Cellular thymidine kinase

In particular, acyclovir 1 has become important because of its potent antiherpes activity. Interest in this compound was further enhanced by the finding that, in HS V infected cells, its mechanism of action is due to its specific phosphorylation by the viral-encoded (but not cellular) thymidine kinase to acyclo GMP followed by further phosphorylation by cellular GMP and GDP kinases to the triphosphate, acyclo GTP, which is a selective and potent inhibitor of the HSV DNA polymerase.DHPA (3) and DHPG (2) "" also show significant antiviral activity. These findings have stimulated a widespread interest in. and led to the synthesis of. a large number of purine (and pyrimidine) acyclo nucleosides. " "... 

BVdU differs from IdU and F TdU by being specifically phosphorylated in the 5 -position by herpes simplex vims type-1 (HSV-1) induced thymidine kinase. This restricts its action to cells infected by HSV-1. It is less active against genital herpes (HSV-2). HSV-l-induced thymidine kinase converts BVdU to the corresponding 5 -mono- and diphosphate, but HSV-2-induced thymidine kinase stops at the stage of the 5 -phosphate of BVdU. Apparendy, cellular kinases phosphorylate BVdU-5 -diphosphate to the corresponding 5 -triphosphate, which inhibits HSV-1 DNA polymerase to a greater extent than similar cellular DNA polymerases. 

FIAC also strongly inhibits HCMV and Epstein-Barr vims (EBV) in vitro the two vimses known not to induce a specific viral thymidine kinase for their repHcation. However, HCMV may stimulate cellular kinases that can anabolize FIAC to its 5 -triphosphate, which specifically inhibits the HCMV-encoded DNA polymerase. This selective activity suggests that FIAC should be evaluated against HCMV infections. FIAC-ttiphosphate incorporated into DNA has shown strong in vitro activity against the DNA polymerases of human hepatitis B vims (HBV) and of woodchuck hepatitis vims (WHV) (37). 

The antiviral mechanism of action of acyclovir has been reviewed (72). Acyclovir is converted to the monophosphate in herpes vims-infected cells (but only to a limited extent in uninfected cells) by viral-induced thymidine kinase. It is then further phosphorylated by host cell guanosine monophosphate (GMP) kinase to acyclovir diphosphate [66341 -17-1], which in turn is phosphorylated to the triphosphate by unidentified cellular en2ymes. Acyclovir triphosphate [66341 -18-2] inhibits HSV-1 viral DNA polymerase but not cellular DNA polymerase. As a result, acyclovir is 300 to 3000 times more toxic to herpes vimses in an HSV-infected cell than to the cell itself. Studies have shown that a once-daily dose of acyclovir is effective in prevention of recurrent HSV-2 genital herpes (1). HCMV, on the other hand, is relatively uninhibited by acyclovir. 

All NRTIs, as exemplified for AZT (Fig. 7), act in a similar fashion following their uptake by the cells, they are phosphorylated successively to their 5 -monophosphate, 5 -diphosphate, and 5 -triphosphate form (De Clercq 2002). Unlike the first phosphorylation step in the metabolic pathway of the acyclic guanosine analogues (see above), which is carried out by a virus-encoded enzyme (thymidine kinase), the first as well as the subsequent phosphorylations of the 2, 3 -dideoxynucleosides are carried out by cellular enzymes, that is, a 2 -deoxynucleoside (e.g., dThd) kinase, a 2 -deoxynucleotide (e.g., dTMP) kinase, and a (2 -deoxy)nucleoside 5 -diphosphate (NDP) kinase. 

Famciclovir (Famvir) is the diacetyl ester prodrug of the acyclic guanosine analogue 6-deoxypenciclovir Dena-vir). Penciciovir has activity against HSV-1, HSV-2, VZV, and HB V. After oral administration, famciclovir is converted to penciciovir by first-pass metabolism. Penciciovir has a mechanism of action similar to that of acyclovir. It is first monophosphorylated by viral thymidine kinase then it is converted to a triphosphate by cellular kinases. Penciciovir triphosphate acts as a competitive inhibitor of viral DNA polymerase, but unlike acyclovir, it does not cause chain termination. 

L D. The conversion of penciclovir to its active form requires initial monophosphorylation by viral thymidine kinases, then conversion to its active triphosphate form by cellular enzymes. Thus, the concentration of penciclovir triphosphate is particularly high in cells infected with its target viruses (e.g., HSV, VZV, HBV). Foscarnet is a pyrophosphate analogue that does not require activation. Oseltamivir is a neuraminidase inhibitor that is con-... 

F]-FLT is not or only marginally incorporated into DNA (<2%) and therefore not a direct measure of proliferation [122]. In vitro studies indicated that [ F]-FLT uptake is closely related to thymidine kinase 1 (TK1) activity and respective protein levels [117,118]. p F]-FLT is therefore considered to reflect TK1 activity and hence, S-phase fraction rather than DNA synthesis. Although being a poor substrate for type 1 equilibrative nucleoside transporters (ENT), cellular uptake of [ F]-FLT is further facilitated by redistribution of nucleoside transporters to the cellular membrane after inhibition of endogenous synthesis of thymidylate (TMP) de novo synthesis of TMP) [125]. However, the detailed uptake mechanism of [ F]-FLT is yet unknown and the influence of membrane transporters and various nucleoside metabolizing enzymes remains to be determined. 

Acyclovir (ACV) is not a true nucleoside, because the guanine residue is attached to an open-chain structure, but it mimics deoxyribose well enough for the compound to be accepted as a substrate by a thymidine kinase specified by certain herpes-type viruses. The normal thymidine kinase in mammalian cells does not recognize ACV as a substrate, however, so only virus-infected cells convert ACV to its monophosphate. Once the first phosphate has been added, the second phosphate is added by cellular guanylate kinase several other cellular kinases can add the third phosphate. The triphosphate is a more potent inhibitor of the viral DNA polymerases than of cellular DNA polymerases and also inactivates the former but not the latter. The net result is that ACV has been an effective treatment of, and prophylaxis for, genital herpes. Also it can result in dramatic relief of pain associated with shingles caused by reactivation of latent varicella-zoster virus, and has been successful in many patients with herpes encephalitis. 

Acyclovir requires three phosphorylation steps for activation. It is converted first to the monophosphate derivative by the virus-specified thymidine kinase and then to the di- and triphosphate compounds by the host s cellular enzymes (Figure 49-3). Because it requires the viral kinase for initial phosphorylation, acyclovir is selectively activated and accumulates only in infected cells. Acyclovir triphosphate inhibits viral DNA synthesis by two mechanisms competitive inhibition with deoxyGTP for the viral DNA polymerase, resulting in binding to the DNA template as an irreversible complex and chain termination following incorporation into the viral DNA. 

In the past, various serendipitous discoveries have capitalized on the differential expression of enzymes by host and viral infected cells. For example, the prodrug Acyclovir, used widely for the treatment of herpes simplex and herpes zoster infections, is selectively activated through phosphorylation by viral thymidine kinase to acyclovir monophosphate which is then converted to the triphosphate, which inhibits DNA polymerase, by host cellular enzymes. Similarly several 2, 3 -dideoxynucleoside analogs such as Zidovudine (azidothymidine, AZT) and 2, 3 -didehydro-3 -deoxythymidine (D4T) have potent antiviral activity against human immunodeficiency vims (HIV). These compounds are selectively phosphoiylated intracellularly to the 5 -triphosphate derivatives which inhibit the viral reverse transcriptase. 

Mode of action Like acyclovir, ganciclovir is activated through conversion to the nucleoside triphosphate by viral and cellular enzymes, the actual pathway depending on the virus. Cytomegalovirus is deficient in thymidine kinase, and therefore forms the triphosphate by another route. The nucleotide competitively inhibits viral DNA polymerase and can be incorporated into the DNA to decrease the rate of chain elongation. 

Mode of action AZT must be converted to the corresponding nucleoside triphosphate by mammalian thymidine kinase in order for it to exert its antiviral activity. AZT-triphosphate is then incorporated into the growing chain of viral (but not mammalian nuclear) DNA by reverse transcriptase1. Because AZT lacks a hydroxyl at the 3 position, another 5 -3 phosphodiester linkage cannot be formed. Thus, synthesis of the DNA chain is terminated, and replication of the virus cannot take place. The relative lack of discrimination of the viral reverse transcriptase is believed to favor the introduction of the AZT into the viral-catalyzed process the cellular DNA polymerase is more selective. In addition, the phosphory-... 

Penciclovir [pen SIK lo veer] is an acyclic guanosine nucleoside derivative that is active against herpes simplex virus Types I and II, and against varicella-zoster virus. Penciclovir is only administered topically (Figure 37.8). Penciclovir is monophosphorylated by viral thymidine kinase, and cellular enzymes form the nucleoside triphosphate, which inhibits herpes DNA polymerase. Penciclovir triphosphate has an intracellular half-life 20 to 30 times longer than does acyclovir triphosphate (see p. 365). Penciclovir is negligibly absorbed from topical application, and is well tolerated. Both healing and pain are shortened approximately one-half day in duration, compared to placebo-treated subjects. 

The MLA is used for the detection of point mutations, structural aberrations and aneugenicity. The principle of the assay is that cells deficient in thymidine kinase (TK) due to the tk+/" or tk/ mutation are resistant to the cytotoxic effects of the pyrimidine analogue trifluoro-thymidine (TFT). TK proficient cells are sensitive to TFT. This influences the cellular metabolism and leads finally to an inhibition of further cell division. Thus mutant cells are able to proliferate in the presence of TFT, whereas normal cells, which contain TK, are not. The major advantage of the assay is its ability to detect a broad range of mutagenic events represented by optimal detection of both large and small colonies. 

STAVUDINE ZIDOVUDINE Possibly 1 efficacy -l cellular activation of stavudine and antagonism in vivo. Both are phosphorylated to the active form by thymidine kinase, which preferentially phosphorylates zidovudine therefore causes i phosphorylation of stavudine Avoid co-administration... 

Acyclovir selectively inhibits viral DNA synthesis. It is preferentially activated in virally infected cells. Cellular uptake and initial phosphorylation are facilitated by the herpes virus thymidine kinase. The affinity of acyclovir for HSV... 

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