Addition of Lithiated 9-Deazapurine Derivatives to a Carbohydrate Cyclic Imine:  Convergent Synthesis of the Aza-<i>C</i>-nucleoside Immucillins

Evans, Gary B.; Furneaux, Richard H.; Hutchison, Tracy L.; Kezar, Hollis S.; Morris, Phillip; Schramm, and Vern L.; Tyler, Peter C.

doi:10.1021/jo0155613

Cited by 95 publications

(77 citation statements)

References 16 publications

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“…were overexpressed in Escherichia coli using the methods described (7)(8)(9). ImmH and MT-ImmH were synthesized as described (7,10,11 (12). S-Adenosylmethionine was converted to MTA by incubating S-adenosylmethionine at pH 3-5 and at 70°C for 3.5 h. All labeled compounds were purified by reverse phase-HPLC (RP-HPLC) with a 15-m, 7.8 ϫ 300-mm C 18 Deltapak column (Waters Associates), eluting isocratically with 50 mM ammonium acetate, pH 5.0, and 25% methanol at 1 ml/min.…”

Section: Methodsmentioning

confidence: 99%

“…In a second AMS experiment, synchronized P. falciparum cultures were incubated in triplicate for 12 h in hypoxanthine-free media and labeled with 100 fmol of the [8][9][10][11][12][13][14] C]purine for 12 h. Labeled cells were washed three times with cold PBS, and nucleic acids were precipitated and washed with 3% cold perchloric acid. Tributyrin was added as carbon carrier to decrease addition of background 14 C. In the first experiment, parasites were mostly trophozoites when label was added, whereas in the second experiment, parasites were late rings/early trophozoites.…”

Section: P Falciparum Culture and Immucillin Kill Curves-humanmentioning

confidence: 99%

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Targeting a Novel Plasmodium falciparum Purine Recycling Pathway with Specific Immucillins

Ting¹,

Shi

Lewandowicz

et al. 2005

Journal of Biological Chemistry

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111

182

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Plasmodium falciparum is unable to synthesize purine bases and relies upon purine salvage and purine recycling to meet its purine needs. We report that purines formed as products of polyamine synthesis are recycled in a novel pathway in which 5 -methylthioinosine is generated by adenosine deaminase. The action of P. falciparum purine nucleoside phosphorylase is a convergent step of purine salvage, converting both 5 -methylthioinosine and inosine to hypoxanthine. We used accelerator mass spectrometry to verify that 5 -methylthioinosine is an active nucleic acid precursor in P. falciparum. Prior studies have shown that inhibitors of purine salvage enzymes kill malaria, but potent malaria-specific inhibitors of these enzymes have not been described previously. 5 -Methylthio-immucillin-H, a transition state analogue inhibitor that is selective for malarial relative to human purine nucleoside phosphorylase, kills P. falciparum in culture. Immucillins are currently in clinical trials for other indications and may also have application as anti-malarials.

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Section: Methodsmentioning

confidence: 99%

Section: P Falciparum Culture and Immucillin Kill Curves-humanmentioning

confidence: 99%

Targeting a Novel Plasmodium falciparum Purine Recycling Pathway with Specific Immucillins

Ting¹,

Shi

Lewandowicz

et al. 2005

Journal of Biological Chemistry

Self Cite

111

182

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“…Accordingly, DADMe-Immucillin-H binds 2,400,000 times tighter than substrate according to the K m /K d ratio (21). Chemical synthesis of Immucillin-H requires the incorporation of four stereochemical centers and is inherently difficult (23,24). In contrast, DADMe-Immucillin-H has two stereocenters and can be made using the Mannich reaction, a three-way condensation of 9-deazahypoxanthine, 3-hydroxy-4-methoxy-pyrrolidine and formaldehyde under mild aqueous conditions (25).…”

Section: Human Purine Nucleoside Phosphorylasementioning

confidence: 99%

Enzymatic Transition State Theory and Transition State Analogue Design

Schramm

2007

Journal of Biological Chemistry

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128

210

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“…Levels of incorporation were measured using a 1450 Microbeta Liquid Scintillation and Luminescence Counter (PerkinElmer Life Science). MT-ImmH, ImmH, 2Ј-deoxy ImmH, and 2Ј-deoxy ImmG were synthesized as previously described (7,21,22). Culture media for studies with various immucillins contained no hypoxanthine supplement.…”

Section: Methodsmentioning

confidence: 99%

Plasmodium falciparum Purine Nucleoside Phosphorylase Is Critical for Viability of Malaria Parasites

Madrid¹,

Ting²,

Waller³

et al. 2008

Journal of Biological Chemistry

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Human malaria infections resulting from Plasmodium falciparum have become increasingly difficult to treat due to the emergence of drug-resistant parasites. The P. falciparum purine salvage enzyme purine nucleoside phosphorylase (PfPNP) is a potential drug target. Previous studies, in which PfPNP was targeted by transition state analogue inhibitors, found that those inhibiting human PNP and PfPNPs killed P. falciparum in vitro. However, many drugs have off-target interactions, and genetic evidence is required to demonstrate single target action for this class of potential drugs. We used targeted gene disruption in P. falciparum strain 3D7 to ablate PNP expression, yielding transgenic 3D7 parasites (⌬pfpnp). Lysates of the ⌬pfpnp parasites showed no PNP activity, but activity of another purine salvage enzyme, adenosine deaminase (PfADA), was normal. When compared with wild-type 3D7, the ⌬pfpnp parasites showed a greater requirement for exogenous purines and a severe growth defect at physiological concentrations of hypoxanthine. Drug assays using immucillins, specific transition state inhibitors of PNP, were performed on wild-type and ⌬pfpnp parasites. The ⌬pfpnp parasites were more sensitive to PNP inhibitors that bound hPNP tighter and less sensitive to MT-ImmH, an inhibitor with 100-fold preference for PfPNP over hPNP. The results demonstrate the importance of purine salvage in P. falciparum and validate PfPNP as the target of immucillins.Each year, Plasmodium species infect 300 to 500 million people and cause nearly two million deaths, mostly in children under the age of five in sub-Saharan Africa (1). Most deaths are due to infection with Plasmodium falciparum. Only AIDS and tuberculosis are more lethal human infectious diseases. No vaccine is available, despite intensive international efforts. At present, control of malaria is dependent on prevention with bed nets, insecticides or chemoprophylaxis, and chemotherapeutic treatment of clinical cases. However, chemotherapy for malaria has been complicated by recent increases in mortality and morbidity due to the emergence of drug-resistant strains (2).Because parasitic protozoa are unable to synthesize purines de novo, purine salvage has been proposed as a potential target for chemotherapy of protozoan parasite infections, including those caused by Plasmodium spp. Malaria parasites are obligate intracellular parasites that carry out their asexual cycle in erythrocytes. Unlike most mammalian cells, erythrocytes have no biochemical machinery for de novo purine synthesis, but act as a rich source of purine salvage enzymes, particularly purine nucleoside phosphorylase (PNP) 4 and adenosine deaminase (ADA). The purine salvage pathway of Plasmodium begins with the deamination of adenosine to inosine by ADA, followed by conversion of inosine to hypoxanthine by PNP. The final enzyme in the pathway is hypoxanthine-guanine-xanthine phosphoribosyltransferase (HGXPRT). Hypoxanthine is a precursor for all purines and is a central metabolite for nucleic acid synthesis in P. ...

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Addition of Lithiated 9-Deazapurine Derivatives to a Carbohydrate Cyclic Imine: Convergent Synthesis of the Aza-C-nucleoside Immucillins

Abstract: Means have been developed for the synthesis and addition of 9-deaza-9-lithiopurine derivatives to the carbohydrate-derived cyclic imine 6 in facile convergent syntheses of biologically active aza-C-nucleosides.

Cited by 95 publications

References 16 publications

Targeting a Novel Plasmodium falciparum Purine Recycling Pathway with Specific Immucillins

Targeting a Novel Plasmodium falciparum Purine Recycling Pathway with Specific Immucillins

Enzymatic Transition State Theory and Transition State Analogue Design

Plasmodium falciparum Purine Nucleoside Phosphorylase Is Critical for Viability of Malaria Parasites

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