A novel peptide targeting CCR7 inhibits tumor cell lymph node metastasis

Cancer Immunology, Immunotherapy (2025) 74:153 https://doi.org/10.1007/s00262-025-03995-4 RESEARCH A novel peptide targeting CCR7 inhibits tumor cell lymph node metastasis Yixuan Sun1 · Yuzhen Qian2 · Lu Qiu1 · Xueqin Zhu1 · Haoming Ning1 · Liwei Pang2 · Xiaoshuang Niu1 · Yi Liu1 · Xiuman Zhou1 · Guanyu Chen1 · Wenjie Zhai2 · Yanfeng Gao1 Received: 27 August 2024 / Accepted: 21 February 2025 © The Author(s) 2025 Abstract Lymph nodes are the most common metastasis sites for tumor cells, which are intimately linked to patient prognosis. It has been reported that cancer cells can upregulate CC Chemokine Receptor 7 (CCR7) expression and hijack its normal functions, enabling them to migrate along the gradient of CCL19 and CCL21 toward the lymph nodes and colonies as the initial stage of distant metastasis. In tumor patients, the metastatic tumor in the lymph nodes exhibited higher expression of CCR7, as well as inhibitory immune checkpoints PD-1, LAG-3, and TIM-3 compared to the primary tumors with the analysis of TCGA and GEO databases. Also, in mouse tumor model, tumor cells with elevated CCR7 expression were more susceptible to develop popliteal lymph node metastasis. Subsequently, we successfully identified a CCR7 binding peptide TC6 by phage display biopanning, which specifically blocks the interaction of CCR7/CCL19 and CCR7/CCL21. Further, the D-amino acids were introduced to substitute the N- and C-terminus of TC6 peptide to obtain the proteolysis-resistant TC6-D3 peptide, which decreased tumor cell migration in vitro via ERK1/2 pathway and inhibited tumor growth and lymph nodes metastasis in vivo, as well as effectively restored T cells cytotoxicity in both primary tumors and lymph nodes. In conclusion, CCR7 promoted tumor cell metastasis to lymph node and inhibited the anti-tumor immune responses in lymph nodes. Specific blockade of the CCR7 pathway with TC6-D3 peptide can significantly reduce lymph node tumor burden, promoting CD8+ T cell infiltration in primary tumors, meanwhile, enhancing anti-tumor immune responses in lymph nodes. Keywords CCR7 · Lymph node metastasis · Peptide · Blockade · Immune responses Introduction Lymph node (LN) metastasis represents a common feature across a wide range of solid tissue malignancies and closely associated with poor survival [1–3]. Cancer cells are often prior to regionally metastasize to adjacent LN before metastasizing systemically through the lymphatics and blood, which induces tumor-specific immune tolerance and thus Yixuan Sun and Yuzhen Qian have contributed equally to this work. * Wenjie Zhai * Yanfeng Gao 1 School of Pharmaceutical Sciences (Shenzhen), Sun Yat-sen University, Shenzhen 518107, China 2 School of Life Sciences, Zhengzhou University, Zhengzhou 450001, China renders distant tissues amenable to metastatic colonization [4]. Recent studies have revealed the decreased oxidative stress and ferroptosis in lymph, including higher levels of glutathione and oleic acid, lesser free iron in LN-protected melanoma cells from ferroptosis and increased their capacity to form metastatic tumors [5, 6]. Engleman et al. previously found that metastatic tumor cells in LN initially upregulated MHC-I and PD-L1 to evade cytotoxicity of natural killing (NK) cells and T cells in response to chronic interferon (IFN) signaling, altering the immune compartment within LN to induce antigen-specific Treg differentiation and immune tolerance, further promoting metastasis to distant tissues [4]. These findings indicate that tumor cells alter the metabolic pathways and phenotypes to escape immune surveillance in LNs. In addition, the specifically expressed chemokines in LNs induced the prior metastasis of tumor cells [7]. Since the chemokine receptors regulate immune cell migration and survival, that often stimulated by inflammatory signals, their expression on tumor cells may reflect Vol.:(0123456789) 153 Page 2 of 16 aberrant inflammatory signals and lead to enhanced tumor growth, invasion, and metastasis. Chemokines are a family of cytokines that mediate leukocyte trafficking. Among them, chemokine receptor 7 (CCR7) with C–C chemokine ligand 19/21 pair (CCL19/CCL21), which belong to the highly druggable G-protein-coupled receptor (GPCR) family, plays a vital role in the migration of dendritic cells (DCs) to LNs [8]. Since overexpressed CCR7 dramatically correlates with larger primary tumors, deeper lymphatic invasion, and poorer survival rates, CCR7 has emerged as an important biomarker for predicting the axillary LNs metastasis in melanoma [9], colorectal [10], breast [11], and esophageal squamous cell carcinoma [12], etc. Recently, it has been reported that cancer cells could upregulate CCR7 expression and hijacked its normal functions, enabling them to migrate along the gradient of CCL19 and CCL21 secreted by the lymphatic endothelial cells and high endothelial venules and toward the LN to colonize them as the first step toward metastasis [13] such as B16-F10, 4T1, and MCF7 mouse model [14–16]. In vivo studies revealed that the tumor growth, angiogenesis, and metastatic formation are decreased when downregulating CCR7 or applying CCR7 antagonists [15, 17]. Therefore, CCR7 is suggested as a promising therapeutic target and blocking the CCR7CCL21/CCL19 axis can effectively inhibit the tumor growth and occurrence of LN metastasis. Due to the central role of CCR7 in tumor metastasis, several therapeutic strategies have been developed to exploit this axis for cancer treatment, including siRNA interfering the CCR7 expression, antibodies, or small molecule inhibitors against CCR7. Recently, Joerg Standfuss et al. screened the small molecule inhibitor Cmp2105, specifically binding to the intracellular domain of CCR7 and inhibiting its activation [18], but its biological activity and the effects on inhibiting tumor cell metastasis to LNs in vivo remain obscure. In addition, Leaf Huang et al. delivered a plasmid encoding an antagonistic CCR7 protein trap by tumor-targeting nanoparticles in a highly metastatic 4T1 mouse model, which locally disrupted the CCR7 signaling pathways in the tumor site and efficiently inhibited 4T1 lymphatic metastasis [19]. Besides, it has been reported that CCR7 therapeutic antibody can effectively block the invasion and metastasis of tumor cells. Carlos Cuesta-Mateos et al. generated CAP-100, an antibody specifically binds to human CCR7 and neutralizes its ligand binding site and intracellular signaling pathways, as well as strongly inhibits the CCR7-induced migration, extravasation, homing, and survival in CLL samples in vitro and in vivo preclinical models, and the clinical trial (NCT04704323) to evaluate this novel therapeutic approach in CLL patients is pending [20]. Accordingly, all results validated that targeting CCR7 is an effective therapeutic strategy to prevent the access of tumor cells into the LN niches, which has a vital clinical application value. Cancer Immunology, Immunotherapy (...truncated)