Abstract PR-010: Dual targeting of IDO1/TDO2 inhibits tumor progression and attenuates the immune suppressive tumor microenvironment
Authors: Crump Lyndsey , Floyd Jessica , Woodruff Elizabeth , Bickerdike Mike , Kuo Li-Wei , Post Miriam , Itescu Silviu , Richer Jennifer , Bitler Benjamin G. 
Published: March 2024

Introduction: High-grade serous carcinoma (HGSC) tumors of the ovary, fallopian tube, or peritoneum are predicted to be immunogenic since the presence of tumor-infiltrating lymphocytes conveys a better prognosis. However, the efficacy of immunotherapies has been limited due to the immune-suppressed tumor immune microenvironment (TIME), which is established through tumor metabolism and immune-suppressive metabolites that directly affect immune cell function by depleting nutrients and activating immune-suppressive transcriptional programs. The tryptophan (TRP) catabolism pathway is a critical regulator of immune-suppressed TIME; however, clinical trials targeting the pathway to date have demonstrated limited response. Note there are two structurally distinct rate-limiting TRP catabolizing enzymes, Indoleamine 2,3-Dioxygenase (IDO1) and Tryptophan 2,3-Dioxygenase (TDO2), that evolved separately to catabolize tryptophan into Kynurenine (KYN). Most TRP catabolism inhibitors (e.g., Epacadostat) have primarily targeted IDO1, which may explain the mild anti-tumor effects of Epacadostat. Thus, we hypothesize that targeting both enzymes involved in TRP catabolism will alleviate the immune-suppressed TIME. Methods: We assessed primary human ascites fluid (IRB #07-0395) with global untargeted metabolomics. Using a panel of HGSC cell lines, we assessed genetic (small hairpin-mediated knockdown and overexpression) and pharmacologic (Epacadostat [IDO1 inhibitor], AT-0174 [IDO1/TDO2 inhibitor], 66cl4 [TDO2 inhibitor], StemRegenin 1 [AhR inhibitor]) modulation of IDO1, TDO2, and aryl hydrocarbon receptor (AhR, signal transducer of KYN). Proliferation assays in 2D and 3D. We examined the response of IDO1/TDO2 inhibition in two syngeneic murine models (ID8 Tp53-/- and BR-Luc Tp53-/-, Brca1-/-, myr-AKT, MYC). Multispectral immunohistochemistry (mIHC) and flow cytometry defined the TIME. Results: Elevated KYN levels are correlated to an inflamed TIME. The depletion of TRP and increased KYN negatively affect the anti-tumor immune response. HGSC cell lines and clinical outcomes depend more on TDO2 than IDO1. In contrast to knocking down IDO1 and AhR, TDO2 knockdown inhibited HGSC cell growth, and overexpression of TDO2 induced cancer cell proliferation. Targeting both IDO1 and TDO2 significantly inhibited TRP catabolism. Notably, only the combined IDO1/TDO2 inhibition resulted in the downregulation of PD-L1. In a syngeneic HGSC murine model, a first-in-class orally available dual IDO1/TDO2 inhibitor (AT-0174) significantly inhibited tumor progression, reduced tumor-associated macrophages, and reduced expression of immune-suppressive proteins, including PD-L1, on immune and tumor cells. Conclusions: These studies demonstrate the importance of TDO2 and the potential of AT-0174 to overcome an immune-suppressed TIME. The findings represent a novel approach to targeting TRP catabolism that may improve tumor response rates to immune therapy.

Citation Format: Lyndsey Crump, Jessica Floyd, Elizabeth Woodruff, Mike Bickerdike, Li-Wei Kuo, Miriam Post, Silviu Itescu, Jennifer Richer, Benjamin G. Bitler. Dual targeting of IDO1/TDO2 inhibits tumor progression and attenuates the immune suppressive tumor microenvironment [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr PR-010.