Abstract Background: Cancer-associated fibroblasts (CAFs) are the major sources of secreted cytokines and chemokines contributing to the shaping of immunosuppressive tumor microenvironment (TME). Dysregulation of glycogen metabolism is integral to cancer cell proliferation and metastasis. The purpose of this study was to identify whether targeting glycogen metabolism in CAFs could alleviate immunosuppression and inhibit colorectal cancer (CRC) growth. Methods: Human CAFs (hCAFs) were isolated from CRC patient samples. APCf/f; CDX2-Cre/ERT2 mice were injected with tamoxifen to induce adenoma formation. Mouse CAFs (mCAFs) were isolated and cultured. The expression of glycogen phosphorylase liver form (PYGL), phospho-PYGL, hexokinase 2 (HK2), GLUT1, IL6, CLCF1, LIF, and CXCL6 was determined by either real time (RT)-PCR or western blot. Glycogen levels were determined using a Glycogen Analysis Kit. Cell proliferation was determined using WST-1. Extracellular acidification rate (ECAR) was determined utilizing an Agilent Seahorse XFe96 extracellular flux analyzer. To determine whether targeting PYGL would inhibit tumor growth, APCf/f; CDX2-Cre/ERT2 and MC38 mouse tumor models were used. Results: (i) Elevated glucose metabolism in CAFs was found compared to normal fibroblasts. (ii) Treatment of hCAFs with either conditioned medium (CM) derived from HCT116 CRC cells or TGFβ resulted in activated glycogen metabolism as noted by increased expression of p-PYGL/PYGL and the reduced level of glycogen in hCAFs. Moreover, CM increased the expression of IL6 family cytokines such as IL6, CLCF1, LIF and CXCL6 in hCAFs. (iii) Inhibition of glycogen metabolism by knockdown of PYGL or treatment with the PYG inhibitor, CP-91149, significantly repressed hCAFs proliferation and reduced the expression of these cytokines in hCAFs. Knockdown of PYGL decreased glycolytic activity as noted by decreased ECAR. (iv) Treatment with CP-91149 significantly inhibited tumor growth. In addition, mCAF cells co-implanted with MC38 cells increased MC38 tumor growth; this increase was attenuated by knockdown of PYGL in mCAFs. Conclusion: Our results demonstrate that glycogen metabolism is crucial for promoting CAF immunosuppressive functions. Importantly, our findings suggest that targeting glycogen metabolism in CAFs alleviates immunosuppression and inhibits CRC malignancy. Citation Format: Xudong Zhu, Yuning Zhou, Haoxiang Zhang, Yinping Jiang, Jinpeng Liu, Chi Wang, B. Mark Evers, Qingding Wang. Targeting glycogen metabolism in cancer-associated fibroblasts alleviates immunosuppression and inhibits colorectal cancer malignancy abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 4713.
Zhu et al. (Fri,) studied this question.