Christopher K. H. Tseng

Identification of active antiviral compounds against a New York isolate of West Nile virus

The recent West Nile virus (WNV) outbreak in the United States has increased the need to identify effective therapies for this disease. A chemotherapeutic approach may be a reasonable strategy because the virus infection is typically not chronic and antiviral drugs have been identified to be effective in vitro against other flaviviruses. A panel of 34 substances was tested against infection of a recent New York isolate of WNV in Vero cells and active compounds were also evaluated in MA-104 cells. Some of these compounds were also evaluated in Vero cells against the 1937 Uganda isolate of the WNV. Six compounds were identified to be effective against virus-induced CPE with 50% effective concentrations (EC50) less than 10 microg/ml and with a selectivity index (SI) of greater than 10. Known inhibitors of orotidine monophosphate decarboxylase and inosine monophosphate dehydrogenase involved in the synthesis of GTP, UTP, and TTP were most effective. The compounds 6-azauridine, 6-azauridine triacetate, cyclopententylcytosine (CPE-C), mycophenolic acid and pyrazofurin appeared to have the greatest activities against the New York isolate, followed by 2-thio-6-azauridine. Anti-WNV activity of 6-azauridine was confirmed by virus yield reduction assay when the assay was performed 2 days after initial infection in Vero cells. The neutral red assay mean EC50 of ribavirin was only 106 microg/ml with a mean SI of 9.4 against the New York isolate and only slightly more effective against the Uganda isolate. There were some differences in the drug sensitivities of the New York and Uganda isolates, but when comparisons were made by categorizing drugs according to their modes of action, similarities of activities between the two isolates were identified.

3-Deazaneplanocin: A new and potent inhibitor of S-adenosylhomocysteine hydrolase and its effects on human promyelocytic leukemia cell line HL-60

3-Deazaneplanocin, a new carbocyclic analog of adenosine, was synthesized as an inhibitor of S-adenosylhomocysteine hydrolase. The Ki of 3-deazaneplanocin for a purified hamster liver preparation of S-adenosylhomocysteine hydrolase was 5 X 10(-11) M, making this inhibitor 250-fold more potent than the previously known most potent inhibitor of this enzyme, 3-deazaaristeromycin. Inhibition was competitive with the substrate adenosine. Human promyelocytic leukemia (HL-60) cells treated with 10(-5) M 3-deazaneplanocin showed a pronounced elevation in S-adenosylhomocysteine which was 4-fold greater than that produced by an equimolar concentration of 3-deazaaristeromycim. This effect preceded a moderate reduction in cell growth and viability following continuous exposure for 6 days. Cellular differentiation as monitored by the reduction of nitroblue tetrazolium was not markedly affected except after 4 days exposure to 10(-5) M 3-deazaneplanocin where 60% of the viable cells were positive. These results indicate that 3-deazaneplanocin may have therapeutic potential as an anticancer or antiviral drug.

3-deazaneplanocin A: A new inhibitor of S-adenosylhomocysteine synthesis and its effects in human colon carcinoma cells

The mechanism of action of the cyclopentenyl analogue of 3-deazaadenosine (3-deazaneplanocin A or c3Nep) was investigated in the human colon carcinoma cell line HT-29. Upon exposure of cells for 24 hr to 3-deazaneplanocin A (c3Nep), neplanocin A (Nep) or 3-deazaaristeromycin (c3Ari), significant toxicity was noted only for Nep, wherein an 87% reduction in viability was produced at a 100 microM concentration. c3Nep and c3Ari at 100 microM reduced viability by 34 and 21%, respectively. Intracellular levels of S-adenosylhomocysteine (AdoHcy) were elevated by a 24-hr exposure to 100 microM c3Nep, Nep and c3Ari and were 120, 75 and 25 pmoles/10(6) cells respectively. Only Nep was metabolized to an S-adenosylmethionine-like metabolite, and its formation was dose-related to its cytotoxicity. The t1/2 for the disappearance of elevated levels of AdoHcy following drug removal was 1.6 to 2.5 hr for all drugs. rRNA and tRNA methylation was inhibited significantly by Nep, but c3Nep and c3Ari inhibited tRNA methylation but not rRNA methylation to a lesser degree. These results demonstrate that c3Nep is a potent inhibitor of AdoHcy synthesis with a low degree of cytotoxicity.

An improved method of synthesis of neplanocin and related cyclopentenyl-containing nucleosides

Abstract A facile one-step condensation of the cyclopentenyl α,-tosylate 2c with the alkali metal salts of 6-chloropurine and uracil, afforded the protected carbocyclic nucleosides 5a and 8 , respectively, which were converted to neplanocin a ( 6 ) and cyclopentenyl uracil ( 9 ) by established methods.

Synthesis of 2′,3′-Dideoxycyclopentenyl Carbocyclic Nucleosides as Potential Drugs for the Treatment of AIDS

Abstract The recently discovered antiviral activity of 2′,3′-dideoxy-nucleosides against the AIDS virus in vitro prompted the synthesis of 2′,3′-dideoxycyclopentenyl adenine and cytosine. A sequence of two consecutive reductive deoxygenations using 1,1′-thiocarbonyldiimidazole followed by reduction with tributyl tin hydride afforded the desired target compounds. Neither compound showed activity in the in vitro assay against the HTLV-III/LAV virus.

Plant-derived and semi-synthetic calanolide compounds with in vitro activity against both human immunodeficiency virus type 1 and human cytomegalovirus.

Plant-derived and semi-synthetic calanolide compounds with anti-human immunodeficiency virus type 1 (HIV-1) activity were tested for anti-human cytomegalovirus (HCMV) activity in both cytopathic effect inhibition and plaque reduction assays. The results indicated that the anti-HCMV activity of calanolide compounds does not correlate with their activity against HIV-1. The semi-synthetic 12-keto derivatives tended to be more active against HCMV than the corresponding 12-OH congeners, which were more active against HIV-1. It appeared that the 7,8-unsaturated double bond in the chromene ring played a certain role in maintaining activities against both HCMV and HIV-1. Saturation of the double bond increased the EC50 values against both viruses, with concomitant increase in toxicity. The calanolide compounds reported here are the first non-nucleoside analogues capable of inhibiting both HIV-1 and HCMV and, therefore, may be useful chemoprophylactic agents for HCMV in HIV-infected people or vice versa.

NIAID resources for developing new therapies for severe viral infections

Abstract Severe viral infections, including hemorrhagic fever and encephalitis, occur throughout the world, but are most prevalent in developing areas that are most vulnerable to infectious diseases. Some of these can also infect related species as illustrated by the threatened extinction of gorillas by Ebola infection in west and central Africa. There are no safe and effective treatments available for these serious infections. In addition to the logistical difficulties inherent in developing a drug for infections that are sporadic and occur mainly in the third world, there is the overwhelming barrier of no hope for return on investment to encourage the pharmaceutical industry to address these unmet medical needs. Therefore, the National Institute of Allergy and infectious Disease (NIAID) has developed and supported a variety of programs and resources to provide assistance and lower the barrier for those who undertake these difficult challenges. The primary programs relevant to the development of therapies for severe viral infections are described and three case studies illustrate how they have been used. In addition, contact information for accessing these resources is supplied.