Abstract 4338: The role of IMPACT in survival of cancer cells during tryptophan deprivation by immunosuppressive enzyme indoleamine 2,3-dioxygenase (IDO1)
Authors: Petr Tomek, Ariane Hallermayr, Michael Kilian, Cristin Gregor Print, Lai-Ming Ching
Journal: Cancer Research (2017)

Abstract A broad range of human malignancies overexpress an enzyme called indoleamine 2,3-dioxygenase 1 (IDO1) to suppress antitumor immunity which associates with poor patient prognosis. One mechanism whereby IDO1 activity suppresses the host’s immune cells is mediated by deprivation of the essential amino acid tryptophan (Trp) which raises an intriguing question. How do the cancer cells overcome low levels of the essential amino acid when the immune T-cells are triggered to self-destruct? A protein called IMPACT (product of IMPrinted gene with AnCienT domain) was reported to confer skin cells and neuronal cells increased survival in Trp-deprived environments. Based on these reports, we hypothesise that cancer cells hijack IMPACT to gain survival advantage in Trp-deprived environments. To obtain evidence for this hypothesis, we analysed publicly available genomics datasets and performed experimental assays. Murine GL261 glioblastoma cells overexpressing a full-length mouse IMPACT gene (GL261-IMPACT) were generated by lipofection. IMPACT overexpression was confirmed by Western blot. Metabolic activity and survival of the GL261 cells in Trp deprivation experiments were evaluated using MTT assay and staining with live/dead indicators fluorescein diacetate and propidium iodide, respectively. Meta-analysis of the publicly available RNA-sequencing datasets, The Cancer Genome Atlas (TCGA) and Genotype Tissue Expression (GTEx), provided evidence that IMPACT was overexpressed in the majority of the 24 analysed human cancers compared to their non-malignant tissue counterparts. Consistent with the high IMPACT expression in prostate cancer patients, 12-25% prostate tumors show amplification of the IMPACT gene (source: cBioPortal for Cancer Genomics). In addition, we found an association of high IMPACT expression with poorer survival for certain leukemia and lung cancer patients in the Gene Expression Omnibus (GEO) portal. To obtain experimental evidence that IMPACT protects cancer cells during Trp deprivation, GL261-IMPACT cells were cultured in media containing 2.5 - 50 μM Trp. Metabolic activity of GL261-IMPACT cells decreased at a slower rate than that of GL261-wild-type cells after 3 to 5 days in media containing limiting Trp concentration (2.5 - 10 μM). On day 5, GL261-wild-type and GL261-IMPACT cells cultivated in 10 μM Trp showed 4.5% and 23% metabolic activity (p = 2.8x10-8), respectively, relative to the same cells grown in 50 μM Trp. Fluorescence microscopy utilising live/dead staining showed that after 5 days of incubation at limiting Trp concentrations (5 - 10 μM), the majority of GL261-wild-type cells were non-viable whereas GL261-IMPACT cells were predominantly viable. Our initial results support the hypothesis that IMPACT aids cancer cells to overcome periods of tryptophan deprivation. Citation Format: Petr Tomek, Ariane Hallermayr, Michael Kilian, Cristin Gregor Print, Lai-Ming Ching. The role of IMPACT in survival of cancer cells during tryptophan deprivation by immunosuppressive enzyme indoleamine 2,3-dioxygenase (IDO1) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4338. doi:10.1158/1538-7445.AM2017-4338