Sun Bo, Liu Jiayun, Liu Xiaocui, Li Jing, Zhang Guilin, Sun Tao, Zheng Chuansheng, Kan Xuefeng
Department of Radiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
Hubei Provincial Clinical Research Center for Precision Radiology & Interventional Medicine, Wuhan, China.
Cell Death Dis. 2025 Apr 13;16(1):288. doi: 10.1038/s41419-025-07622-z.
Preventing tumor recurrence after radiofrequency ablation (RFA) of malignant solid tumors with large size or in high-risk locations represents a great challenge. In this study, we explored the feasibility of using oncolytic peptide LTX-315 plus an anti-cytotoxic T lymphocyte antigen-4 (CTLA-4) antibody for inhibiting residual tumors after RFA of hepatocellular carcinoma (HCC). In in vitro experiment, the CD8T cells from Hepa1-6 tumors, after being subjected to three different treatments (control, iRFA, iRFA + LTX-315), were extracted and were then co-cultured with Hepa1-6 cells and an anti-CTLA-4 antibody. The enzyme-linked immunospot, flow cytometry, and cell counting kit-8 assay were employed to assess the cytotoxicity of extracted CD8T cells on Hepa1-6 cells. In in vivo experiment, different murine orthotopic HCC models were variously treated by: (1) pseudo iRFA + phosphate-buffered saline (PBS); (2) iRFA + PBS; (3) iRFA + LTX-315; (4) iRFA + anti-CTLA-4 antibody; and (5) iRFA + LTX-315 + anti-CTLA-4 antibody. The treatment effects were compared among different groups and were pathologically confirmed. The possible mechanisms of the combination treatment (LTX-315+anti-CTLA-4 antibody) for residual tumors after iRFA of HCC were explored. LTX-315 significantly reduced the PD-1 expression and significantly increased CTLA-4 expression of CD8T cells in residual tumors, and additional treatment of anti-CTLA-4 antibody could significantly enhance the cytotoxicity of CD8T cells for Hepa1-6 cells in vitro experiments. Compared with the other treatments, the combined treatment of LTX-315 with anti-CTLA-4 antibody achieved a better tumor response and longer survival, and it could synergistically activate the cGAS-STING pathway and elicit an immunogenic cell death, leading to a strong anti-tumor immunity after iRFA of HCC. The immunosuppressive microenvironment of residual tumors was significantly improved by the combination therapy with a significantly increased ratio of M1-like tumor-associated macrophages to M2-like tumor-associated macrophages, a significantly decreased infiltration of regulatory T cells and myeloid-derived suppressor cells, and a significantly lower expression of PD-1 and CTLA-4. Overall, the results of this study demonstrated that LTX-315 plus anti-CTLA-4 antibody could synergistically improve the immunosuppressive microenvironment of residual tumors and induce a strong anti-tumor immunity after iRFA of HCC. This combination treatment strategy may offer a new alternative to reduce the tumor recurrence after RFA of malignant solid tumors with large sizes or in high-risk locations.
对于大尺寸或高危部位的恶性实体瘤，预防射频消融（RFA）后肿瘤复发是一项巨大挑战。在本研究中，我们探讨了使用溶瘤肽LTX-315联合抗细胞毒性T淋巴细胞抗原4（CTLA-4）抗体抑制肝细胞癌（HCC）RFA后残留肿瘤的可行性。在体外实验中，提取经三种不同处理（对照、iRFA、iRFA + LTX-315）后的Hepa1-6肿瘤中的CD8T细胞，然后将其与Hepa1-6细胞和抗CTLA-4抗体共培养。采用酶联免疫斑点法、流式细胞术和细胞计数试剂盒-8法评估提取的CD8T细胞对Hepa1-6细胞的细胞毒性。在体内实验中，不同的小鼠原位HCC模型接受以下不同处理：（1）假iRFA + 磷酸盐缓冲盐水（PBS）；（2）iRFA + PBS；（3）iRFA + LTX-315；（4）iRFA + 抗CTLA-4抗体；（5）iRFA + LTX-315 + 抗CTLA-4抗体。比较不同组的治疗效果并进行病理确认。探讨了联合治疗（LTX-315 + 抗CTLA-4抗体）对HCC iRFA后残留肿瘤的可能机制。LTX-315显著降低残留肿瘤中CD8T细胞的PD-1表达并显著增加CTLA-4表达，在体外实验中，额外使用抗CTLA-4抗体可显著增强CD8T细胞对Hepa1-6细胞的细胞毒性。与其他治疗相比，LTX-315与抗CTLA-4抗体联合治疗取得了更好的肿瘤反应和更长的生存期，并且它可以协同激活cGAS-STING通路并引发免疫原性细胞死亡，从而在HCC iRFA后产生强大的抗肿瘤免疫力。联合治疗显著改善了残留肿瘤的免疫抑制微环境，M1样肿瘤相关巨噬细胞与M2样肿瘤相关巨噬细胞的比例显著增加，调节性T细胞和髓源性抑制细胞的浸润显著减少，PD-1和CTLA-4的表达显著降低。总体而言，本研究结果表明，LTX-315联合抗CTLA-4抗体可协同改善残留肿瘤的免疫抑制微环境，并在HCC iRFA后诱导强大的抗肿瘤免疫力。这种联合治疗策略可能为减少大尺寸或高危部位恶性实体瘤RFA后肿瘤复发提供一种新的选择。
