Longterm gene therapy of xeroderma pigmentosum group A mice using HVJ-liposomes.

着色性干皮病（XP）是一种常染色体不可避免的遗传性疾病，其核苷酸切除修复缺陷，因此XP患者引起日晒区皮肤癌的频率很高。在XP补体组中，XP补体A组（XPA）表现出最严重的临床表型，伴有神经异常，XPA是日本XPA患者中最常见的类型。本项目的目的是以hvj脂质体的形式治疗人XPA基因，挑战XPA模型小鼠体内的基因治疗，因为我们的研究目标之一是建立XPA患者的基因治疗。转染含有人XPA基因的ppcaggs -XPA或pcHA-XPA两个质粒，以血凝素基因为标记，在UVC照射24或48小时后通过非计划DNA合成测量其修复能力。两者都恢复了XPA小鼠细胞的修复效率。用日本血凝病毒（HVJ）脂质体将含人XPA基因的质粒导入小鼠皮肤。hvj -脂质体- xpa在4 kJ/m^2 UVB照射前24小时皮下注射或在UVB照射前48小时局部涂漆。注射XPA-HVJ脂质体小鼠表皮核的非计划性DNA合成高于对照脂质体。接下来，我们在UVB照射前24小时或48小时用或不使用hvj - xpa脂质体，每周1-2次用500 J/m^2的UVB照射小鼠，持续3个月。包封在hvj脂质体中的pcHA-XPA和pCAGGS-XPA均能减小紫外线诱导肿瘤的大小和数量，延长生存时间。然而，两种治疗方法都不能完全抑制肿瘤的形成。肿瘤组织学为鳞状细胞癌。由此可见，HVJ-XPA脂质体对XPA模型小鼠具有降低紫外线诱导肿瘤的作用。

Xeroderma pigmentosum (XP) is an autosomalrecessibely inheritated disorder in which nucleotide excision repair is deficient, thus the patients with XP cause skin cancer of sun-exposed area in high frequency. Among XP complementation groups, XP complementation group A (XPA) reveals the severest clinical phenotype with neurological abnormalities and XPA is the most common type among Japanese XPA patients. The aim of this project is to challenge the gene therapy of XPA model mice in vivo with treating human XPA gene in the form of HVJ-liposomes, since one of our research goal is to establish the gene therapy for XPA patients. Two plasmids containing human XPA gene, pCAGGS-XPA or pcHA-XPA, which contains hemagultinin gene as a tag, was transfected and the repair ability was measured by unscheduled DNA syntshesis at post 24 or 48 hr UVC irradiation. Both recovered the repair efficiency in XPA mice cells. In in vivo study HVJ (Hemagluutinating Virus of Japan)-liposomes was used for introducing the plasmid containing human XPA gene into mice skin. HVJ-liposomes-XPA was injected intradermaly 24 hr before 4 kJ/m^2 of UVB irradiation or topically painted 48 hr before UVB irradiation. Unscheduled DNA synthesis of epidermal nuclei in mice injected XPA-HVJ liposmes was higher than that of control liposomes. Next we irradiated the mice with 500 J/m^2 of UVB 1-2 times per week for 3 months with or without treating HVJ-XPA-liposmes 24 or 48 hr prior to UVB irradiation. Both pcHA-XPA and pCAGGS-XPA encapsulated in HVJ-liposmes reduced the size, the number of the UV-induced tumors and elongate the survival duration. However, both treatment did not completely inhibit the tumor formation. The histology of the tumor was squamous cell carcinoma. In conclusion, the treatment of HVJ-XPA liposomes has effect on reducing the UV-induced tumors in XPA model mice.