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Published on: June 15, 2016
VISTA expressed on tumor cells is regulated by m6A and influences immune microenvironment through STAT3/CCL22 in
Hao Xu1,2, Kaikai Shen3, Bei Jiang4
1Department of Respiratory and Critical Care Medicine, Jinling Hospital, Nanjing Medical University, Nanjing, China.
Background:
Immunotherapy is playing an increasingly vital role in the treatment of non-small cell lung cancer (NSCLC), yet the challenge of immune evasion remains prevalent. Therefore, investigating the regulatory mechanisms of the tumor immune microenvironment could yield significant benefits.
Methods:
This study utilized differential expression analysis, WGCNA, and Lasso-Cox to analyze the GSE126044 dataset, identifying N6-methyladenosine (m6A)-related molecules that play significant roles in immunotherapy. The m6A modification of the molecule were validated through m6A dot blot, MeRIP, RNA pull-down and RNA stability assays. Their impact on the biological behavior of tumor cells (TCs) was evaluated by CCK-8 assays, wound healing assays, transwell assays, co-culture experiments and flow cytometry. Furthermore, RNA sequencing was conducted to uncover downstream pathways and cytokines, followed by subsequent validation. Lastly, validation was carried out in vivo.
Results:
Through bioinformatics analysis, we identified that V-domain Immunoglobulin Suppressor of T cell Activation (VISTA) serves a crucial role in immunotherapy and that its expression is regulated by m6A. It was subsequently confirmed that METTL3 enhances the m6A modification levels of VISTA mRNA, which is recognized by YTHDF1. This interaction improved the stability of VISTA mRNA, leading to increased expression of VISTA proteins on the surface of TCs. The elevated expression of VISTA boosted the cytotoxic response of CD8 + T cells toward TCs. RNA sequencing analysis indicated that the JAK/STAT pathway significantly contributes to this process. Overexpression of VISTA on TCs decreased the phosphorylation of STAT3, which in turn reduced the expression and release of CCL22. This reduction alleviated immune suppression within the tumor microenvironment, resulting in increased enrichment and activity of CD8 + T cells and decreased enrichment of regulatory T cells (Tregs), ultimately enhancing their cytotoxic effects against TCs.
Conclusion:
We conduct a pioneering systematic study on the expression, function, and molecular mechanisms of the VISTA molecule on NSCLC TCs. We propose that in the tumor immune microenvironment, VISTA functions as both an immune checkpoint and a regulator of CD8 + T cell activation/recruitment and Treg recruitment via its expression on TCs.
Insights
V-domain Immunoglobulin Suppressor of T cell Activation (VISTA) expression on non-small cell lung cancer cells is regulated by m6A modification. This enhances CD8+ T cell responses and reduces immune suppression, offering a novel immunotherapy target.
Area of Science:
- Immunology
- Molecular Biology
- Oncology
Background:
- Immunotherapy is crucial for non-small cell lung cancer (NSCLC) treatment.
- Immune evasion remains a significant challenge in NSCLC treatment.
- Understanding tumor immune microenvironment regulation is vital for improving immunotherapy efficacy.
Purpose of the Study:
- To investigate the role of N6-methyladenosine (m6A)-related molecules in NSCLC immunotherapy.
- To elucidate the regulatory mechanisms of V-domain Immunoglobulin Suppressor of T cell Activation (VISTA) in the tumor immune microenvironment.
- To explore VISTA's function in T cell activation, recruitment, and immune suppression.
Main Methods:
- Differential expression analysis, Weighted Gene Co-expression Network Analysis (WGCNA), and Lasso-Cox regression on the GSE126044 dataset.
- Validation of m6A modification using m6A dot blot, MeRIP, RNA pull-down, and RNA stability assays.
- Assessment of VISTA's impact on tumor cell behavior and in vivo validation.
Main Results:
- V-domain Immunoglobulin Suppressor of T cell Activation (VISTA) expression is regulated by m6A modification, enhanced by METTL3 and recognized by YTHDF1.
- Increased VISTA on tumor cells boosts CD8+ T cell cytotoxicity and alleviates immune suppression via the JAK/STAT pathway.
- VISTA overexpression reduces regulatory T cell (Treg) recruitment and enhances anti-tumor immunity.
Conclusions:
- VISTA plays a critical role in NSCLC immunotherapy by regulating T cell responses and immune suppression.
- VISTA functions as an immune checkpoint and a regulator of CD8+ T cell and Treg recruitment on tumor cells.
- Targeting VISTA expression on tumor cells presents a promising strategy for enhancing NSCLC immunotherapy.
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