The Combination of CD300c Antibody with PD-1 Blockade Suppresses Tumor Growth and Metastasis by Remodeling the Tumor Microenvironment in Triple-Negative Breast Cancer Soyoung Kim, Ik-Hwan Han, Suin Lee, DaeHwan Park, Hyunju Lee, et al. International Journal of Molecular Sciences, 2025 Triple-negative breast cancer (TNBC) is an aggressive cancer characterized by a high risk of recurrence, invasiveness, metastatic potential, and poor prognosis. Tumor-associated macrophages (TAMs), particularly M2-like TAMs, contribute to TNBC progression by promoting an immunosuppressive tumor microenvironment (TME), highlighting the need for TME remodeling. This study aimed to evaluate the therapeutic efficacy of co-administering CL7, a CD300c monoclonal antibody that induces M1 macrophage polarization, and anti-PD-1, an immune checkpoint inhibitor, in TNBC. To establish a TNBC model, 4T1 cells were inoculated into the fourth left mammary gland of mice. CL7 and anti-PD-1 were intravenously administered twice a week. Flow cytometry and RT-PCR were performed to assess the immunotherapeutic effects, and lung metastases were evaluated by the Hematoxylin and Eosin staining of lung tissues. Tumor growth was significantly reduced in the combination treatment group (CL7 and anti-PD-1) compared to both the PBS and monotherapy groups. Additionally, the combination treatment increased M1 macrophages and activated CD8+ T and NK cells in the tumor, while significantly suppressing lung metastases. These findings suggest that the combination of CL7 and anti-PD- therapy has the potential to treat TNBC by remodeling the TME.
Conformation-sensitive targeting of CD18 depletes M2-like tumor-associated macrophages resulting in inhibition of solid tumor progression Ik-Hwan Han, Ilseob Choi, Hongseo Choi, Soyoung Kim, Chanmi Jeong, Juwon Yang, Yingying Cao, Jeongyoon Choi, Heekyung Lee, Jin Sun Shin, Hye Duck Yeom, Eun-Ji Lee, Nari Cha, Hyemin Go, Se Eun Lim, Songah Chae, Won-Jun Lee, Minjin Kwon, Hongsung Kim, Hyojung Choi, Sehyun Pak, Namgyeong Park, Eunbin Ko, Deok-Sang Hwang, Junho H Lee, Hwan-Suck Chung, Seong Ho Kang, Hyunsu Bae Journal for Immunotherapy of Cancer, 2025 BackgroundTumor-associated macrophages (TAMs) primarily exist in the M2-like phenotype in the tumor microenvironment (TME). M2-TAMs contribute to tumor progression by establishing an immunosuppressive environment. However, TAM targeting is hindered, mainly owing to a lack of specific biomarkers for M2-TAMs. Previously, we demonstrated that a novel peptide drug conjugate (TB511) consisting of a TAM-binding peptide and the apoptosis-promoting peptide targets M2-TAMs. This was achieved through M2-TAM targeting, although the target mechanism of action remained elusive. Herein, we elucidate the anticancer efficacy of TB511 by identifying new target proteins that preferentially bind to M2-TAMs and clarifying the apoptosis-inducing mechanism in these cells.MethodsWe investigated the target proteins and binding site of TB511 using LC-MS/MS analyses, surface plasmon resonance and peptide–protein interaction 3D modeling. Activated CD18 expression in M2 TAMs was assessed using Quantibrite PE beads in PBMCs. The anticancer efficacy of TB511 was tested using colorectal cancer (CRC) and non-small cell lung cancer (NSCLC) mouse model. The immunotherapeutic effect of TB511 was investigated through spatial transcriptomics in human pancreatic ductal adenocarcinoma (PDAC) model.ResultsActivated CD18 was highly expressed in human tumor tissues and was significantly higher in M2 TAMs than in other immune cells. TB511 showed high binding affinity to CD18 among the cell membrane proteins of M2 macrophages and appeared to bind to the cysteine-rich domain in the activated form. Moreover, TB511 specifically induced apoptosis in M2 TAMs, but its targeting ability to M2 macrophages was inhibited in CD18 blockade or knockout model. In mouse or humanized mouse models of solid tumors such as CRC, NSCLC, and PDAC, TB511 suppressed tumor growth by targeting M2-TAMs via CD18 and enhancing the presence of CD8+T cells in the TME.ConclusionsCollectively, our findings suggest that activated CD18 holds promise as a novel target protein for cancer therapy, and TB511 shows potential as a therapeutic agent for tumor treatment.
Immunomodulatory peptide–drug conjugate MEL-dKLA suppresses progression of prostate cancer by eliminating M2-like tumor-associated macrophages Ik-Hwan Han, Ilseob Choi, Soyoung Kim, Minjin Kwon, Hyojung Choi, Hyunsu Bae Frontiers in Immunology, 2025 Prostate cancer is one of the most common malignancies in men and is frequently associated with tumor-promoting inflammation. Tumor-associated macrophages (TAMs) are known to facilitate cancer progression by suppressing antitumor immunity. Therefore, targeting TAMs represents a promising strategy for cancer therapy. This study aimed to investigate whether melittin-dKLA, a conjugated peptide consisting of melittin (MEL), which selectively binds M2-like macrophages, and the pro-apoptotic peptide d(KLAKLAK)2 (dKLA), can inhibit prostate cancer progression by targeting M2 macrophages. Human monocytic cells (THP-1 cells) were differentiated into TAMs using tumor-conditioned medium (TCM), and the conditioned medium from these TAMs was termed M-TCM. MEL-dKLA binding affinity was assessed using FITC-labeled melittin. A prostate cancer mouse model was established by subcutaneous injection of TRAMP-C2 cells, followed by MEL-dKLA administration every three days. As a result, THP-1-derived macrophages stimulated with TCM exhibited elevated expression of M2 markers (ARG1, CD206, and CD163). Prostate cancer cells (PC-3) stimulated with M-TCM showed increased proliferation and expression of epithelial-mesenchymal transition (EMT) markers. MEL-dKLA preferentially bound to M2 macrophages and TAMs, and inducing selective cytotoxicity. Conditioned media from MEL-dKLA-treated M2 macrophages and TAMs resulted in markedly decreased PC-3 cell proliferation, migration, and invasion. In vivo, MEL-dKLA treatment significantly reduced tumor growth, decreased the number of CD163+ M2 macrophages, and increased CD8+ T cell infiltration in tumor tissues. These findings demonstrate that MEL-dKLA suppresses prostate cancer progression by targeting M2-like TAMs both in vitro and in vivo. MEL-dKLA may serve as a promising therapeutic agent to modulate the tumor microenvironment in prostate cancer.
The CD300c antibody CL7 suppresses tumor growth by regulating the tumor microenvironment in non-small cell lung carcinoma Soyoung Kim, Ik-Hwan Han, Suin Lee, Sujin Park, Jae-Won Jeon, Hyunsu Bae Frontiers in Oncology, 2025 Despite advances in therapy, non-small cell lung cancer (NSCLC) continues to rank among the deadliest cancers worldwide. Targeting immunosuppressive components within the tumor microenvironment (TME) has emerged as a promising therapeutic strategy. Unlike M1 tumor-associated macrophages (TAMs), M2-like TAMs contribute to NSCLC progression by promoting an immunosuppressive tumor microenvironment (TME), highlighting the need for tumor microenvironment remodeling. CL7, a monoclonal antibody that targets the activating receptor CD300c on human monocytes and macrophages, was selected as a therapeutic candidate because CD300c engagement triggers MAPK and NF-κB signaling pathways, promoting M1 macrophage polarization and antitumor immune activation. To evaluate the therapeutic potential of CL7, we established an orthotopic NSCLC model by inoculating LLC-luc cells into the left lung of mice. We administered CL7 intraperitoneally at doses of 5 or 10 mg/kg twice a week. Only representative data from the 10 mg/kg CL7 group are shown to maintain consistency with subsequent analyses (flow cytometry, RT-qPCR, and IHC). Tumor growth was significantly suppressed in the CL7-treated group compared to the PBS control group. CL7 treatment also modulated the tumor microenvironment by increasing the population of M1 macrophages and CD8 + T cells, while decreasing the population of regulatory T cells. Our findings suggest that CL7 exerts antitumor effects in NSCLC by reprogramming the immunosuppressive landscape of the TME and enhancing antitumor immunity.
Adoptive Transfer of CX3CR1-Transduced Tregs Homing to the Forebrain in Lipopolysaccharide-Induced Neuroinflammation and 3xTg Alzheimer’s Disease Models Hyejin Yang, Juwon Yang, Namgyeong Park, Deok-Sang Hwang, Seon-Young Park, Soyoung Kim, Hyunsu Bae International Journal of Molecular Sciences, 2024 CX3CR1-transduced regulatory T cells (Tregs) have shown potential in reducing neuroinflammation by targeting microglial activation. Reactive microglia are implicated in neurological disorders, and CX3CR1-CX3CL1 signaling modulates microglial activity. The ability of CX3CR1-transduced Tregs to inhibit LPS-induced neuroinflammation was assessed in animal models. CX3CR1 Tregs were administered to LPS-induced and 3xTg Alzheimer’s mouse models, resulting in reduced proinflammatory marker expression in both the cortices and hippocampi. In the 3xTg Alzheimer’s model, neuroinflammation was significantly reduced, demonstrating the efficacy of CX3CR1 Tregs even in chronic neuroinflammatory conditions. These findings highlight the therapeutic potential of CX3CR1 Treg therapy in modulating microglial activity and offer promising treatment strategies for neurodegenerative diseases.
A preclinical and phase I clinical study of ex vivo-expanded amyloid beta-specific human regulatory T cells in Alzheimer's disease Hyejin Yang, Min Soo Byun, Na-Yeon Ha, Juwon Yang, Seon-Young Park, Jee Eun Park, Dahyun Yi, Young-Tae Chang, Woo Sang Jung, Jae Yoon Kim, Jinsung Kim, Dong Young Lee, Hyunsu Bae Biomedicine and Pharmacotherapy, 2024 INTRODUCTION: Despite advancements in adoptive regulatory T cell (Treg) therapy, its application in Alzheimer's disease (AD) remains constrained by challenges in ex vivo Treg selection and expansion with antigen specificity. Our previous findings demonstrated the bystander suppressive immunomodulatory mechanism of ex vivo expanded amyloid β-specific mouse Tregs in AD models, prompting inquiry into the efficacy of ex vivo expanded human Tregs in AD. METHODS: We developed an effective ex vivo expansion method for manufacturing amyloid β-specific human Tregs (Aβ-hTreg) and evaluated their safety and efficacy in 3xTg mouse models of AD and a phase 1 clinical trial with six AD patients. The phenotype of Aβ-hTreg was analyzed using single-cell transcriptomics. The clinical trial involved intravenous administration of Aβ-hTreg, with three patients receiving a low dose and three receiving a high dose. Exploratory assessments of effectiveness, including cognitive tasks and functional evaluations, were conducted ninety days post-treatment. RESULTS: Behavioral spatial learning and memory impairment, neuroinflammatory and amyloid pathology were dramatically ameliorated by single intrathecal administration of ex vivo expanded Aβ-hTreg to 3xTg AD mice. Single cell transcriptomics analysis revealed alterations in five key genes within a cluster of Tregs under antigen-specific manufacturing conditions. In the clinical trial with six AD patients, dose-limiting toxicity was experienced by none of the participants within five days of receiving GMP-grade Aβ-hTreg (VT301), indicating its good tolerability. Although exploratory assessments of effectiveness did not reach statistically significant values among the groups, these findings offer valuable insights for AD treatment and management, guiding the planning of the next phase of clinical trials. DISCUSSION: This study suggests that hTregs may modulate Alzheimer's disease pathology by suppressing neuroinflammation, while VT301 shows promise as a safe treatment option. However, further research is necessary to confirm its clinical efficacy and optimize treatment strategies. TRIAL REGISTRATION: Title: A Study of Possibility of Using Regulatory T Cells (VT301) for Treatment of Alzheimer's Disease, ClinicalTrials.gov NCT05016427, Study approval date: Ministry of Food and Drug Safety of the Republic of Korea (MFDS) - August 31st, 2020, Institutional Review Board (IRB) of Seoul National University Hospital, Republic of Korea - September 29th, 2020, The date of first patient enrollment: December 7th, 2020. https://clinicaltrials.gov/study/NCT05016427.
Therapeutic effects of MEL-dKLA by targeting M2 macrophages in pulmonary fibrosis Ilseob Choi, Ik-Hwan Han, Nari Cha, Hye Yeon Kim, Hyunsu Bae Biomedicine and Pharmacotherapy, 2024 Idiopathic pulmonary fibrosis is a progressive lung disease characterized by excessive extracellular matrix accumulation and myofibroblast proliferation with limited treatment options available. M2 macrophages are pivotal in pulmonary fibrosis, where they induce the epithelial-to-mesenchymal and fibroblast-to-myofibroblast transitions. In this study, we evaluated whether MEL-dKLA, a hybrid peptide that can eliminate M2 macrophages, could attenuate pulmonary fibrosis in a cell co-culture system and in a bleomycin-induced mouse model. Our findings demonstrated that the removal of M2 macrophages using MEL-dKLA stimulated reprogramming to an antifibrotic environment, which effectively suppressed epithelial-to-mesenchymal and fibroblast-to-myofibroblast transition responses in lung epithelial and fibroblast cells and reduced extracellular matrix accumulation both in vivo and in vitro. Moreover, MEL-dKLA exhibited antifibrotic efficacy without damaging tissue-resident macrophages in the bleomycin-induced mouse model. Collectively, our findings suggest that MEL-dKLA may be a new therapeutic option for the treatment of idiopathic pulmonary fibrosis. • MEL-dKLA alleviates pulmonary fibrosis by targeting Fra-2 + M2 macrophages in a mouse model. • MEL-dKLA inhibits epithelial-mesenchymal and fibroblast-myofibroblast transitions. • MEL-dKLA reprograms the microenvironment into an anti-fibrotic state by eliminating M2 macrophages.
Blocking Microglial Proliferation by CSF-1R Inhibitor Does Not Alter the Neuroprotective Effects of Adoptive Regulatory T Cells in 3xTg Alzheimer’s Disease Mice Seon-Young Park, Nari Cha, Soyoung Kim, Songah Chae, Won-jun Lee, Hyunjae Jung, Hyunsu Bae Current Issues in Molecular Biology, 2024 Alzheimer’s disease (AD) is a chronic neurodegenerative disease that causes cognitive impairment. Neuroinflammation induced by activated microglia exacerbates AD. Regulatory T cells (Tregs) play roles in limiting neuroinflammation by converting microglial polarization. Therefore, adoptive Treg therapy is considered an attractive option for neurodegenerative disorders. However, the mechanism underlying Treg therapy via microglial modulation is not fully understood. In this study, we sought to determine whether adoptively transferred Tregs were effective when microglia proliferation was inhibited by using GW2580, which is an inhibitor of CSF1R. We found that inhibition of microglial proliferation during Treg transfer did not alter the therapeutic effects of Tregs on cognitive deficits and the accumulation of Aβ and pTAU in 3xTg-AD mice. The expression of pro- and anti-inflammatory markers in the hippocampus of 3xTg mice showed that GW2580 did not affect the inhibition of neuroinflammation by Treg transfer. Additionally, adoptively transferred Tregs were commonly detected in the brain on day 7 after transfer and their levels decreased slowly over 100 days. Our findings suggest that adoptively transferred Tregs can survive longer than 100 days in the brain, suppressing microglial activation and thus alleviating AD pathology. The present study provides valuable evidence to support the prolonged efficacy of adoptive Treg therapy in AD.
Therapeutic Effects of Aβ-Specific Regulatory T Cells in Alzheimer’s Disease: A Study in 5xFAD Mice Seon-Young Park, Juwon Yang, Hyejin Yang, Inhee Cho, Jae Yoon Kim, Hyunsu Bae International Journal of Molecular Sciences, 2024 The aging global population is placing an increasing burden on healthcare systems, and the social impact of Alzheimer’s disease (AD) is on the rise. However, the availability of safe and effective treatments for AD remains limited. Adoptive Treg therapy has been explored for treating neurodegenerative diseases, including AD. To facilitate the clinical application of Treg therapy, we developed a Treg preparation protocol and highlighted the therapeutic effects of Tregs in 5xFAD mice. CD4+CD25+ Tregs, isolated after Aβ stimulation and expanded using a G-rex plate with a gas-permeable membrane, were adoptively transferred into 5xFAD mice. Behavioral analysis was conducted using Y-maze and passive avoidance tests. Additionally, we measured levels of Aβ, phosphorylated tau (pTAU), and nitric oxide synthase 2 (NOS2) in the hippocampus. Real-time RT-PCR was employed to assess the mRNA levels of pro- and anti-inflammatory markers. Our findings indicate that Aβ-specific Tregs not only improved cognitive function but also reduced Aβ and pTAU accumulation in the hippocampus of 5xFAD mice. They also inhibited microglial neuroinflammation. These effects were observed at doses as low as 1.5 × 103 cells/head. Collectively, our results demonstrate that Aβ-specific Tregs can mitigate AD pathology in 5xFAD mice.
Alpha-Synuclein-Specific Regulatory T Cells Ameliorate Parkinson’s Disease Progression in Mice Seon-Young Park, HyeJin Yang, Soyoung Kim, Juwon Yang, Hyemin Go, Hyunsu Bae International Journal of Molecular Sciences, 2023 Parkinson’s disease (PD) is a long-term neurodegenerative disease characterized by dopaminergic neuronal loss and the aggregation of alpha-synuclein (α-syn) in the brain. Cell therapy using regulatory T cells (Tregs) has therapeutic potential on PD progression in a mouse model; however, several challenges were associated with its applications. Here, we propose a strategy for α-syn specific Treg expansion (α-syn Treg). We presented α-syn to T cells via dendritic cells. This method increased the mobility of Tregs towards the site of abundant α-syn in vitro (p < 0.01; α-syn Tregs versus polyclonal Tregs (poly Tregs)) and in vivo. Consequently, α-syn Tregs showed noteworthy neuroprotective effects against motor function deficits (p < 0.05, p < 0.01; α-syn Tregs versus poly Tregs), dopaminergic neuronal loss (p < 0.001; α-syn Tregs versus poly Tregs), and α-syn accumulation (p < 0.05; α-syn Tregs versus poly Tregs) in MPTP-induced PD mice. Furthermore, the adoptive transfer of α-syn Tregs exerted immunosuppressive effects on activated microglia, especially pro-inflammatory microglia, in PD mice. Our findings suggest that α-syn presentation may provide a significant improvement in neuroprotective activities of Tregs and suggest the effective clinical application of Treg therapy in PD.
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