Dipeptidyl-peptidase IV (DPP IV/CD26)-activated prodrugs: a successful strategy for improving water solubility and oral bioavailability.

In the search of novel enzyme-based prodrug approaches to improve pharmacological properties of therapeutic drugs such as solubility and bioavailability, dipeptidyl-peptidase IV (DPP IV, also termed as CD26) enzyme activity provides a previously unexplored successful prodrug strategy. This review covers key aspects of the enzyme useful for the design of CD26-directed prodrugs. The proof-of-concept of this prodrug technology is provided for amine-containing agents by directly linking appropriate di- (or oligo)peptide moieties to a free amino group of a non-peptidic drug through an amide bond which is specifically hydrolized by DPP IV/CD26. Special emphasis is also made in discussing the design and synthesis of more elaborated tripartite prodrug systems, for further extension of the strategy to hydroxy-containing drugs. The application of this technology to improve water solubility and oral bioavailability of prominent examples of antiviral nucleosides is highlighted.

Novel water-soluble prodrugs of acyclovir cleavable by the dipeptidyl-peptidase IV (DPP IV/CD26) enzyme.

We herein report for the first time the successful use of the dipeptidyl peptidase IV (DPPIV/CD26) prodrug approach to guanine derivatives such as the antiviral acyclovir (ACV). The solution- and solid-phase synthesis of the tetrapeptide amide prodrug 3 and the tripeptide ester conjugate 4 of acyclovir are reported. The synthesis of the demanding tetrapeptide amide prodrug of ACV 3 was first established in solution and successfully transferred onto solid support by using Ellman's dihydropyran (DHP) resin. In contrast with the valyl ester prodrug (valacyclovir, VACV), the tetrapeptide amide prodrug 3 and the tripeptide ester conjugate 4 of ACV proved fully stable in PBS. Both prodrugs converted to VACV (for 4) or ACV (for 3) upon exposure to purified DPPIV/CD26 or human or bovine serum. Vildagliptin, a potent inhibitor of DPPIV/CD26 efficiently inhibited the DPPIV/CD26-catalysed hydrolysis reaction. Both amide and ester prodrugs of ACV showed pronounced anti-herpetic activity in cell culture and significantly improved the water solubility in comparison with the parent drug.

Dipeptidyl peptidase IV-activated prodrugs of anti-varicella zoster virus bicyclic nucleoside analogues containing different self-cleavage spacer systems.

A new type of double prodrug of the antiviral family of bicyclic nucleoside analogues (BCNA) bearing cyclization self-cleavage spacers between the Val-Pro dipeptide sequence as well as the parent compound were synthesized and evaluated with regard to activation by the DPPIV/CD26 enzyme and for their stability in human and bovine serum. In buffer solution, carbamate and ester prodrugs were found to be chemically stable. Most prodrugs containing a dipeptidyl linker efficiently converted into the BCNA parent drug. In contrast, the Val-Pro alkyldiamino prodrugs converted predominantly into their alkyldiamino prodrug intermediates in the presence of CD26 and human serum. A marked increase in water solubility was observed for all prodrugs. In contrast to the parent compound, a tetrapeptide prodrug containing the Val-Val dipeptide as a self-cleavage spacer released substantial amounts of the BCNA parent drug at the basolateral side of Caco-2 cell cultures and exhibited 15- to 20-fold increased bioavailability in mice relative to the poorly bioavailable parent compound.

Dipeptidyl peptidase IV (DPPIV/CD26)-based prodrugs of hydroxy-containing drugs.

We previously described a novel prodrug approach in which a di- or tetrapeptide moiety is linked to a wide variety of amine-containing drugs through an amide bond, which is specifically cleaved by dipeptidyl peptidase IV (DPPIV/CD26) activity. Herein we report the application of this prodrug approach to a variety of hydroxy-containing drugs (primary, secondary, tertiary, or aromatic hydroxy groups). We designed and studied tripartite prodrugs containing a dipeptide moiety (cleavable by DPPIV/CD26) and a valine as a hetero-bifunctional connector to link the dipeptide to the hydroxy group of the drug through a metabolically labile ester bond. The hydroxy-containing prodrugs showed various susceptibilities to hydrolysis by DPPIV/CD26 and serum, depending on the nature of the compound. Prodrugs of compounds containing a primary hydroxy group (as in didanosine) or a hydroxy moiety on an aromatic entity (as in acetaminophen) were most efficiently converted. In contrast, a tertiary hydroxy group was much less susceptible to conversion into its parent drug by DPPIV/CD26 or serum. A number of the prodrugs showed remarkable increases in water solubility relative to their parent drugs.

Dipeptidyl peptidase IV dependent water-soluble prodrugs of highly lipophilic bicyclic nucleoside analogues.

We present the first report of the application of the dipeptidyl peptidase IV (DPPIV/CD26) based prodrug approach to hydroxy-containing drug derivatives. In particular, we applied this strategy to the highly lipophilic antiviral drug family of bicyclic furanopyrimidine nucleoside analogues (BCNA) in order to improve their physicochemical and pharmacokinetic properties. Our stability data demonstrated that the prodrugs efficiently release the parent BCNA drug upon selective conversion by purified DPPIV/CD26 and by soluble DPPIV/CD26 present in bovine, murine, and human serum. Vildagliptin, a specific inhibitor of DPPIV/CD26, was able to completely block the hydrolysis of the prodrugs in the presence of purified DPPIV/CD26 human, murine, and bovine serum. Several novel prodrugs showed remarkable increases in water solubility (up to more than 3 orders of magnitude) compared to the poorly soluble parent drug. We also demonstrated a markedly enhanced oral bioavailability of the prodrugs versus the parent drug in mice.

Application of the dipeptidyl peptidase IV (DPPIV/CD26) based prodrug approach to different amine-containing drugs.

Here we explore the applicability of the dipeptidyl peptidase IV (DPPIV/CD26) based prodrug approach to a variety of amine-containing drugs. Efficient procedures have been developed for the synthesis of dipeptide and tetrapeptide amide prodrugs including N-acylation protocols of the exocyclic amino function of cytidine and adenosine nucleosides. Our studies demonstrated that XaaPro dipeptides linked to a free amino group present on an aromatic ring or on a sugar entity are prodrugs that efficiently release the parent drug upon conversion by purified DPPIV/CD26 as well as soluble DPPIV/CD26 in bovine and human serum. Vildagliptin, a specific inhibitor of DPPIV/CD26, was able to completely block the hydrolysis of the prodrugs in the presence of purified CD26 but also in human and bovine serum. When the amino group is present on a pyrimidine or purine ring, the dipeptide derivatives are chemically unstable, whereas the tetrapeptide derivatives (i.e., ValProValPro or ValAlaValPro) were much more stable in solution and efficiently converted to the parent drug by the action of DPPIV/CD26. This DPPIV/CD26-directed prodrug technology can be useful to increase solubility of the parent drug molecules and/or to allow better formulation properties.

Preclinical development of bicyclic nucleoside analogues as potent and selective inhibitors of varicella zoster virus.

OBJECTIVES: To progress the anti-varicella-zoster-virus (VZV) aryl bicyclic nucleoside analogues (BCNAs) to the point of Phase 1 clinical trial for herpes zoster. METHODS: A new chromatography-free synthetic access to the lead anti-VZV aryl BCNAs is reported. The anti-VZV activity of lead Cf1743 was evaluated in monolayer cell cultures and organotypic epithelial raft cultures of primary human keratinocytes. Oral dosing in rodents and preliminary pharmacokinetics assessment was made, followed by an exploration of alternative formulations and the preparation of pro-drugs. We also studied uptake into cells of both parent drug and pro-drug using fluorescent microscopy and biological assays. RESULTS: Cf1743 proved to be significantly more potent than all reference anti-VZV compounds as measured either by inhibition of infectious virus particles and/or by viral DNA load. However, the very low water solubility of this compound gave poor oral bioavailability (approximately 14%). A Captisol admixture and the 5'-monophosphate pro-drug of Cf1743 greatly boosted water solubility but did not significantly improve oral bioavailability. The most promising pro-drug to emerge was the HCl salt of the 5'-valyl ester, designated as FV-100. Its uptake into cells studied using fluorescent microscopy and biological assays indicated that the compound is taken up by the cells after a short period of incubation and limited exposure to drug in vivo may have beneficial effects. CONCLUSIONS: On the basis of its favourable antiviral and pharmacokinetic properties, FV-100 is now being pursued as the clinical BCNA candidate for the treatment of VZV shingles.

Efficient conversion of tetrapeptide-based TSAO prodrugs to the parent drug by dipeptidyl-peptidase IV (DPPIV/CD26).

A novel prodrug approach has been evaluated using the anti-HIV-active TSAO molecule as the prototype drug to prove the kinetics with purified enzyme and the principles of conversion to the parent compound in sera and cell culture. When a variety of tetrapeptidyl amide prodrugs of NAP-TSAO were synthesized and exposed to purified dipeptidyl-peptidase IV (DPPIV/CD26) as well as human and bovine sera, they are converted to the parent NAP-TSAO drug in two successive steps by both purified CD26 and human and bovine serum. The efficiency of conversion strongly depends on the nature of the amino acid that has to be cleaved-off from the prodrug molecule. The tetrapeptidyl prodrug 20 showed a more than 10-fold improved water-solubility in comparison to that of the parent compound NAP-TSAO. The antiviral activity of the prototype NAP-TSAO could also be modulated by introducing different tetrapeptide moieties on the molecule resulting, in some cases, in a superior antiviral potential in cell culture than the parent drug.