Abstract 1790: Pyruvate carboxylase regulates tumor progression through central carbon metabolism and immunosuppression
Authors: Coleman Michael F. , Cotul Eylem Kulkoyluoglu , Pfeil Alexander J. , Ho Alyssa N. , Devericks Emily N. , Safdar Muhammad H. , Chen Hao , Attaar Numair , Kiesel Violet A. , Teegarden Dorothy , Hursting Stephen D. , Wendt Michael K. 
Published: March 2024

Abstract
The tumor microenvironment (TME) is characterized by metabolic reprogramming and immune evasion, essential features in tumor progression. Tumor metabolic dysregulation, particularly the high production of lactate, plays a pivotal role in suppressing antitumor immunity within the TME. Pyruvate carboxylase (PC), responsible for converting pyruvate to oxaloacetate, is a critical determinant for lung metastasis in triple-negative breast cancer (TNBC). Herein we test the hypothesis that PC-mediated regulation of central carbon metabolism controls TME immunosurrveilance. Wild-type C57BL6/J and NSG mice received orthotopic injections of E0771 or M-Wnt tumor cells with shRNA targeting PC or a scramble control. RNA from tumors underwent transcriptomic profiling using Affymetrix Clariom D microarray (n=3-6/group per experiment). Gene set enrichment analysis identified pathways enriched in each condition, including a PC-associated hypoxia signature. Immunohistochemistry with anti-CD3 antibody and full spectrum flow cytometry assessed T cell infiltration and TME immune cell composition. Untargeted metabolomics, extracellular flux analysis, and high-resolution respirometry determined how loss of PC altered cellular metabolism. We found that suppressing PC expression increased primary tumor growth (~2 fold), potently drove immunosuppression, and remodeled cellular metabolism towards lactate production. Injection of PC-suppressed cells increased regulatory T cell levels in wild type mice. Importantly, PC-suppressed cells injected into NSG mice, which lack T, B, and NK cells, demonstrated slower, rather than accelerated, tumor growth. PC-suppression reduced oxygen consumption rate (OCR), but supply of anaplerotic carbon through exogenous malate or oxaloacetate restored OCR. Conversely, both the rate of and reliance on lactate production was increased by PC suppression. Notably, loss of immunosurveillance following downregulation of PC was reversed by inhibiting lactate transport. Additionally, lower PC expression was associated with increased hypoxia in tumors, supporting in vitro evidence of PC-repression by hypoxia. In summary, hypoxia-induced PC suppression emerges as a crucial mechanism through which primary tumors hinder antitumor immunity, partly via increased lactate production. These findings underscore the significance of PC-directed tumor metabolism as an understudied nexus in the interplay between tumor progression and antitumor immunity.
Citation Format: Michael F. Coleman, Eylem Kulkoyluoglu Cotul, Alexander J. Pfeil, Alyssa N. Ho, Emily N. Devericks, Muhammad H. Safdar, Hao Chen, Numair Attaar, Violet A. Kiesel, Dorothy Teegarden, Stephen D. Hursting, Michael K. Wendt. Pyruvate carboxylase regulates tumor progression through central carbon metabolism and immunosuppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1790.