Antiviral effect in human cytomegalovirus-infected cells, pharmacokinetics, and intravitreal toxicology in rabbits of acyclovir diphosphate dimyristoylglycerol

Abstract

Acyclovir diphosphate dimyristoylglycerol (ACVDP-DG) is a lipid prodrug which is active against ACVresistant strains of herpes simplex virus because of its intracellular metabolism to ACV monophosphate. In human cytomegalovirus (HCMV)-infected MRC-5 cells, ACVDP-DG was ninefold more active than ACV. When liposomal [8-3 H]ACVDP-DG was injected intravitreally at the maximum nontoxic dose of 1 mol in rabbits, the drug remained above its estimated 90% HCMV-inhibitory concentration for 18 days. Intravitreal ganciclovir persists above its 90% inhibitory concentration for only 1 to 2 days. ACVDP-DG may be useful as a local treatment for HCMV retinitis.Human cytomegalovirus (HCMV) retinitis occurs in 15 to 42% of patients with AIDS (9,10,12,14,17). Two antiviral drugs currently available for the treatment of HCMV retinitis are ganciclovir (GCV) and foscarnet, both of which are associated with significant toxicities (5) and the emergence of resistance (4, 16). Intravitreal injections with GCV have been used in patients not tolerating systemic therapy (11,23). The main disadvantage of this local therapy is the need for four to six injections per eye per month (1,8,13,21).Acyclovir diphosphate dimyristoylglycerol (ACVDP-DG) is an analog of CDP diacylglycerol and is very active in herpes simplex virus type 1-and type 2-infected cells in vitro (15). ACVDP-DG is metabolized to ACV monophosphate intracellularly and retains substantial in vitro antiviral activity in ACVresistant thymidine kinase-deficient mutants of herpes simplex virus type 1 and type 2 because of its unique metabolism (15). In this paper, we report the antiviral activity of ACVDP-DG in HCMV-infected MCR-5 cells and the ocular toxicity and pharmacokinetics after intravitreal injections of liposomal ACVDP-DG in rabbit eyes.Human embryonic lung fibroblast cells (MRC-5) and HCMV (strain AD169) were obtained from the American Type Culture Collection (Rockville, Md.). Virus stocks were prepared by infecting subconfluent MRC-5 cells; the titers of HCMV in the cell-free supernatant were determined, and aliquots were stored in liquid nitrogen. Subconfluent MRC-5 cells in 24-well culture dishes were pretreated for 24 h with drugs in minimal essential medium containing 2% fetal bovine serum and antibiotics. The medium was removed, and virus was added, absorbed for 1 h at 37ЊC, aspirated, and replaced with the drug dilutions. After 5 days of incubation, HCMV DNA was quantified in triplicate by nucleic acid hybridization with an HCMV antiviral susceptibility test kit from Diagnostic Hybrids, Inc. (Athens, Ohio). Results are expressed as percentages of the amount of HCMV-specific DNA in untreated HCMV-infected control cells.The concentration of ACVDP-DG which inhibited the replication of the AD169 strain of HCMV in MRC-5 cells by 50% (IC 50 ) was at 2.4 M, compared with IC 50 s of 22 and 1.1 M for ACV and GCV, respectively (Fig. 1). The concentration of GCV required to reduce HCMV DNA by 90% was 28 M, compared with 36 M for ACVDP-DG and 316 M for ACV. The 50% toxic dose of...

Synthesis of Nucleoside Phosphate and Phosphonate Prodrugs

“…The coupling of a morpholidate activated phosphatidic acid was also used for the preparation of a myristoyl glyceride DP derivative of acyclovir . This compound (not shown) was found to be active on ACV resistant herpes TK – , indicating an efficient delivery of ACV-MP …”

“…268b This compound (not shown) was found to be active on ACV resistant herpes TK – , indicating an efficient delivery of ACV-MP. 270 …”

Ganciclovir Release Rates in Vitreous from Different Formulations of 1-O-Hexadecylpropanediol-3-Phospho-Ganciclovir

“…In this way, the drug can be slowly released into vitreous or directly delivered into retinal cells, as the liposome membrane fuses with retinal cell membranes and cellular metabolism converts the lipid conjugate to active drug. Drug that is incorporated into the cell membrane can serve as a secondary drug depot to further extend the drug vitreous half-life [11]. Our results demonstrated a longer vitreous half-life and better vitreous clarity compared with traditional liposomes [6].…”

Nano and microtechnologies for ophthalmic administration, an overview

“…Related to intraocular route, liposomes have demonstrated to protect and increase the half-life of fragile active substances avoiding repetitive administrations to achieve therapeutic levels in the site of action [21,22]. In the case of intravitreal administration, liposomes have demonstrated to be useful in relatively long-term release of lipidic pro-drugs [23][24][25]. Liposomes have been already used in the ophthalmic clinical practice [26].…”