Champney K J, Lauter C B, Bailey E J, Lerner A M
J Clin Invest. 1978 Dec;62(6):1142-53. doi: 10.1172/JCI109233.
The minimum inhibitory concentration (MIC) of adenine arabinoside (ara-A) in rabbit kidney microtiter tissue cultures (RK-13) to a prototype strain of herpes simplex virus, type 1 (E115) based upon inhibition of cytopathic effects is 1.5 mug/ml. In this system, the MIC of arabinosylhypoxanthine (ara-Hx), the major in vivo metabolic derivative of ara-A, is 75 mug/ml. Inhibition of cytopathic effects of herpes simplex virus, type 1 (HSV-1) in microtiter wells of RK-13 cells varies directly with the concentrations of ara-A or ara-Hx, and inversely with residual HSV-1. The MIC of ara-A for HSV-1 in RK-13 cells is 5-20 times lower than similar measures with vero renal, mouse embryo, or human foreskin cultures. With RK-13 tissue cultures in microtiter plates, an assay for "ara-A equivalents" in human body fluids was developed which compares in sensitivity with high pressure liquid chromatography and has the advantage of simultaneously measuring combined antiherpesvirus effects of ara-A and its major metabolic derivative, ara-Hx. In vitro checkerboard studies in RK-13 cells confirmed that ara-A and ara-Hx in combination had antiviral effects which are synergistic. The total of the fractional MIC of ara-A plus ara-Hx in combination also varies inversely with residual HSV-1 in microtiter wells. Because virus adsorption is complete at 2 h before specimens to be tested are added in this assay, and because human interferon is not measured in rabbit cells, the antiviral assay is not affected by the presence of type-specific antiherpesvirus antibody or human interferon.Antiviral activity (AVA) was assayed as ara-A equivalents in sera and urines from 10 patients with serious herpesvirus infections who received 2.5-20 mg/kg daily of ara-A by intramuscular or intravenous routes. When a dosage schedule of 10 mg/kg per day or more was used, sustained concentrations of AVA that ranged from 0.8 to 14.4 mug/ml were found. When an inhibitor of adenosine deaminase (covidarabine) was not added to the specimens, mean serum concentrations were congruent with3.0 mug/ml (10 mg/kg per day, i.v.), and 4.1 mug/ml (20 mg/kg per day). However, in a single patient given 20 mg/kg of ara-A daily with covidarabine immediately added to the sera, the mean concentration of AVA was 12.9 mug/ml. Urines contained even higher AVA. Assays of 19 sera were performed both by microbiologic assay for AVA and by high pressure liquid chromatography for ara-A and ara-Hx. AVA was greater by microbiologic assay, and was greater than that which could be accounted for by stoichiometric chromatographic measures of ara-A and ara-Hx. These results with sera of treated patients are consistent both with the in vitro synergy of ara-A and ara-Hx found by checkerboard titrations, and with the beneficial responses to ara-A of patients with herpesvirus infections reported here and elsewhere.
基于对细胞病变效应的抑制作用,阿糖腺苷(ara - A)在兔肾微量滴定组织培养(RK - 13)中对单纯疱疹病毒1型原型株(E115)的最低抑菌浓度(MIC)为1.5微克/毫升。在该系统中,阿糖腺苷的主要体内代谢衍生物阿糖次黄嘌呤(ara - Hx)的MIC为75微克/毫升。RK - 13细胞微量滴定孔中单纯疱疹病毒1型(HSV - 1)的细胞病变效应抑制作用与ara - A或ara - Hx的浓度直接相关,与残余的HSV - 1呈反比。RK - 13细胞中ara - A对HSV - 1的MIC比在非洲绿猴肾、小鼠胚胎或人包皮培养物中的类似测定结果低5至20倍。利用微量滴定板中的RK - 13组织培养,开发了一种用于检测人体体液中“阿糖腺苷当量”的方法,其灵敏度与高压液相色谱法相当,且具有同时测量ara - A及其主要代谢衍生物ara - Hx联合抗疱疹病毒效应的优势。在RK - 13细胞中进行的体外棋盘格研究证实,ara - A和ara - Hx联合具有协同抗病毒作用。ara - A加ara - Hx联合的分数MIC总和也与微量滴定孔中残余的HSV - 1呈反比。由于在该测定中,在加入待测标本前2小时病毒吸附已完成,且兔细胞中不检测人干扰素,所以抗病毒测定不受型特异性抗疱疹病毒抗体或人干扰素存在的影响。对10例严重疱疹病毒感染患者进行了抗病毒活性(AVA)检测,这些患者通过肌肉注射或静脉注射途径每日接受2.5 - 20毫克/千克的ara - A。当采用每日10毫克/千克或更高的给药方案时,发现AVA的持续浓度范围为0.8至14.4微克/毫升。当未向标本中添加腺苷脱氨酶抑制剂(阿糖胞苷)时,平均血清浓度分别为3.0微克/毫升(每日静脉注射10毫克/千克)和4.1微克/毫升(每日静脉注射20毫克/千克)。然而,在一名每日给予20毫克/千克ara - A且立即向血清中添加阿糖胞苷的患者中,AVA的平均浓度为12.9微克/毫升。尿液中AVA含量甚至更高。对19份血清进行了AVA的微生物学测定以及ara - A和ara - Hx的高压液相色谱测定。微生物学测定的AVA更高,且高于ara - A和ara - Hx的化学计量色谱测量所能解释的值。这些治疗患者血清的结果与棋盘格滴定法发现的ara - A和ara - Hx体外协同作用一致,也与本文及其他地方报道的疱疹病毒感染患者对ara - A的有益反应一致。