Aduma P, Connelly M C, Srinivas R V, Fridland A
Department of Infectious Diseases, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Mol Pharmacol. 1995 Apr;47(4):816-22.
The acyclic nucleoside phosphonates (S)-1-(3-hydroxy-2-phosphonylmethoxypropyl)cytosine (HPMPC), (S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine (HPMPA), and 9-(2-phosphonylmethoxyethyl)adenine (PMEA) inhibited herpes simplex virus-1 replication in Vero cells, and the IC50 values ranged from 4 microM (for HPMPC and HPMPA) to 40 microM (for PMEA). Pretreatment of cells with HPMPC for 12-24 hr induced an effective antiviral state, and the cells maintained this antiviral state for > 7 days. In contrast, much larger amounts (approximately 2.5-5 x IC50 doses) of PMEA or HPMPA were required to establish an antiviral state, which lasted for only approximately 24 or 72 hr, respectively. A 12-hr treatment of the cells with the phosphonates was required for the establishment of optimal antiviral activity; surprisingly, longer durations of exposure to PMEA (but not HPMPA or HPMPC) resulted in diminished antiviral effect. We investigated the metabolism of PMEA and HPMPC to determine the cellular basis for these differences. The cellular uptake of HPMPC was approximately 8-fold greater than that of PMEA. The levels of the PMEA metabolites PMEA monophosphate and PMEA diphosphate increased for approximately 12 hr and plateaued thereafter. PMEA and its metabolites were cleared from the cells with a half-life of 4.9 hr. In contrast, the HPMPC metabolites HPMPC monophosphate (HPMPCp) and HPMPC diphosphate (HPMPCpp) accumulated throughout the 24-hr study period and, at equimolar drug concentrations (25 microM), reached intracellular levels approximately 2-3-fold greater than those of the PMEA metabolites. HPMPC also differed from PMEA in its capacity to generate a phosphodiester metabolite (HMPCp-choline), which was a predominant metabolite in HPMPC-treated cells. In addition, the rates of disappearance of intracellular metabolites of the two drugs were significantly different. Thus, the decay of HPMPCpp was quite slow and biphasic (t1/2 = 24 and 65 hr) and that of HMPCp-choline was monophasic (t1/2 = 87 hr). Together, these factors can explain the differing antiviral potencies seen with PMEA and HPMPC. The possible role of the choline adduct in the expression of antiviral activity of the drug remains to be elucidated, but the adduct may serve as an intracellular store for the long term maintenance of active HPMPCpp in cells. The results also highlight the extent of diversity in the cellular pharmacology and antiviral activities of the acyclic nucleoside phosphonates.
无环核苷膦酸酯（S）-1-（3-羟基-2-膦酰甲氧基丙基）胞嘧啶（HPMPC）、（S）-9-（3-羟基-2-膦酰甲氧基丙基）腺嘌呤（HPMPA）和9-（2-膦酰甲氧基乙基）腺嘌呤（PMEA）可抑制单纯疱疹病毒1型在Vero细胞中的复制，其半数抑制浓度（IC50）值范围为4微摩尔/升（针对HPMPC和HPMPA）至40微摩尔/升（针对PMEA）。用HPMPC预处理细胞12至24小时可诱导有效的抗病毒状态，且细胞维持该抗病毒状态超过7天。相比之下，需要大量（约2.5至5倍IC50剂量）的PMEA或HPMPA才能建立抗病毒状态，且该状态分别仅持续约24小时或72小时。用膦酸酯处理细胞12小时是建立最佳抗病毒活性所必需的；令人惊讶的是，长时间暴露于PMEA（但不包括HPMPA或HPMPC）会导致抗病毒效果减弱。我们研究了PMEA和HPMPC的代谢，以确定这些差异的细胞基础。HPMPC的细胞摄取量比PMEA大约高8倍。PMEA的代谢产物单磷酸PMEA和二磷酸PMEA的水平在约12小时内升高，此后趋于平稳。PMEA及其代谢产物从细胞中清除的半衰期为4.9小时。相比之下，在整个24小时的研究期间，HPMPC的代谢产物单磷酸HPMPC（HPMPCp）和二磷酸HPMPC（HPMPCpp）不断积累，并且在等摩尔药物浓度（25微摩尔/升）下，其细胞内水平比PMEA代谢产物高约2至3倍。HPMPC在生成磷酸二酯代谢产物（HMPCp-胆碱）的能力方面也与PMEA不同，该产物是HPMPC处理细胞中的主要代谢产物。此外，两种药物细胞内代谢产物的消失速率存在显著差异。因此，HPMPCpp的衰减非常缓慢且呈双相性（半衰期分别为24小时和65小时），而HMPCp-胆碱的衰减呈单相性（半衰期为87小时）。这些因素共同可以解释PMEA和HPMPC抗病毒效力的差异。胆碱加合物在该药物抗病毒活性表达中的可能作用仍有待阐明，但该加合物可能作为细胞内储存库，用于细胞中活性HPMPCpp的长期维持。这些结果还突出了无环核苷膦酸酯在细胞药理学和抗病毒活性方面的多样性程度。
