Department of Gastroenterology, Hepatology and Endocrinology, Medical School Hannover, Carl Neuberg Strasse 1, 25 Hannover, Germany.
Gut. 2010 Oct;59(10):1416-26. doi: 10.1136/gut.2009.196519. Epub 2010 Jul 30.
Viral infection of a dying cell dictates the immune response against intracellular antigens, suggesting that virotherapy may be an effective tool to induce immunogenic cell death during systemic cancer treatment. Since viruses and proteasome inhibitors both induce accumulation of misfolded proteins, endoplasmic reticulum (ER) stress and immune responses during treatment of hepatocellular carcinoma (HCC) with bortezomib and the tumour-specifically replicating virus hTert-Ad (human telomerase reverse transcriptase promoter-regulated adenovirus) were investigated.
Unfolded protein response (UPR) pathways and ER stress-mediated apoptosis were investigated by western blots, caspase-3 assays, 4',6-diamidino-2-phenylindole (DAPI) and Annexin V staining in HCC cells following hTert-Ad/bortezomib treatment. Oncolysis was assessed in subcutaneous HCC mouse models. Antiviral/antitumoural immune responses were characterised in immunocompetent HCC mouse models by ELISA, ELISpot assays and pentamer staining. Systemic efficacy of antitumoural immunity was investigated by determination of lung metastases burden.
Bortezomib and hTert-Ad trigger complementary UPR pathways but negatively interfere with important recovery checkpoints, resulting in enhanced apoptosis of HCC cells in vitro and improved oncolysis in vivo. In immunocompetent mice, bortezomib inhibited antiviral immune responses, whereas ER stress-induced apoptosis of infected HCC resulted in caspase-dependent triggering of antitumoural immunity. In therapeutic settings in immunocompetent, but not in immunodeficient or CD8-depleted mice, virotherapy-induced antitumoural immunity efficiently inhibited outgrowth of non-infected lung metastases. Immunotherapeutic efficacy could be significantly improved by bortezomib in experiments with low viral doses.
Proteasome inhibition during virotherapy disrupts the UPR, leading to enhanced ER stress-induced apoptosis, improved local oncolysis and antitumoural immunity. The results suggest that combining intratumoural virotherapy with adjuvant systemic therapies, which specifically support the function of the virotherapy as an antitumoural vaccine, is a promising immunotherapeutic strategy against HCC.
垂死细胞中的病毒感染决定了针对细胞内抗原的免疫反应，这表明病毒疗法可能是在全身癌症治疗中诱导免疫原性细胞死亡的有效工具。由于病毒和蛋白酶体抑制剂在治疗肝细胞癌（HCC）时都会诱导错误折叠蛋白的积累、内质网（ER）应激和免疫反应，因此研究了硼替佐米和肿瘤特异性复制病毒 hTert-Ad（人类端粒酶逆转录酶启动子调控的腺病毒）联合治疗时的未折叠蛋白反应（UPR）途径和 ER 应激介导的细胞凋亡。
Western blot、caspase-3 测定、4'，6-二脒基-2-苯基吲哚（DAPI）和 Annexin V 染色检测 hTert-Ad/硼替佐米处理后 HCC 细胞中的 UPR 途径和 ER 应激介导的细胞凋亡。皮下 HCC 小鼠模型评估溶瘤作用。通过 ELISA、ELISpot 测定和五聚体染色在免疫活性 HCC 小鼠模型中表征抗病毒/抗肿瘤免疫反应。通过测定肺转移瘤负担研究抗肿瘤免疫的系统疗效。
硼替佐米和 hTert-Ad 触发互补的 UPR 途径，但会相互干扰重要的恢复检查点，导致 HCC 细胞在体外凋亡增强，体内溶瘤作用改善。在免疫活性小鼠中，硼替佐米抑制抗病毒免疫反应，而感染的 HCC 细胞中 ER 应激诱导的细胞凋亡导致 caspase 依赖性抗肿瘤免疫反应的触发。在免疫活性的、而非免疫缺陷的或 CD8 耗竭的小鼠中，病毒治疗诱导的抗肿瘤免疫在治疗设置中有效抑制未感染的肺转移瘤的生长。在低病毒剂量的实验中，硼替佐米可显著提高免疫治疗的疗效。
病毒治疗期间的蛋白酶体抑制会破坏 UPR，导致增强的 ER 应激诱导的细胞凋亡、改善局部溶瘤作用和抗肿瘤免疫。结果表明，将肿瘤内病毒治疗与辅助全身治疗相结合，特别是支持病毒治疗作为抗肿瘤疫苗的功能，是一种有前途的 HCC 免疫治疗策略。
