Following the groundbreaking success of PD-1/PD-L1 therapy in solid tumors, numerous immune checkpoints have been harnessed in clinical settings to combat cancer. However, the infiltration of regulatory T cells (Tregs) within tumor tissues contributes to the formation of an immunosuppressive microenvironment, compromising the efficacy of such treatments.

In the quest to address this challenge, multiple attempts have been made to specifically target or eliminate Tregs in the tumor microenvironment, but with limited success. The emergence of Chemokine Receptor 8 (CCR8) as a highly expressed marker on tumor-infiltrating T cells has kindled new hope. By specifically inhibiting the function of Tregs at the tumor site, CCR8 holds the potential to bolster cancer therapy, transforming from an overlooked target to a fervently pursued immunotherapeutic agent.

What is CCR8?

CCR8 is a crucial protein belonging to the beta chemokine receptor family, a group of seven transmembrane proteins that function as G protein-coupled receptors. This unique receptor is primarily expressed in the thymus and is activated by its ligands, CCL1 and CCL8, which are chemokines responsible for attracting and activating various immune cells. One of CCR8's essential roles is its involvement in regulating monocyte chemotaxis and thymic cell apoptosis, which are crucial processes in immune system modulation. Additionally, CCR8 plays a vital role in guiding activated T cells to sites of inflammation and lymphoid tissues, thereby orchestrating immune responses.

An intriguing aspect of CCR8's function lies in its association with regulatory T cells (Tregs). These specialized T cells are essential for suppressing immune responses and maintaining immune tolerance. CCR8 is significantly expressed in tumor-infiltrating Tregs, suggesting its potential involvement in tumor immune evasion and progression.

The signaling mechanism of CCR8 involves the binding of its ligands to the extracellular domain of the receptor. This binding triggers a conformational change that activates intracellular G proteins, initiating a cascade of downstream pathways. These pathways, including phospholipase C, calcium mobilization, protein kinase C, MAP kinases, and NF-kappaB, are responsible for various cellular responses, such as chemotaxis, adhesion, proliferation, survival, and cytokine production^[1].

Recent research has shed light on CCR8's implications in cancer therapy. In the context of the tumor microenvironment, CCL1, one of CCR8's ligands, can activate CCR8 on tumor cells, promoting tumor cell proliferation, migration, and apoptosis resistance. Therefore, drug development targeting the CCL1/CCR8 pathway has shown promise for enhancing anti-tumor immune responses. Moreover, CCL1 also recruits Treg cells to the tumor microenvironment and induces the transformation of CD4+ T cells into Tregs, further promoting tumor immune escape. Hence, therapeutic antibodies targeting CCR8 not only function to block its activity but also leverage the Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC) effect to deplete FOXp3+ CCR8+ Treg cells infiltrating the tumor. This strategy has demonstrated significant tumor growth inhibition in preclinical studies.

The significance of the CCL1→CCR8 axis in cancer processes^[2]

In a research report published by Campbell JR, et al^[3], it was confirmed that CCR8 therapeutic antibodies with a robust ADCC profile (anti-mCCR8-IgG2a) effectively suppressed the growth of MC38 tumors. On the other hand, antibodies with a weak ADCC profile (anti-mCCR8-IgG1-D265A) showed less pronounced tumor inhibition.

Anti-mouse CCR8 antibody activity in MC38 tumor model^[3]

Currently, ongoing research on CCR8 therapy mainly focuses on ADCC-dependent antibody drugs, highlighting the promising potential of CCR8-targeted treatments in combating cancer and modulating immune responses.

Although there are no approved CCR8-targeted drugs worldwide at present, numerous CCR8-targeted drugs are in various stages of research, underscoring the intense competition within the development field. Among them, Bristol-Myers Squibb's BMS-986340, Shionogi's S-531011, and Lixen Pharmaceuticals' LM-108 are relatively advanced in their development progress. To better support CCR8-related research, GemPharmatech has employed gene editing technology to replace the CCR8 gene in BALB/c mice entirely with its human counterpart, resulting in the development of the BALB/c-hCCR8 humanized mouse model. This model effectively expresses the human CCR8 protein and has successfully validated the in vivo efficacy of CCR8 antibody drugs.

In vivo efficacy evaluation of anti-hCCR8 antibody in BALB/c-hCCR8 mouse

We established a robust preclinical model for evaluating the efficacy of anti-human CCR8 antibodies by transplanting CT26 colon cancer cells into BALB/c-hCCR8 homozygous mice subcutaneously. Mice were grouped into a control group (treated with PBS) and three treatment groups (treated with TA-1, TA-2, and TA-3 antibodies) when the average tumor volume reached approximately 80mm³. The results demonstrated varying inhibitory effects on tumor growth among the three different anti-human CCR8 antibodies. Specifically, TA-2 and TA-3 antibodies exhibited more pronounced tumor growth inhibition, with TA-3 showing the most significant anti-tumor efficacy (TGI = 56%). Figure A depicts the average tumor volume ± SEM, and Figure B illustrates the average mouse weight ± SEM during the experiment.

These findings underscore the potential of BALB/c-hCCR8 mice as a robust in vivo model for preclinical evaluation of anti-human CCR8 antibodies, offering valuable insights into their therapeutic effectiveness against CCR8-associated diseases, including colon cancer.

Flow Cytometric Analysis of Tumor-Infiltrating Lymphocytes (TILs)

At the end of the experiment, tumor tissues from the control group and treatment groups were collected for flow cytometric analysis to investigate the immune cell subpopulations. The treatment groups with anti-human CCR8 antibodies (TA-1, TA-2, and TA-3) exhibited a significant reduction in the proportion of total Treg cells and CCR8+ Treg cells compared to the control group. These results indicate that antibody drugs targeting CCR8 can enhance anti-tumor immunity by depleting Tregs expressing CCR8, which possess immunosuppressive capabilities. These results highlight the potential of CCR8-targeted therapeutic antibodies as a promising strategy to bolster anti-tumor immune responses by targeting the immunosuppressive Treg subset infiltrating the tumor microenvironment.

Reference:

1. Haque NS, Fallon JT, Taubman MB, Harpel PC. The chemokine receptor CCR8 mediates human endothelial cell chemotaxis induced by I-309 and Kaposi sarcoma herpesvirus-encoded vMIP-I and by lipoprotein(a)-stimulated endothelial cell conditioned medium. Blood. 2001 Jan 1;97(1):39-45. doi: 10.1182/blood.v97.1.39. PMID: 11133740.

2. Korbecki J, Grochans S, Gutowska I, Barczak K, Baranowska-Bosiacka I. CC Chemokines in a Tumor: A Review of Pro-Cancer and Anti-Cancer Properties of Receptors CCR5, CCR6, CCR7, CCR8, CCR9, and CCR10 Ligands. Int J Mol Sci. 2020 Oct 15;21(20):7619.

3. Campbell JR, McDonald BR, Mesko PB, Siemers NO, Singh PB, Selby M, Sproul TW, Korman AJ, Vlach LM, Houser J, Sambanthamoorthy S, Lu K, Hatcher SV, Lohre J, Jain R, Lan RY. Fc-Optimized Anti-CCR8 Antibody Depletes Regulatory T Cells in Human Tumor Models. Cancer Res. 2021 Jun 1;81(11):2983-2994.