Tumor-intrinsic CD47 signal regulates glycolysis and promotes colorectal cancer cell growth and metastasis.
肿瘤固有 CD47 信号调节糖酵解，促进结直肠癌细胞生长和转移。
- 作者列表："Hu T","Liu H","Liang Z","Wang F","Zhou C","Zheng X","Zhang Y","Song Y","Hu J","He X","Xiao J","King RJ","Wu X","Lan P
:Rationale: CD47 plays a vital role in the immune escape of tumor cells, but its role in regulating immune-unrelated biological processes such as proliferation and metastasis remains unclear. We seek to explore the immune-independent functions of CD47 in colorectal cancer (CRC). Methods: The expression of CD47 in CRC was determined by immunohistochemistry. The biological effect of CD47 signaling on tumor cell proliferation and metastasis was evaluated in vitro and in vivo. RNA sequencing analysis was performed to identify pivotal signaling pathways modulated by CD47. The interaction between CD47 and ENO1 was verified by co-immunoprecipitation (co-IP). The effect of CD47 on glycolytic metabolites was analyzed by seahorse XF and targeted metabolomics. Results: The expression of CD47 was upregulated and correlated to poor prognosis in CRC patients. Functional assays revealed that CD47 promoted CRC cell growth and metastasis in vitro and in vivo. Our mechanistic investigations demonstrated that CD47 interacted with ENO1 and protected it from ubiquitin-mediated degradation, subsequently promoting glycolytic activity and phosphorylation of ERK in CRC cells. Inhibition of ENO1 diminished CD47-mediated cell growth and migration. Clinically, the combined expression of CD47 and ENO1 provided reliable predictive biomarkers for the prognosis of CRC patients. Conclusions: CD47 is overexpressed in CRC, and its expression is associated with poor prognosis. Through stabilizing ENO1, CD47 enhances the aerobic glycolysis and ERK activity in CRC cells, thereby promoting the progression of CRC. Our studies reveal an unconventional role of CD47, suggesting that targeting the CD47-ENO1 axis may provide a novel therapeutic avenue for CRC.
: 理论基础: CD47 在肿瘤细胞的免疫逃逸中起着至关重要的作用，但其在调节增殖和转移等免疫无关的生物学过程中的作用仍不清楚。我们试图探索 CD47 在结直肠癌 (CRC) 中的免疫非依赖性功能。方法: 采用免疫组织化学方法检测 CD47 在结直肠癌中的表达。在体外和体内评价了 CD47 信号对肿瘤细胞增殖和转移的生物学作用。进行 RNA 测序分析，以确定 cd47 调控的关键信号通路。通过免疫共沉淀 (co-IP) 验证 CD47 与 ENO1 的相互作用。通过海马 XF 和靶向代谢组学分析 CD47 对糖酵解代谢产物的影响。结果: CD47 表达上调，与 CRC 患者的不良预后相关。功能检测发现 CD47 在体内外促进 CRC 细胞生长和转移。我们的机制研究证明，CD47 与 ENO1 相互作用，保护其免受泛素介导的降解，随后促进 CRC 细胞的糖酵解活性和 ERK 的磷酸化。抑制 ENO1 减少细胞生长和迁移 CD47-mediated。临床上，CD47 和 ENO1 的联合表达为 CRC 患者的预后提供了可靠的预测生物标志物。结论: CD47 在结直肠癌中呈高表达，其表达与预后不良有关。通过稳定 ENO1，CD47 增强 CRC 细胞的有氧糖酵解和 ERK 活性，从而促进 CRC 的进展。我们的研究揭示了 CD47 的非常规作用，提示靶向 CD47-ENO1 轴可能为 CRC 提供一种新的治疗途径。
METHODS::Macrophages (MΦ) play a critical role in tumor growth, immunosuppression and inhibition of adaptive immune responses in cancer. Hence, targeting signaling pathways in MΦs that promote tumor immunosuppression will provide therapeutic benefit. PI3Kγ has been recently established by our group and others as a novel immuno-oncology target. Herein, we report that a macrophage Syk-PI3K axis drives polarization of immunosuppressive MΦs which establish an immunosuppressive tumor microenvironment in in vivo syngeneic tumor models. Genetic or pharmacological inhibition of Syk and/or PI3Kγ in MΦs promotes a pro-inflammatory MΦphenotype, restores CD8+ T cell activity, destabilizes HIF under hypoxia, and stimulates an antitumor immune response. Assay for Transposase-accessible Chromatin using Sequencing (ATAC-seq) analyses on the bone marrow derived macrophages (BMDMs) show that inhibition of Syk kinase promotes activation and binding of NF-κB motif in SykMC-KO BMDMs, thus stimulating immunostimulatory transcriptional programming in MΦs to suppress tumor growth. Finally, we have developed in silico the "first in class" dual Syk/PI3K inhibitor, SRX3207, for the combinatorial inhibition of Syk and PI3K in one small molecule. This chemotype demonstrates efficacy in multiple tumor models and represents a novel combinatorial approach to activate antitumor immunity.
METHODS::Dysfunction of invariant natural killer T (iNKT) cells in tumor microenvironment hinders their anti-tumor efficacy, and the underlying mechanisms remain unclear. Here we report that iNKT cells increase lipid biosynthesis after activation, and that is promoted by PPARγ and PLZF synergically through enhancing transcription of Srebf1. Among those lipids, cholesterol is required for the optimal IFN-γ production from iNKT cells. Lactic acid in tumor microenvironment reduces expression of PPARγ in intratumoral iNKT cells and consequently diminishes their cholesterol synthesis and IFN-γ production. Importantly, PPARγ agonist pioglitazone, a thiazolidinedione drug for type 2 diabetes, successfully restores IFN-γ production in tumor-infiltrating iNKT cells from both human patients and mouse models. Combination of pioglitazone and alpha-galactosylceramide treatments significantly enhances iNKT cell-mediated anti-tumor immune responses and prolongs survival of tumor-bearing mice. Our studies provide a strategy to augment the anti-tumor efficacy of iNKT cell-based immunotherapies via promoting their lipid biosynthesis.
METHODS::Near-infrared photoimmunotherapy (NIR-PIT) is a newly developed and selective cancer treatment that induces necrotic and immunogenic cell death and utilizes a monoclonal antibody conjugated to a photo-absorber dye, IR700DX, activated by NIR light. Although CD44 is surface cancer marker associated with drug resistance, anti-CD44-IR700 NIR-PIT results in inhibited cell growth and prolonged survival in multiple tumor types. Meanwhile, anti-CD25-IR700-targeted NIR-PIT has been reported to achieve selective and local depletion of FOXP3+CD25+CD4+ regulatory T cells (Tregs), which are primary immunosuppressive cells in the tumor microenvironment (TME), resulting in activation of local antitumor immunity. Combined NIR-PIT with CD44- and CD25-targeted agents has the potential to directly eliminate tumor cells and also amplify the immune response by removing FOXP3+CD25+CD4+ Tregs from the TME. We investigated the difference in therapeutic effects of CD44-targeted NIR-PIT alone, CD25-targeted NIR-PIT alone, and the combination of CD44- and CD25-targeted NIR-PIT in several syngeneic tumor models, including MC38-luc, LL/2, and MOC1. The combined NIR-PIT showed significant tumor growth inhibition and prolonged survival compared with CD44-targeted NIR-PIT alone in all tumor models and showed prolonged survival compared with CD25-targeted NIR-PIT alone in MC38-luc and LL/2 tumors. Combined CD44/CD25 NIR-PIT also resulted in some complete remissions, whereas this was not achieved with either type of NIR-PIT alone. Therefore, combined NIR-PIT simultaneously targeting cancer antigens and immunosuppressive cells in the TME may be more effective than either type of NIR-PIT alone and may have potential to induce prolonged immune responses in treated tumors.