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Lysophosphatidylserine Receptor LPS1 Signaling Enhances the Anti-Tumor Immunity of Macrophages
Shun Nishikado1, Jumpei Omi1, Akiharu Uwamizu1, Tomohiko Ohwada1 and Junken Aoki11Graduate School of Pharmaceutical Sciences, Tokyo University, 7-3-1, hongo, Bunkyo-ku, Tokyo, Japan
Background: Lysophosphatidylserine (LysoPS) is a unique lysophospholipids with serine as a polar head and augments various immune responses through activating its specific receptors, LPS1/GPR34, LPS2/P2Y10 and LPS3/GPR174 [1][2] (doi: 10.1111/imr.13204, doi.org/10.1101/2023.02.15.528751). We previously found that LPS1-KO mice showed significantly increased tumor growth and reduced Granzyme B positive cytotoxic CD8+ T lymphocyte population in the tumor compared to wild-type mice in the tumor-bearing model. These findings suggest that LPS1 signaling enhances anti-tumor immunity, but the detailed mechanisms for LPS1 signaling in anti-tumor immunity remain unclear. Our qRT-PCR analysis and previously published single-cell RNA-seq dataset [3] showed that LPS1 is highly expressed in tumor-associated macrophages (TAM). Thus, we hypothesized that TAM had an important role in LPS1 signaling in anti-tumor immunity. In this study, we performed a functional analysis of LPS1 signaling using TAM and bone marrow derived-macrophage (BMDM).
Methods: In the tumor-bearing model, murine colon adenocarcinoma MC38 cells were subcutaneously injected into both flanks of mice. Tumor growth was monitored from day 7 to 16 after the injection. On day 16, the tumor tissues were harvested and subjected to flow cytometry analysis for the evaluation for the inflammatory capacity of the TAMs (CD45+ F4/80+ CD11b+) or the population of Granzyme B-expressing cytotoxic CD8+ T lymphocytes in the tumor. In some experiments, the tumor-bearing mice were treated with the LPS1 agonist [2][4] (100µM x 200µl/i.p.) twice per day. The expression of cell surface M1/M2 markers (MHCⅠ/Ⅱ, CD80/86, PD-L1, iNOS, and CD206) on these TAMs was evaluated by FACS. We also analyzed the mRNA levels of pro-inflammatory cytokines (Il-6, Tnfa, Il1b, and Il12) and chemokines (Cxcl1/9/10 and Ccl3/5) in BMDMs after they were treated with lipopolysaccharide (LPS) to induce their M1 phenotypes. To investigate whether LPS1 signaling on macrophages enhances the anti-tumor immunity, we intratumorally (i.t.) injected BMDMs and monitored the tumor growth over time. We also determined the immune cell population in the tumor tissues.
Results: TAMs from LPS1-KO mice showed the reduced expression of M1 markers (MHCI, MHCII, CD80, PD-L1, and iNOS) compared to TAMs from wild-type mice. We also found the attenuated M1 phenotypes in BMDMs prepared from LPS1-KO bone marrow after they were stimulated with LPS, as judged by reduced expression of pro-inflammatory cytokines (Il-6, Tnfa, Il1b and Il12) and chemokines (Cxcl1/9/10 and Ccl3/5). The i.t. injection of wild-type BMDMs but not LPS1-KO BMDMs significantly suppressed the tumor growth. Consistent with this, the i.t. injection of wild-type BMDMs, but not LPS1-KO BMDMs, increased the Granzyme B-positive cytotoxic CD8+ T lymphocyte in tumor tissues. Finally, we found that LPS1 agonist significantly suppressed the tumor growth in wild-type mice, which was not observed in LPS1-KO mice.
Conclusion: Present study revealed that LPS1 signaling on macrophages has a role to enhance tumor immunity. LPS1 signaling in macrophages may exert its anti-tumor activity by inducing the cells to have an M1 phenotype and by shaping the proinflammatory tumoricidal niche, resulting in enhancing the anti-tumor immunity. LPS1 agonist administration dramatically reduced the tumor size in an LPS1-specific manner. Thus, LPS1 is a promising drug target for novel cancer therapeutics.
Reference
[1]Shun Y., et al. Immunologycal Reviews. 2023
[2]Tamaki I., et al. bioRxiv. 2023
[3]Lei Zhang. et al. Cell. 2020
[4]Sho N., et al. J Med Chem. 2020