Synthetic Biology in Cancer Immunotherapy: Engineering Bacteria to Inhibit IDO Activity and Reprogram CD8+ T Cell Responses

Academic Background

In recent years, significant progress has been made in cancer immunotherapy, particularly through the activation of T cells to combat tumors. However, metabolic adaptations in the tumor microenvironment (TME) often impair T cell function, limiting the efficacy of immunotherapy. Tryptophan (Trp) metabolism plays a crucial role in T cell function. Tumor cells consume tryptophan by expressing indoleamine 2,3-dioxygenase (IDO), leading to the accumulation of its metabolite kynurenine (Kyn), which suppresses effector T cell function and creates an immunosuppressive microenvironment. Although IDO inhibitors have shown promise in preclinical models, they face numerous challenges in clinical trials, such as off-target effects and toxicity.

To address these issues, researchers have begun exploring synthetic biology techniques to genetically engineer bacteria to modulate metabolic signals in the tumor microenvironment, thereby enhancing the anti-tumor activity of T cells. This study genetically engineered Clostridium butyricum (CB) to continuously release tryptophan and butyrate, thereby inhibiting IDO activity and reprogramming CD8+ T cell metabolism, ultimately achieving tumor growth suppression.

Source of the Paper

This paper was co-authored by Heng Wang, Fang Xu, Chenlu Yao, and others, affiliated with Soochow University and other research institutions. The paper was published on December 18, 2024, in PNAS (Proceedings of the National Academy of Sciences), titled “Engineering bacteria for cancer immunotherapy by inhibiting IDO activity and reprogramming CD8+ T cell response.”

Research Process and Results

1. Genetic Engineering of Clostridium butyricum (L-Trp CB)

The researchers first genetically engineered Clostridium butyricum by inserting tryptophan synthesis-related genes (trpE, trpD, trpC, trpB, trpA) into the plasmid of CB, constructing an engineered strain (L-Trp CB) capable of continuously releasing tryptophan. Quantitative PCR and high-performance liquid chromatography (HPLC) analysis confirmed that L-Trp CB efficiently expressed tryptophan synthase and released large amounts of tryptophan. Additionally, L-Trp CB retained the ability to produce butyrate, which inhibits IDO activity, preventing tryptophan catabolism and kynurenine accumulation.

2. Effects of L-Trp CB on Tumor Cells and T Cells

In vitro experiments showed that butyrate released by L-Trp CB significantly reduced IDO expression in various tumor cells, thereby decreasing tryptophan consumption. Meanwhile, tryptophan released by L-Trp CB significantly promoted the proliferation and function of CD8+ T cells, enhancing their effector phenotype. Metabolic analysis revealed that L-Trp CB significantly increased oxidative phosphorylation and glycolysis in CD8+ T cells, thereby enhancing their energy metabolism and anti-tumor activity.

3. Anti-Tumor Effects of L-Trp CB in Animal Models

In multiple mouse and rabbit tumor models, L-Trp CB selectively colonized tumor tissues and significantly inhibited tumor growth. Bioluminescence imaging and flow cytometry analysis showed that L-Trp CB significantly increased the proportion and function of tumor-infiltrating CD8+ T cells while reducing immunosuppressive signals in the tumor microenvironment. Furthermore, the combination of L-Trp CB with PD-L1 blockade produced synergistic anti-tumor effects, significantly prolonging animal survival.

4. Reprogramming CD8+ T Cell Metabolism by L-Trp CB

Through single-cell RNA sequencing and gene set enrichment analysis (GSEA), researchers found that L-Trp CB significantly upregulated genes related to glycolysis, the tricarboxylic acid (TCA) cycle, and oxidative phosphorylation in CD8+ T cells, thereby enhancing their energy metabolism and effector function. Additionally, L-Trp CB activated the mTORC1 signaling pathway, further promoting metabolic reprogramming and anti-tumor activity in CD8+ T cells.

Conclusions and Significance

This study genetically engineered Clostridium butyricum to develop a strain (L-Trp CB) capable of continuously releasing tryptophan and butyrate. This strain selectively colonizes tumor tissues, inhibits IDO activity, and reprograms CD8+ T cell metabolism, significantly suppressing tumor growth. This research provides a novel synthetic biology strategy for cancer immunotherapy, with significant scientific and application value.

Research Highlights

1. Innovation: This study is the first to genetically engineer Clostridium butyricum to simultaneously release tryptophan and butyrate, thereby inhibiting IDO activity and reprogramming CD8+ T cell metabolism.
2. Efficacy: L-Trp CB demonstrated significant anti-tumor effects in multiple animal models and showed synergistic effects when combined with PD-L1 blockade.
3. Safety: L-Trp CB exhibited good tolerance in animal models, with no significant side effects observed.

Future Prospects

Although this study achieved significant results, some issues remain to be explored. For example, the safety and efficacy of L-Trp CB in humans need to be validated through clinical trials. Additionally, the effects of L-Trp CB on other immune cells (e.g., myeloid cells) require further investigation. In the future, researchers can optimize the design of engineered strains to further enhance their anti-tumor efficacy and clinical application potential.

This study provides a novel synthetic biology strategy for cancer immunotherapy, with broad application prospects.