Inna L. Karpenko

Synthesis and antiherpetic activity of acyclovir phosphonates.

Abstract Phosphonate derivatives of acyclovir containing phosphorous acid and ethoxycarbonylphosphonic acid residues as well as their isopropyl esters were prepared. They selectively inhibited the herpes simplex virus 1 reproduction in Vero cell culture, the efficacy of esters being 3–4 times higher than that of ACV. The hydrolysis of the synthesized compounds was studied in the PBS buffer and human blood serum.

Antiviral Properties, Metabolism, and Pharmacokinetics of a Novel Azolo-1,2,4-Triazine-Derived Inhibitor of Influenza A and B Virus Replication

ABSTRACT Influenza viruses of types A and B cause periodic pandemics in the human population. The antiviral drugs approved to combat influenza virus infections are currently limited. We have investigated an effective novel inhibitor of human influenza A and B viruses, triazavirine {2-methylthio-6-nitro-1,2,4-triazolo[5,1-c]-1,2,4-triazine-7(4Í)-one} (TZV). TZV suppressed the replication of influenza virus in cell culture and in chicken chorioallantoic membranes, and it protected mice from death caused by type A and B influenza viruses. TZV was also effective against a rimantadine-resistant influenza virus strain and against avian influenza A virus H5N1 strains. The pharmacokinetic parameters and bioavailability of TZV were calculated after the administration of TZV to rabbits. The TZV metabolite AMTZV {2-methylthio-6-amino-1,2,4-triazolo[5,1-s]-1,2,4-triazin(e)-7(4Í)-one} was discovered in ÍÅK 293T and Huh7 cell cultures, a liver homogenate, and rabbit blood after intragastric administration of TZV. AMTZV was nontoxic and inactive as an inhibitor of influenza virus in cell culture. Most likely, this metabolite is a product of TZV elimination.

HCV Core Protein Uses Multiple Mechanisms to Induce Oxidative Stress in Human Hepatoma Huh7 Cells

Hepatitis C virus (HCV) infection is accompanied by the induction of oxidative stress, mediated by several virus proteins, the most prominent being the nucleocapsid protein (HCV core). Here, using the truncated forms of HCV core, we have delineated several mechanisms by which it induces the oxidative stress. The N-terminal 36 amino acids of HCV core induced TGFβ1-dependent expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases 1 and 4, both of which independently contributed to the production of reactive oxygen species (ROS). The same fragment also induced the expression of cyclo-oxygenase 2, which, however, made no input into ROS production. Amino acids 37–191 of HCV core up-regulated the transcription of a ROS generating enzyme cytochrome P450 2E1. Furthermore, the same fragment induced the expression of endoplasmic reticulum oxidoreductin 1α. The latter triggered efflux of Ca2+ from ER to mitochondria via mitochondrial Ca2+ uniporter, leading to generation of superoxide anions, and possibly also H2O2. Suppression of any of these pathways in cells expressing the full-length core protein led to a partial inhibition of ROS production. Thus, HCV core causes oxidative stress via several independent pathways, each mediated by a distinct region of the protein.

ANTI-HIV ACTIVITY OF NOVEL PHOSPHONATE DERIVATIVES OF AZT, d4T, AND ddA

Anti-HIV activity and cytotoxicity were tested for novel phosphonate derivatives of AZT, d4T and ddA. For d4T phosphonate derivatives the most active was 2′,3′-Dideoxy-2′,3′-didehydrothymidine 5′-isopropylphosphite and among the AZT phosphonate derivatives highest activity was shown by 2′,3′-Dideoxy-3′-azidothymidine 5′-cyclohexylphosphite.

1,2,4-Triazoloazine derivatives as a new type of herpes simplex virus inhibitors

A new class of inhibitors of herpes simplex virus replication was found. The compounds under study are derived from condensed 1,2,4-triazolo[5,1-c][1,2,4]triazines and 1,2,4-triazolo[1,5-a]pyrimidines, structural analogues of natural nucleic bases. Antiherpetic activity and cytotoxicity of the compounds were studied. The corresponding triphosphates of several active compounds were prepared and tested as inhibitors of DNA synthesis catalyzed by herpes simplex virus polymerase. The potential mechanism of their action is blocking of DNA dependent DNA polymerase, a key enzyme of viral replication.

Antiherpetic Properties of Acyclovir 5′-Hydrogenphosphonate and the Mutation Analysis of Herpes Virus Resistant Strains

In this study, we continued to study antiherpetic properties of acyclovir 5′‐hydrogenphosphonate (Hp‐ACV) in cell cultures and animal models. Hp‐ACV was shown to inhibit the development of herpetic infection in mice induced by the HSV‐1/L2 strain. The compound suppressed replication of both ACV‐sensitive HSV‐1/L2 and ACV‐resistant HSV‐1/L2/R strains in Vero cell culture. Viral population resistant to Hp‐ACV (HSV‐1/L2/RHp‐ACV) was developed much slower than ACV‐resistant population. The analysis of Hp‐ACV‐resistant clones isolated from the HSV‐1/L2/RHp‐ACV population demonstrated their partial cross‐resistance to ACV. The mutations determining the resistance of HSV‐1 clones to Hp‐ACV were partly overlapped with mutations defining ACV resistance but did not always coincide. HSV‐1/L2/RHp‐ACV herpes virus thymidine kinase is shortened from the C‐terminus by 100 amino acid residues in length.

Cell Metabolism of Acyclovir Phosphonate Derivatives and Antiherpesvirus Activity of their Combinations with α2-Interferon

The combinational use of acyclovir (ACV) phosphonate esters and α2‐interferon was shown to produce a synergistic effect on inhibition of HSV‐1 replication in Vero cell cultures. Unlike other acyclovir phosphonate derivatives studied earlier, ACV H‐phosphonate is not an ACV prodrug. On penetrating into the cells, it may be directly converted into ACV monophosphate escaping dephosphonylation–phosphorylation steps.

Synthesis and Anti‐HIV Properties of New Carbamate Prodrugs of AZT

A series of new 5′‐O‐carbamate prodrugs of AZT have been prepared. The stability in biological media, anti‐HIV properties and pharmacokinetic parameters in dogs were evaluated. The compounds display moderate anti‐HIV activity in cell culture. After oral administration of carbamate IV in dogs, both intact prodrug IV and released AZT were discovered in dog blood. Pharmacokinetic parameters of the compound IV were estimated. Half‐life (T1/2) of AZT released after oral administration of IV in dogs was close to that after administration of AZT itself, and time to the maximum concentration (Tmax) of AZT released from IV was two and three times longer compared with that of AZT and H‐phosphonate AZT, respectively. Acute toxicity was more than five times less if compared with AZT. As a result, we consider this series of carbamate derivatives of AZT as perspective for development of anti‐HIV agents.

A new antiviral: Chimeric 3TC–AZT phosphonate efficiently inhibits HIV-1 in human tissues ex vivo

Although more-recently developed antivirals target different molecules in the HIV-1 replication cycle, nucleoside reverse transcriptase inhibitors (NRTIs) remain central for HIV-1 therapy. Here, we test the anti-HIV activity of a phosphonate chimera of two well-known NRTIs, namely AZT and 3TC. We show that this newly synthesized compound suppressed HIV-1 infection in lymphoid tissue ex vivo more efficiently than did other phosphonates of NRTIs. Moreover, the new compound was not toxic for tissue cells, thus making the chimeric phosphonate strategy a valid approach for the development of anti HIV-1 compound heterodimers.

P-(alkyl)-nucleoside 5'-hydrogenphosphonates as depot forms of antiviral nucleotide analogues.

Abstract P-(Alkyl)esters of AZT 5′-hydrogenphosphonate were synthesized and their stabilities in the phosphate buffer and human serum were evaluated. The esters bearing residues of primary and secondary alcohols were degraded to give AZT, whereas those containing tertiary alkyl groups yielded AZT 5′-hydrogenphosphonate. The corresponding derivatives of d2A and d4T showed the same properties.