G-CSF and G-CSFR Modulate CD4 and CD8 T Cell Responses to Promote Colon Tumor Growth and Are Potential Therapeutic Targets.

Ioannis Karagiannidis,Eric R Prossnitz,Brandon B Phinney,Damian Jacenik,Ruoxin Yao,Stephanie J Jerman,Ellen J Beswick

doi:10.3389/fimmu.2020.01885

Abstract

Cytokines are known to shape the tumor microenvironment and although progress has been made in understanding their role in carcinogenesis, much remains to learn regarding their role in tumor growth and progression. We have identified granulocyte colony-stimulating factor (G-CSF) as one such cytokine, showing that G-CSF is linked with metastasis in human gastrointestinal tumors and neutralizing G-CSF in a mouse model of colitis-associated cancer is protective. Here, we set out to identify the role of G-CSF and its receptor, G-CSFR, in CD4+ and CD8+ T cell responses in the tumor microenvironment. MC38 colon cancer cells were injected into WT, G-CSFR−/− mice, or Rag2−/− mice. Flow cytometry, Real Time PCR and Multiplex cytokine array analysis were used for in vitro T cell phenotype analysis. Adoptive transfer of WT or G-CSFR−/− CD4+ of CD8+ T cells were performed. Mouse tumor size, cytokine expression, T cell phenotype, and cytotoxic activity were analyzed. We established that in G-CSFR−/− mice, tumor growth of MC38 colon cancer cells is significantly decreased. T cell phenotype and cytokine production were also altered, as both in vitro and in vivo approaches revealed that the G-CSF/G-CSFR stimulate IL-10-producing, FoxP3-expressing CD4+ and CD8+ T cells, whereas G-CSFR−/− T cells exhibit increased IFNγ and IL-17A production, leading to increased cytotoxic activity in the tumor microenvironment. Furthermore, peritumoral injection of recombinant IFNγ or IL-17A inhibited colon and pancreas tumor growth compared to controls. Taken together, our data reveal an unknown mechanism by which G-CSF, through its receptor G-CSFR, promotes an inhibitory Treg phenotype that limits tumor immune responses and furthermore suggest that targeting this cytokine/receptor axis could represent a novel therapeutic approach for gastrointestinal, and likely other tumors with high expression of these factors.

Highlights

Colorectal cancer (CRC) is the third most common human malignancy worldwide, and is a leading cause of cancer deaths in the United States [1, 2]
We showed that G-CSF neutralization in the colitis-associated cancer model led to an increase in CD4+ and CD8+ T cells in Abbreviations: CRC, colorectal cancer; G-CSF, granulocyte colony-stimulating factor; Regulatory T cells, Tregs; WT, wild type; Fas ligand (FasL), FAS Ligand; PMNs, polymorphonuclear neutrophils
Since G-CSF has been shown to have pro-tumorigenic properties, we sought to examine how the tumor microenvironment and cytokine production are affected by G-CSF/G-CSF receptor (G-CSFR)

Summary

Introduction

Colorectal cancer (CRC) is the third most common human malignancy worldwide, and is a leading cause of cancer deaths in the United States [1, 2]. G-CSF is best known for its function in signaling to promote the survival, proliferation, differentiation and function of neutrophil precursors via interactions with the G-CSF receptor (G-CSFR) found on neutrophils. Increased expression of G-CSF and its receptor is associated with various human malignancies, including lung [5], brain [6], breast, ovarian, bladder [7], gastric and colon cancers [8, 9]. We have shown G-CSF and G-CSFR to be associated with metastasis in human gastric and colon cancer [10]. Tumors with high expression of G-CSF and G-CSFR are associated with increased tumor cell proliferation, migration and invasion as well as poor patient prognosis [10, 11]. Details of the mechanisms by which GCSF/G-CSFR promote tumor progression and poor outcome remain elusive

Methods

Results

Conclusion