PO.IM01.01 · 免疫学
SRC3-KO调节性T细胞在小鼠乳腺癌模型中的临床前研究
Preclinic study of src3-ko regulatory T cells in mouse breast cancer model
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
引言：调节性T细胞（Treg）通过其免疫抑制活性在平衡免疫应答以预防自身免疫性疾病中发挥关键作用。然而，Treg的抑制活性可被癌症利用以逃避免疫监视，从而导致肿瘤进展。类固醇受体共激活因子3（SRC-3）在Treg中高表达，我们已发现基因工程改造的、在Treg中特异性敲除SRC-3的小鼠对乳腺癌生长具有抵抗力，提示SRC-3-KO Treg是乳腺癌患者的一种潜在疗法。在本研究中，我们探讨了SRC-3 KO Treg在三阴性乳腺癌（TNBC）小鼠模型中的治疗效果。
方法：通过将表达荧光素酶的E0771 TNBC细胞注射到第四乳腺垫，建立了小鼠同基因、免疫完整的乳腺癌模型。为尽可能模拟临床情景，从荷瘤C57BL/6J供体小鼠的脾脏中分离Treg细胞。通过CRISPR/Cas9靶向破坏SRC-3基因后，让Treg在体外恢复并增殖4天，然后收集用于过继性细胞治疗（ACT），转移至已建立乳腺癌的C57BL/6J受体小鼠。未经基因编辑的野生型Treg细胞用作对照。通过生物发光成像每周两次监测肿瘤生长，持续3周。实验结束时，处死小鼠并收集脾脏、血液和肿瘤用于流式细胞术分析以及免疫荧光染色。
结果：SRC-3 KO Treg的ACT在所有接受治疗的小鼠中显著减少了肿瘤生长或清除了肿瘤。SRC-3KO Treg治疗还延长了小鼠的总生存期。在受体小鼠的脾脏、血液和肿瘤中均检测到经ACT的WT和SRC-3KO Treg。此外，与WT Treg治疗的对照组相比，SRC-3-KO Treg治疗的荷瘤小鼠中发现了多得多的细胞毒性T细胞，包括CD8和granzyme B+细胞。
结论：在来自荷瘤供体的Treg中删除SRC-3基因，在免疫完整的小鼠乳腺癌模型中具有强效的抗肿瘤作用。这些SRC-3 KO Treg通过增强肿瘤微环境中效应性CD8+ T细胞的细胞毒性来发挥其作用。我们的结果强化了使用SRC-3 KO Treg细胞作为TNBC有效免疫治疗的临床潜力。
查看英文原文 English abstract
Introduction: Regulatory T cells (Tregs) play a critical role in balancing immune responses to prevent autoimmune disease through their immunosuppressive activities. However, the suppressive activity of Tregs can be coopted by cancers to evade immune surveillance, leading to tumor progression. Steroid receptor coactivator 3 (SRC-3) is highly expressed in Tregs and we have discovered that genentically engineered mice with SRC-3 specifically knocked out in Tregs are resistant to breast cancer growth, suggesting SRC-3-KO Tregs are a potential therapy for breast cancer patients. In this study, we explored the therapeutic effect of SRC-3 KO Tregs in a mouse model of triple negative breast cancer (TNBC).
Method: A mouse syngeneic, immune-intact breast cancer model was produced by injecting luciferase expressing E0771 TNBC cells into the fourth mammary pad. To closely mimic a clinical scenario, Treg cells were isolated from the spleens of tumor-bearing C57BL/6J donor mice. After disrupting the SRC-3 gene by CRISPR/Cas9 targeting, Tregs were allowed to recover and proliferate in vitro for 4 days before collecting them for adoptive cell therapy (ACT) into C57BL/6J recipient mice with established breast cancer. Wild type Treg cells without gene editing were used as control. Tumor growth was monitored by bioluminescence imaging twice a week for 3 weeks. By the end of experiment, mice were sacrificed and spleen, blood, and tumors were collected for flow cytometry analysis as well as immunofluorescence staining.
Results: ACT of SRC-3 KO Tregs significantly reduced tumor growth or cleared tumosr in all treated mice. SRC-3KO Treg treatment also prolonged mice overall survival. Both ACTed WT and SRC-3KO Tregs were detected in the spleen, blood, and tumors of recipient mice. Moreover, many more cytotoxic T cells, including CD8 and granzyme B+ cells were found in SRC-3-KO Treg treated tumor-bearing mice, compared to WT Treg treated controls.
Conclusion: Deleting the SRC-3 gene in Tregs derived from tumor-bearing donors possess potent anti-tumor effects in an immune-intact mouse breast cancer model. These SRC-3 KO Tregs exert their effects by enhancing the cytotoxicity of effector CD8+ T cells in the tumor microenvironment. Our results reinforce the clinical potential of using SRC-3 KO Treg cells as an effective immunotherapy for TNBCs.
利益披露 Disclosure
Y. Xia,
Coregen Independent Contractor, Stock Option, ).
Y. Gilad,
Coregen Independent Contractor, Stock Option, ).
D. M. Lonard,
Coregen g., Board of Directors, non-salaried role), Stock, ), Travel, Patent.