van Leeuwen E B M, Cloosen S, Senden-Gijsbers B L M G, Germeraad W T V, Bos G M J
Department of Internal Medicine, Division of Hemato-Oncology, University Hospital Maastricht, Maastricht, The Netherlands.
Cytotherapy. 2006;8(1):36-46. doi: 10.1080/14653240500508166.
DC-presenting tumor Ag are currently being developed to be used as a vaccine in human cancer immunotherapy. To increase the chances for successful therapy it is important to deliver full-length tumor Ag instead of loading single peptides. Methodologically, several recombinant DNA delivery techniques have been used.
In this study we compared nucleofection, an optimized form of electroporation, and adenoviral transduction regarding their efficiency to transduce human monocyte-derived (Mo-) DC in vitro. Expression of the tumor-associated Ag mucin-1 (MUC1) after adenoviral transduction (rAd5Fib35-MUC1) was determined using two MAb.
We showed that the viability of cells and percentage of green fluorescent protein (GFP)-positive cells after transduction with a fiber-modified adenoviral vector (rAd5F35-GFP) was much higher than after nucleofection. Furthermore, phenotype and function of DC were not impaired by infection with adenovirus particles. Cells matured normally; up-regulation of CD40, CD80, CD83, CD86 and HLA-DR was not affected by adenoviral transduction. The capacity to stimulate naive T-cell proliferation was preserved and no change in IL-10 production was observed. Production of IL-12 increased up to 500-fold upon adenoviral transduction, considered to contribute positively to an anti-tumor immune response. Non-transduced mature DC expressed low levels of endogenous MUC1. After transduction with the rAd5F35-MUC1 adenoviral vector, a 100-fold increase in MUC1 expression by DC was observed.
The use of the fiber-modified adenoviral vector presented here may therefore be favorable compared with non-viral gene delivery systems for DC that will be used in cancer immunotherapy.
目前正在研发递呈肿瘤抗原的树突状细胞（DC），以用作人类癌症免疫治疗中的疫苗。为提高成功治疗的几率，递送全长肿瘤抗原而非加载单个肽段很重要。在方法上，已使用了多种重组DNA递送技术。
在本研究中，我们比较了核转染（一种优化形式的电穿孔）和腺病毒转导在体外转导人单核细胞来源的（Mo-）DC的效率。使用两种单克隆抗体测定腺病毒转导（rAd5Fib35-MUC1）后肿瘤相关抗原粘蛋白-1（MUC1）的表达。
我们发现，用纤维修饰的腺病毒载体（rAd5F35-GFP）转导后，细胞活力和绿色荧光蛋白（GFP）阳性细胞百分比远高于核转染后。此外，腺病毒颗粒感染不会损害DC的表型和功能。细胞正常成熟；CD40、CD80、CD83、CD86和HLA-DR的上调不受腺病毒转导的影响。刺激初始T细胞增殖的能力得以保留，且未观察到白细胞介素-10产生的变化。腺病毒转导后白细胞介素-12的产生增加了多达500倍，被认为对抗肿瘤免疫反应有积极贡献。未转导的成熟DC表达低水平的内源性MUC1。用rAd5F35-MUC1腺病毒载体转导后，观察到DC的MUC1表达增加了100倍。
因此，与将用于癌症免疫治疗的DC的非病毒基因递送系统相比，本文所使用的纤维修饰腺病毒载体可能更具优势。
