University Medical Center of the Johannes Gutenberg University Mainz, 1st Department of Internal Medicine, Langenbeckstrasse, Mainz, Germany.
Onco Targets Ther. 2013 Aug 20;6:1119-27. doi: 10.2147/OTT.S49371. eCollection 2013.
Tumor-directed and immune-system-stimulating therapies are of special interest in cancer treatment. Here, we demonstrate the potential of parvovirus H-1 (H-1PV) to efficiently kill colorectal cancer cells and induce immunogenicity of colorectal tumors by inducing maturation of dendritic cells (DCs) alone and also in combination with cytostatic drugs in vitro. Using our cell culture model, we have additionally investigated the effects of anti-CTLA-4 (cytotoxic T-lymphocyte-associated antigen 4) receptor antibody tremelimumab on this process.
Colon carcinoma cell lines were treated with different concentrations of cytostatic drugs or tremelimumab or were infected with H-1PV in different multiplicities of infection (MOIs), and viability was determined using MTT assays. Expression of CTLA-4 in colon carcinoma cell lines was measured by FACScan™. For the coculture model, we isolated monocytes using adherence, and differentiation into immature DCs (iDCs) was stimulated using interleukin-4 and granulocyte-macrophage colony-stimulating factor. Maturation of iDCs into mature DCs (mDCs) was induced by a cytokine cocktail. SW480 colon carcinoma cells were infected with H-1PV or treated with cytostatic drugs. Drug treated and H-1PV-infected SW480 colon carcinoma cells were cocultured with iDCs and expression of maturation markers was measured using FACScan™. Cytokine measurements were performed using enzyme-linked immunosorbent assay.
Colon carcinoma cells SW480 were potently infected and killed by H-1PV. CTLA-4 expression in SW480 cells increased after infection with H-1PV and also after treatment with cytostatic drugs. Tremelimumab had no influence on viability of the colon carcinoma cell line. There was no maturation of iDCs after coculture with SW480; instead, H-1PV-infected or drug pretreated SW480 induced maturation. Cytokine production was higher for H-1PV-infected cells but was not significantly enhanced by tremelimumab treatment alone or in combination. Addition of tremelimumab did not interfere with the maturation process as measured by markers of maturation as well as by determination of cytokine levels.
By enhancing both cell death and immunogenicity of tumors, H-1PV is of special interest for tumor-directed therapy. These features make it a promising candidate for clinical application in human colorectal cancer. As tremelimumab does not significantly interfere with this process, an interesting therapeutic combination of active enhancement of tumor immunogenicity and independent masking of the CTLA-4 silencing process on tumor cells is highlighted.
在癌症治疗中，针对肿瘤的治疗和免疫系统刺激疗法具有特殊的意义。在这里，我们证明了细小病毒 H-1（H-1PV）通过单独诱导树突状细胞（DC）成熟，以及与细胞抑制剂联合使用，在体外有效杀死结直肠癌细胞并诱导结直肠肿瘤的免疫原性的潜力。使用我们的细胞培养模型，我们还研究了抗 CTLA-4（细胞毒性 T 淋巴细胞相关抗原 4）受体抗体 tremelimumab 对该过程的影响。
用不同浓度的细胞抑制剂或 tremelimumab 处理结肠癌细胞系，或用不同感染复数（MOI）感染 H-1PV，并用 MTT 法测定细胞活力。通过 FACScan™测量结肠癌细胞系中 CTLA-4 的表达。对于共培养模型，我们使用贴壁法分离单核细胞，并使用白细胞介素-4 和粒细胞-巨噬细胞集落刺激因子刺激其分化为未成熟的树突状细胞（iDC）。用细胞因子鸡尾酒诱导 iDC 成熟为成熟的树突状细胞（mDC）。用 H-1PV 感染 SW480 结肠癌细胞或用细胞抑制剂处理。用细胞抑制剂处理和 H-1PV 感染的 SW480 结肠癌细胞与 iDC 共培养，并通过 FACScan™测量成熟标志物的表达。用酶联免疫吸附试验（ELISA）测定细胞因子测量。
结肠癌细胞系 SW480 可被 H-1PV 有效感染和杀死。SW480 细胞感染 H-1PV 或用细胞抑制剂处理后 CTLA-4 表达增加。Tremelimumab 对结肠癌细胞系的活力没有影响。SW480 共培养后 iDC 无成熟；相反，H-1PV 感染或药物预处理的 SW480 诱导成熟。H-1PV 感染细胞的细胞因子产生更高，但单独用 tremelimumab 处理或联合处理并不显著增强。加入 tremelimumab 不会通过成熟标志物的测定和细胞因子水平的测定来干扰成熟过程。
通过增强肿瘤的细胞死亡和免疫原性，H-1PV 对肿瘤定向治疗具有特殊意义。这些特征使其成为人类结直肠癌临床应用的有前途的候选者。由于 tremelimumab 不会显著干扰该过程，因此突出了主动增强肿瘤免疫原性和独立掩盖肿瘤细胞 CTLA-4 沉默过程的有趣治疗组合。
