Feitelson Mark A, Clayton Marcia M, Sun Bill, Schinazi Raymond F
Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, PA, USA.
Antivir Chem Chemother. 2007;18(4):213-23. doi: 10.1177/095632020701800405.
Woodchuck hepatitis virus (WHV)-infected woodchucks have been used for preclinical development of drugs against hepatitis B virus (HBV). However, there is no simple in vivo model to evaluate small amounts of compounds against HBV. To develop such a model, HepAD38 cells, in which HBV replication is regulated by tetracycline (tet), were grown as subcutaneous tumours in nude mice. Mice developing viraemia were then left untreated or given tet in the drinking water. In some of the mice given tet, it was removed and the mice were injected intraperitoneally with phosphate buffer saline (PBS), lamivudine (3TC), clevudine (CLV) or tenofovir dipivoxil fumarate (TDF). Virus DNA titres were measured by real-time PCR during and after drug treatment. In water-fed and PBS-injected mice, virus titres reached approximately 10(9) copies/ml serum within 35 days of HepAD38 injection, whereas in tet-treated mice, virus titres remained at 10(4)-10(5) copies/ml. HBV DNA levels were suppressed by 3TC, TDF and CLV, with the latter two drugs showing more sustained virus suppression compared with 3TC. Combination therapy with CLV plus TDF was much more effective than either drug alone in suppressing virus titre for at least 3 weeks after the end of treatment. There was no demonstrable toxicity to HepAD38 cells in drug-treated mice. Hence, a robust tet-controlled system for HBV replication in vivo was demonstrated, validated with monotherapies against HBV and shown to be useful in assessing combination therapy. This system will be useful for preclinical assessment of small amounts of single or multiple compounds against HBV in vivo.
感染土拨鼠肝炎病毒（WHV）的土拨鼠已被用于抗乙型肝炎病毒（HBV）药物的临床前开发。然而，目前尚无简单的体内模型来评估针对HBV的少量化合物。为了建立这样一个模型，将HBV复制受四环素（tet）调控的HepAD38细胞作为皮下肿瘤在裸鼠体内生长。然后，让出现病毒血症的小鼠不接受治疗或饮用含tet的水。在一些饮用含tet水的小鼠中，停止给予tet，并给小鼠腹腔注射磷酸盐缓冲盐水（PBS）、拉米夫定（3TC）、氯夫定（CLV）或替诺福韦酯富马酸盐（TDF）。在药物治疗期间及之后，通过实时PCR测量病毒DNA滴度。在饮用普通水和注射PBS的小鼠中，注射HepAD38细胞后35天内病毒滴度达到约10⁹拷贝/ml血清，而在接受tet治疗的小鼠中，病毒滴度维持在10⁴ - 10⁵拷贝/ml。3TC、TDF和CLV均可抑制HBV DNA水平，与3TC相比，后两种药物对病毒的抑制作用更持久。在治疗结束后至少3周内，CLV加TDF联合治疗在抑制病毒滴度方面比单独使用任何一种药物都有效得多。药物治疗的小鼠中未发现对HepAD38细胞有明显毒性。因此，证明了一种强大的体内HBV复制tet调控系统，通过针对HBV的单一疗法进行了验证，并显示可用于评估联合治疗。该系统将有助于在体内对针对HBV的少量单一或多种化合物进行临床前评估。
