexpressed by ES, and that menin inhibition results in impaired ES cell proliferation, survival, and tumorgenicity. Additionally, our recent studies revealed a previously undescribed role for menin in the activation of the serine biosynthetic pathway (SSP), a critical metabolic pathway that is aberrantly activated in many human cancers. The biologic functions of menin are largely determined by its protein-binding partners, the best characterized of which is the histone methyltransferase MLL. In the current study, we are investigating the mechanistic link between menin and the SSP to determine whether menin activates the SSP via epigenetic trithorax complexes.
Our data show that the core SSP enzymes, PHGDH, PSAT1, and PSPH are highly expressed by ES. Moreover, expression of all three genes is highly correlated in primary tumors, suggesting that they may be coordinately regulated by an upstream factor. In other cancer types, activation of the SSP has been shown to result from amplification of the PHGDH locus or activation of the master transcriptional regulator, ATF4. We have found that PHGDH is not amplified in ES but ATF4 is overexpressed, and ATF4 loss of function leads to growth inhibition. Q-RT-PCR and western blot of ES cells following ATF4 knockdown reveals down-regulation of PHGDH, PSAT1, and PSPH. In addition, ChIP-qPCR shows enrichment of ATF4 binding at SSP gene promoters, which is diminished by treatment with MI-503, a menin:MLL interaction inhibitor. Menin inhibition with MI-503 also leads to loss of ATF4 expression, coincident with loss of SSP expression. Additionally, ChIP-qPCR shows enrichment of menin binding at the ATF4 gene promoter, which is associated with H3K4me3 enrichment. Preliminary studies show that ATF4 over-expression via lentiviral transduction may rescue the loss of SSP gene expression induced by MI-503. Together these findings support the hypothesis that ATF4 acts downstream of menin to drive SSP activation in ES. Ongoing studies are assessing whether this is mediated by trithorax-dependent or –independent functions of menin.
Citation Format: Jennifer Jimenez, Laurie K. Svoboda, Sudha Sud, Samuel Kerk, Jolanta Grembecka, Costas A. Lyssiotis, Elizabeth R. Lawlor. Menin and ATF4 cooperate to drive serine biosynthesis in Ewing sarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4356.