Novel nucleotide analogues bearing (1H-1,2,3-triazol-4-yl)phosphonic acid moiety as inhibitors of Plasmodium and human 6-oxopurine phosphoribosyltransferases

A novel family of acyclic nucleoside phosphonates (ANPs) bearing a (1H-1,2,3-triazol-4-yl)phosphonic acid group in the acyclic side chain have been prepared in order to study the influence of the hetaryl rigidizing element on the biological properties of such compounds. The key synthetic step consisted of a copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) between diethyl ethynylphosphonate and the corresponding azidoalkyl precursor. Two ANPs in this family, bearing a guanine base, exhibited the highest potency for the human 6-oxopurine phosphoribosyltransferase irrespective of the stereochemistry on the C-2′ atom. Four compounds inhibited Plasmodium falciparum 6-oxopurine phosphoribosyltransferase with little differences in their Kvalues irrespective of whether the base was guanine, hypoxanthine or xanthine but only two, with guanine as base, inhibited PvHGPRT

Acyclic Nucleoside Phosphonates as Inhibitors of Hypoxanthine-Guanine-Xanthine Phosphoribosyltransferase: New Anti-Malarial Chemotherapy Leads

Hypoxanthine-guanine-xanthine phosphoribosyltransferase (HGXPRTase) is a widely recognized target for the discovery of new anti-malarial drugs. Specific acyclic nucleoside phosphonates (ANPs) inhibit HGXPRTase and possess anti-plasmodial activity. Within the framework of a SAR-study, the classical ANPs (e.g. PME-, PMP- and HPMP-derivatives) as well as novel series of compounds were tested to investigate their efficiency and selectivity on the inhibition of P. falciparum, P. vivax and human enzyme

Acyclic nucleoside bisphosphonates as inhibitors of 6-oxopurine phosphoribosyltransferases: Potential antimalarial and antibacterial agents

Acyclic nucleoside phosphonates (ANPs) that contain a 6-oxopurine base are good inhibitors of the human, Plasmodium falciparum, P. vivax, Escherichia coli and Mycobacterium tuberculosis 6-oxopurine phosphoribosyltransferases (PRTases), key enzymes of the purine salvage pathway. Chemical modifications based on the crystal structures of several inhibitors in complex with human HGPRTase have led to the design of new ANPs. These novel compounds contain a second phosphonate group attached to the ANP scaff old. The crystal structures of these inhibitors in complex with human HGPRTase show that they can fill three critical locations in the active site: the binding sites of the purine base, the 5’-phosphate group and pyrophosphate. Prodrugs have been synthesized and have been shown to arrest the growth of P. falciparum in erythrocyte culture. Prodrugs of selected ANPs also inhibit the growth of Mycobacterium tuberculosis in cell-based assays

Structure-based development of Plasmodium hypoxanthine-guanine phosphoribosyltransferase inhibitors: a proof of concept study

Structure-based development of Plasmodium hypoxanthine-guanine phosphoribosyltransferase inhibitors: a proof of concept study Manon Laportea, Dana Hockováb, Dianne T. Keoughc, Luke W. Guddatc, Lieve Naesensa aRega Institute, KU Leuven, Belgium; bInstitute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic, cSchool of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia The malarial parasites Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) are purine auxotrophs that rely on the salvage of host purines for their survival and growth. A critical enzyme in this salvage pathway, hypoxanthine guanine phosphoribosyltransferase (HGPRT), is considered a prime target for antimalarial therapy. Our collaboration between the University of Queensland, the Institute of Organic Chemistry and Biochemistry (Prague) and the Rega institute (KU Leuven), is focussed on rational development of novel acyclic nucleoside phosphonates (ANPs) that are structural analogues of the HGPRT nucleotide reaction products IMP and GMP. Based on available HGPRT crystal structures, ANP derivatives with a second phosphonate group attached (bisANPs) were designed. These compounds proved to be particularly strong HGPRT inhibitors with Ki values as low as 30 nM, and displayed antimalarial activity in Pf-infected erythrocytes with IC50 values as low as 3.8 µM (Keough et al. 2013). However, since the Plasmodium parasite possesses several enzymes that could possibly serve as the target for inhibition by the (bis)ANPs, either directly or after metabolic conversion, it remained to be established whether the observed antiparasitic effect indeed results from HGPRT inhibition. We here present a target validation assay to assess, in a cellular environment, the inhibitory effect of (bis)ANPs towards HGPRT. This method complements the enzymatic assays (in which purified HGPRT enzymes are studied in a cell-free environment), and the Plasmodium cell culture assay (that involves replication of the whole Pf parasite). First, we created adenoviral (Ad) vectors containing the cDNA sequences encoding human, Pf or PvHGPRT. Transduction of these Ad vectors into HGPRT-deficient human 1306 cells generated high HGPRT expression levels, as estimated by a tritium release assay with [2,8-3H]hypoxantine (Balzarini and De Clercq, 1992). Several (bis)ANPs were shown to inhibit the HGPRT reaction by the human or Pv enzyme. Our novel assay allows to validate Plasmodium HGPRT inhibitors in cell culture and will be instrumental to guide further development of this new class of antimalarial drugs.status: submitte

Acyclic nucleoside phosphonates with a branched 2-(2-phosphonoethoxy)ethyl chain: Efficient synthesis and antiviral activity

Series of novel acyclic nucleoside phosphonates (ANPs) with various nucleobases and 2-(2-phosphonoethoxy)ethyl (PEE) chain bearing various substituents in β-position to the phosphonate moiety were prepared. The influence of structural alternations on antiviral activity was studied. Several derivatives exhibit antiviral activity against HIV and vaccinia virus (middle micromolar range), HSV-1 and HSV-2 (lower micromolar range) and VZV and CMV (nanomolar range), although the parent unbranched PEE-ANPs are inactive. Adenine as a nucleobase and the methyl group attached to the PEE chain proved to be a prerequisite to afford pronounced antiviral activity.status: publishe

Crystal structures of Trypanosoma brucei hypoxanthine – guanine – xanthine phosphoribosyltransferase in complex with IMP , GMP and XMP

The 6-oxopurine phosphoribosyltransferases (PRTs) are drug targets for the treatment of parasitic diseases. This is due to the fact that parasites are auxotrophic for the 6-oxopurine bases relying on salvage enzymes for the synthesis of their 6-oxopurine nucleoside monophosphates. In Trypanosoma brucei, the parasite that is the aetiological agent for sleeping sickness, there are three 6-oxopurine PRT isoforms. Two are specific for hypoxanthine and guanine, whilst the third, characterized here, uses all three naturally occurring bases with similar efficiency. Here, we have determined crystal structures for TbrHGXPRT in complex with GMP, XMP and IMP to investigate the structural basis for substrate specificity. The results show that Y201 and E208, not commonly observed within the purine binding pocket of 6-oxopurine PRTs, contribute to the versatility of this enzyme. The structures further show that a nearby water can act as an adaptor to facilitate the binding of XMP and GMP. When GMP binds, a water can accept a proton from the 2-amino group but when XMP binds, the equivalent water can donate its proton to the 2-oxo group. However, when IMP is bound, no water molecule is observed at that location. DATABASE: Coordinates and structure factors were submitted to the Protein Data Bank and have accession codes of 6MXB, 6MXC, 6MXD and 6MXG for the TbrHGXPRT.XMP complex, TbrHGXPRT.GMP complex, TbrHGXPRT.IMP complex, and TbrHGPRT.XMP complex, respectively

6-Oxopurine phosphoribosyltransferase: A target for the development of antimalarial drugs

Malaria remains the most serious parasitic diseases affecting humans in the world today, resulting in 1-2 million fatalities each year. Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) are the predominant causative agents. Both are responsible for widespread mortality and morbidity and are a serious socio-economic burden, especially for countries in the developing world. One of the most important defences against malaria has been the use of chemotherapeutic drugs (e.g. chloroquine, artemisinins, pyrimethamine) but these have mainly been found by serendipity. Their mechanisms was not understood at the time of their discovery and, even today, are still not unequivocal. For many of these compounds, the parasite is now resistant and, hence, there is an urgent need to develop new therapeutic drugs directed to validated targets. One metabolic pathway crucial for the survival and replication and survival of the parasite is the synthesis of the purine nucleoside monophosphates essential for the production of DNA/RNA molecules. A key enzyme in this pathway is the 6-oxopurine phosphoribosyltransferase (PRTase). The focus of this review is on the identification and characterization of inhibitors of the enzymes from both Pf and Pv as antimalarial drug leads. The acyclic nucleoside phosphonates (ANPs) appear to be excellent candidates because they are good inhibitors of the two Plasmodium enzymes, can be selective compared to the human enzyme, can arrest parasitemia in cell based assays, have low cytotoxicity to the human host cell and, because of their stable carbon-phosphorous bond, are stable within the cell