Chemical and enzymatic stability of amino acid derived phosphoramidates of antiviral nucleoside 5′-monophosphates bearing a biodegradable protecting group

Abstract

Ribavirin and 2'-O-methylcytidine 5'-phosphoramidates derived from L-alanine methyl ester bearing either an O-phenyl or a biodegradable O-[3-(acetyloxy)-2,2-bis(ethoxycarbonyl)propyl] or O-[3-(acetyloxymethoxy)-2,2-bis(ethoxycarbonyl)propyl] protecting group were prepared. The kinetics of the deprotection of these pro-drugs by porcine liver esterase and by a whole cell extract of human prostate carcinoma was studied by HPLC-ESI-MS/MS. The 3-(acetyloxymethoxy)-2,2-bis(ethoxycarbonyl)propyl and 3-(acetyloxy)-2,2-bis(ethoxycarbonyl)propyl groups were readily removed releasing the l-alanine methyl ester phosphoramidate nucleotide, the deprotection of the 3-(acetyloxymethoxy) derivative being approximately 20 times faster. The chemical stability of the 2'-O-methylcytidine pro-drugs was additionally determined over a pH range from 7.5 to 10.

Prodrugs of Phosphonates and Phosphates: Crossing the Membrane Barrier

“…For example, in place of an aryl ester, a 3-acyloxymethoxypropyl group has been studied (e.g. 65 ), and hydrolysis of the ester was found to be ~20-fold faster than the corresponding phenyl phosphoramidate [131]. In this work, small differences in the reactivity of the R P and S P isomers were noted, but it was not possible to identify which one was the more reactive.…”

“…Simple phenyl esters may well be the most extensively employed substructure [130,131], especially in nucleotide analogues such of the general structure 62 [116,127,132–136]. However in cases such as the hydroxamic acid 63, an inhibitor of 6-phosphogluconate dehydrogenase for use in trypanosomiasis, addition of simple substituents such as methyl groups to the phenyl ring has resulted in more than a 10-fold increase in potency [137].…”

Synthesis of Nucleoside Phosphate and Phosphonate Prodrugs

“…For example, Leisvuori et al 215 prepared 2′-OMe cytidine aryloxyphosphoramidate prodrug 422 by first 5′-hydroxyl silylation of 418 , tritylation of the N 4 -position, and protection of the 3′-hydroxyl with levulinic acid and DCC in dioxane. Selective 5′-desilylation carried out with TBAF in a mixture of THF and acetic acid afforded the appropriately protected nucleoside 420 .…”

“…In the same vein, Leisvuori and co-workers 215 used levulinate groups to prepare ribavirin phosphoramidate prodrug 426 . 2′,3′-Bis-levulinoylated ribavirin 423 was reacted with 1.5 equiv of diphenyl phosphite in pyridine to allow formation of the nucleoside phosphite phenyl ester and subsequently to alanine methyl ester in the presence of carbon tetrachloride and triethylamine to form the protected nucleoside prodrug 425 in 67% yield.…”

Influence of the Nucleobase and Anchimeric Assistance of the Carboxyl Acid Groups in the Hydrolysis of Amino Acid Nucleoside Phosphoramidates

“…Recently, the kinetics of the hydrolysis of l-alanine methyl esters of NP prodrugs was reported and a mechanism for the hydrolysis reaction in both acidic and basic solutions was proposed. [13][14][15] However, the physicochemical properties and structural determinants that influence the chemical stability and hydrolysis mechanism of this class of compounds are not known in detail. We have synthesized a series of aaNPs, containing either natural or modified nucleobases (Figure 1).…”