In vitro and in vivo activity of 1-O-hexadecylpropanediol-3-phospho-ganciclovir and 1-O-hexadecylpropanediol-3-phospho-penciclovir in cytomegalovirus and herpes simplex virus infections.

Human cytomegalovirus (HCMV) and herpes simplex virus (HSV) can cause a wide variety of clinical manifestations in man. Ganciclovir (GCV) is effective against HCMV infection when administered by the intravenous route and may be used orally in large doses for prophylaxis of HCMV infections in organ transplantation patients and in AIDS patients. In previous studies with acyclovir (ACV), we found that covalent attachment of an alkyl glycerol phosphate moiety greatly increased oral bioavailability and increased antiviral activity against hepatitis B virus. Adducts of ACV with alkyl propanediol phosphate were more active than the alkyl glycerol phosphate analogue in vitro in 2.2.15 cells, which constitutively produce hepatitis B virus. To see if this strategy would work for two other poorly absorbed nucleoside analogues, we synthesized 1-O-hexadecylpropanediol-3-phospho-GCV (HDP-P-GCV) and 1-O-hexadecyl-propanediol-3-phospho-penciclovir (HDP-P-PCV), and evaluated the in vitro antiviral activity, selectivity and oral antiviral activity of both compounds versus GCV or PCV in mice infected with HSV-1 or HDP-P-GCV versus murine cytomegalovirus (MCMV). HDP-P-GCV is orally active in both MCMV and HSV-1 infection in mice with antiviral activity equivalent to (HSV-1) or greater than oral GCV (MCMV). Oral HDP-P-PCV was more active than PCV orally versus intranasal HSV-1 infection in mice.

Intravitreal toxicology and duration of efficacy of a novel antiviral lipid prodrug of ganciclovir in liposome formulation.

PURPOSE: To evaluate the intraocular safety and antiviral treatment efficacy of the sustained lipid prodrug of ganciclovir, 1-O-hexadecylpropanediol-3-phospho-ganciclovir (HDP-P-GCV), as an intravitreal injectable drug system for viral retinitis. METHODS: HDP-P-GCV was synthesized by coupling 1-O-hexadecyl-propanediol-3-phosphate to either free hydroxyl of ganciclovir in pyridine with dicyclohexylcarbodiimide as catalyst. The compound was formulated into liposomes. The antiviral activity was assessed by DNA reduction in vitro, and intraocular safety was assessed by ophthalmoscopy, electrophysiology, and histology after intravitreal injections, with resultant intravitreal concentrations of 0.2, 0.632, 1.12, and 2 mM. The treatment efficacy was evaluated by simultaneous intravitreal injection of HDP-P-GCV and herpes simplex virus type 1 (HSV-1) or by intravitreal injection of HDP-P-GCV at various times before HSV-1 intravitreal inoculation. Retinitis was scored with ophthalmoscopy and compared with controls. RESULTS: In vitro, the IC50 of HDP-P-GCV against HSV-1 and human cytomegalovirus (HCMV) infected cells was 0.02 and 0.6 microM, respectively. In rabbits in vivo, HDP-P-GCV dispersed evenly and maintained a good vitreous clarity at all doses except 2 mM final intravitreal concentration. Although cataracts were observed in some eyes at the higher doses, they were not observed in eyes with 0.2 mM final intravitreal concentration. No other indications of ocular toxicity were observed. Intravitreal injection of HDP-P-GCV with resultant 0.2 mM intravitreal concentration in the HSV-1 retinitis rabbit model demonstrated a complete protection of the retina with the simultaneous treatment strategy and a 4 (P = 0.03) to 6-(P = 0.058) week significant protection of retina with the pretreatment strategies when compared with ganciclovir or blank liposome controls. CONCLUSIONS: In the rabbit model of HSV-1 retinitis HDP-P-GCV acts as a long-lasting intravitreal injectable anti-CMV or anti-HSV compound. This self-assembling liposome system could be applicable for many compounds available for intraocular diseases.

Mitochondrial cardiolipin in diverse eukaryotes. Comparison of biosynthetic reactions and molecular acyl species.

Cardiolipin, a unique dimeric phospholipid of bacteria and mitochondria, can be synthesized by two alternative pathways discovered in rat and Escherichia coli, respectively. In mitochondrial preparations from fungi (Saccharomyces cerevisiae, Neurospora crassa), higher plants (Phaseolus aureus), molluscs (Mytilus edulis) and mammals (rat liver, bovine adrenal gland), cardiolipin was synthesized from CDP-diacylglycerol and phosphatidylglycerol, suggesting a common eukaryotic mechanism of cardiolipin formation which is in contrast to the prokaryotic biosynthesis from two molecules of phosphatidylglycerol. All mitochondrial cardiolipin synthases were inhibited by lysophosphatidylglycerol, were insensitive to N-ethylmaleimide and required divalent cations, although they had different cation specificities. The molecular species of cardiolipin from rat liver, bovine heart, S. cerevisiae and N. crassa were analysed by high-performance liquid chromatography of the derivative 1,3-bis[3'-sn-phosphatidyl]-2-benzoyl-sn-glycerol dimethyl ester. Cardiolipins from these organisms contained mainly monounsaturated or diunsaturated chains with 16 or 18 carbon atoms, resulting in a relatively homogeneous distribution of double bonds and carbon numbers among the four acyl positions. About half of the molecular species were symmetrical, i.e. they combined two identical diacylglycerol moieties. In N. crassa, the same species pattern was found at growth temperatures of 25 degrees C and 37 degrees C. Tentative molecular models were created for the most abundant molecular species and subjected to energy minimization. Geometric data, derived from these models, suggested similarities in the gross structure of the major cardiolipin species from different sources.