Simultaneous TLR2 inhibition and TLR9 activation synergistically suppress tumor metastasis in mice
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Updated: May 24, 2026
Experimental Metastasis and CTL Adoptive Transfer Immunotherapy Mouse Model
Published on: November 26, 2010
Jun Yan1, Fang Hua, Han-zhi Liu
1Molecular Immunology and Pharmacology Laboratory, Institute of Meteria Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
Aim:
To develop a rational immunotherapy against tumor metastasis by combining a Toll-like-receptor 2 (TLR2)-neutralizing antibody with a TLR9 agonist CpG ODN, and then investigate the mechanism of action for this combinational regimen.
Methods:
After mouse melanoma B16-F10 cell inoculation, female C57BL/6 mice were treated with either CpG ODN (0.5 mg/kg) or the anti-TLR2 antibody (200 μg/kg), or with a combination of the two agents. Pulmonary metastases were evaluated by counting metastatic nodes on the lung surface using anatomical microscopy. Flow cytometry was used to evaluate the cytotoxicity of the immune cells in tumor-draining lymph nodes, the cell population in the spleen, and the infiltration of immune cells within the lungs. Cytokine and enzyme expression in the lung tissue was evaluated using ELISA or immunostaining.
Results:
Anti-metastatic effects were detected in mice treated with either CpG ODN or the anti-TLR2 antibody alone. However, treatment with CpG ODN plus the anti-TLR2 antibody synergistically suppressed the metastasis as compared with treatment with either single agent. The combinational treatment resulted in enhanced infiltration of natural killer cells and cytotoxic T cells, reduced recruitment of type 2 macrophages and Tregs, and decreased expression of immunosuppressive factors including TGF-β1, cyclooxygenase-2 and indoleamine 2,3-dioxygenase, thus stimulated tumor cytotoxicity and suppressed metastasis. The anti-metastatic effect of the combinational regimen was further confirmed in spontaneous metastatic mouse model of Lewis lung carcinoma.
Conclusion:
Our studies suggest that combining a TLR9 agonist with an anti-TLR2 antibody, which eliminates immunosuppressive factors from the tumor environment, is critical for an effective anti-metastatic immunotherapy.
Insights
Combining a Toll-like receptor 2 (TLR2) neutralizing antibody with a TLR9 agonist CpG ODN synergistically suppresses tumor metastasis. This immunotherapy enhances anti-tumor immune cells and reduces immunosuppressive factors for effective cancer treatment.
Area of Science:
- Immunology
- Cancer Research
- Pharmacology
Background:
- Tumor metastasis remains a significant challenge in cancer treatment.
- Current immunotherapies often face limitations in overcoming the immunosuppressive tumor microenvironment.
Purpose of the Study:
- To develop a novel immunotherapy for anti-metastasis by combining a TLR2-neutralizing antibody with a TLR9 agonist (CpG ODN).
- To elucidate the mechanism of action of this combination therapy against tumor metastasis.
Main Methods:
- Mice bearing melanoma B16-F10 or Lewis lung carcinoma were treated with CpG ODN, anti-TLR2 antibody, or both.
- Pulmonary metastases were quantified, and immune cell populations, cytotoxicity, and immunosuppressive factors were analyzed via flow cytometry, ELISA, and immunostaining.
Main Results:
- The combination therapy synergistically suppressed pulmonary metastasis compared to single-agent treatments.
- This regimen enhanced natural killer and cytotoxic T cell infiltration while reducing type 2 macrophages and Tregs.
- Expression of immunosuppressive factors like TGF-β1, COX-2, and IDO was decreased, boosting tumor cytotoxicity.
Conclusions:
- Combining a TLR9 agonist with an anti-TLR2 antibody is a critical strategy for effective anti-metastatic immunotherapy.
- This approach neutralizes immunosuppressive factors, creating a more favorable tumor environment for immune attack.
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