Biomedical subjects

T Suzutani

Publications and source records attributed to T Suzutani.

At least 37 records · Page 2Linked to original sources

PubMed1997

Structure-activity relationship of the affinity of 5-substituted uracil nucleoside analogues for varicella-zoster virus thymidine kinase and their activity against varicella-zoster virus.

We investigated structure-activity relationships of 5-substituted uracil nucleoside analogues for their selective antiviral activity against varicella-zoster virus (VZV) and affinity for VZV thymidine kinase (TK). Anti-proliferative activity of the compounds was measured using human lymphoblastoid cells. Most 2'-deoxyribofuranosyluracil, arabinofuranosyluracil (araU) and 2'-deoxy-2'-fluoro-arabinofuranosyluracil derivatives showed selective anti-VZV activity as well as activity against herpes simplex virus types 1 and 2. 2'-Deoxyuridine derivatives showed higher affinity than the corresponding araU analogues. A correlation was seen between the 50% effective doses for VZV and the Ki values for VZV TK, except for 5-ethyl-2'-deoxyuridine and 5-ethyl araU that showed relatively high affinity for VZV TK without showing any activity against VZV. 5-Halogenovinyluracil nucleosides showed the highest affinity and the most potent and selective anti-VZV activity. 2'-Deoxy-2'-fluoro-arabinofuranosyluracil derivatives exhibited high anti-VZV potency though they showed relatively low affinity for VZV TK. Some 3'-deoxythymidine analogues having anti-human immunodeficiency virus activity were inactive against herpesviruses.

Antineoplastic Agents↗

PubMed1997

Role of gamma delta TCR+ lymphocytes in the augmented resistance of trehalose 6,6'-dimycolate-treated mice to influenza virus infection.

Trehalose 6,6'-dimycolate (TDM), an immunomodulator, potentiates non-specific resistance in mice to influenza virus infection. When mice were injected intravenously with TDM, the striking proliferation of a minority of T-lymphocytes bearing gamma/delta T-cell receptors (gamma delta T-cells) that accumulated in granulomatous lungs was thought to be associated with the maintenance of acquired resistance to lethal influenza virus infection. To clarify the cellular basis of the defence against influenza virus, mice were depleted of gamma delta T-cells, alpha/beta (alpha beta) T-cells, or natural killer (NK) cells by in vivo administration of corresponding antibodies prior to influenza virus infection. The depletion of gamma delta T-cells significantly abrogated the augmented resistance of TDM-treated mice to infection, as did depletion of either alpha beta T-cells or NK cells. To gain insight into the functional ability of gamma delta T-cells, we evaluated the cytotoxic activity of this T-cell subset against a panel of target cell lines that were stably transfected with the influenza virus haemagglutinin (HA) gene from A/PR/8/34(H1N1) and A/Aichi/2/68(H3N2) strains. The gamma delta T-cells from TDM-treated mice showed profound cytotoxicity against the target cells expressing HA of either the H1 or H3 subtype, in a non-major histocompatibility complex-restricted manner. Taken together, these results indicate that gamma delta T-cells play a non-specific role, in conjunction with alpha beta T-cells and NK cells, in protecting mice against influenza virus infection, and that the recognition and destruction of HA-expressing target cells by the activated gamma delta T-cells is one of the steps involved in this anti-influenza virus immunosurveillance.

Adjuvants, Immunologic↗

PubMed1996

A case of varicella-zoster myelopathy.

INTRODUCTION: Early diagnosis of neurological complications of varicella-zoster virus (VZV) is important because of its treatability. We performed polymerase chain reaction (PCR) to detect VZV-DNA from the cerebrospinal fluid (CSF) of a patient with myelopathy. PATIENT & METHODS: A 69-year-old man developed sensory disturbances in the lower extremities and bladder-bowel disturbances, followed by cutaneous zoster on his left arm. Polymerase chain reaction was applied to identify the viral DNA in CSF. RESULTS: The increased antibody index of VZV and herpes simplex virus (HSV) in the CSF suggested intrathecal synthesis of IgG antibodies to these viruses. VZV-DNA was detected in the CSF by nested PCR, but neither HSV-1 nor HSV-2 DNA was detected in CSF. He was successfully treated with acyclovir and prednisolone. CONCLUSION: PCR may be a useful tool for the diagnosis of VZV myelopathy.

Aged↗

PubMed1996

Analysis of phosphorylation pathways of antiherpesvirus nucleosides by varicella-zoster virus-specific enzymes.

The inhibitory activities of acyclovir (ACV), 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU), ganciclovir (GCV), 9-(2-deoxy-2-hydroxymethyl-beta-D-erythro-oxetanosyl)guanine (OXT-G), and (+)-9-[(1R,2R,3S)-2,3-bis(hydroxymethyl)Cyclobutyl]guanine (cOXT-G) on the replication of wild-type and thymidine kinase (TK)-negative strains of herpes simplex virus types 1 and 2 and varicella-zoster virus (VZV) and the wild-type strain of human cytomegalovirus were tested to clarity whether the phosphorylation of these compounds is catalyzed by viral TK or other enzymes. ACV and BV-araU had little effect on the replication of TK-negative virus strains. On the other hand, GCV, OXT-G, and cOXT-G inhibited the replication of TK-negative VZV at concentrations 10 times higher than those at which they inhibited wild-type VZV, indicating that a kinase other than TK phosphorylates GCV and OXT-G in VZV-infected cells. GCV phosphorylation activity was not detected in VZV-infected cell lysates; therefore, this activity was evaluated in COS 1 cells expressing viral TK and viral protein kinase (PK). The COS 1 cells expressing VZV TK were shown to be susceptible to all compounds tested. In contrast, VZV Pk-expressing COS 1 cells were susceptible to only GCV, OXT-G, and cOXT-G. These results suggest that VZV PK phosphorylates some nucleoside analogs, for example, GCV, OXT-G, and cOXT-G. This phosphorylation pathway may be important in the anti-VZV activities of some nucleoside analogs.

Acyclovir↗

PubMed1996

Relationship of the strain and the intraocular amount of herpes simplex virus types 1 and 2 in the induction of anterior-chamber-associated immune deviation.

We investigated the induction of anterior-chamber-associated immune deviation (ACAID) by herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2). Two strains of HSV-1 (VR-3 and F) and four strains of HSV-2 (UW-268, 186, SL, and MS) were inactivated with ultraviolet light, and the anterior chamber of BALB/c mice was inoculated with a viral strain. Immunization against delayed-type hypersensitivity (DTH) was done subcutaneously on day 7 after anterior chamber inoculation. On day 10 after DTH immunization, each virus was inoculated into the foot pad. DTH was evaluated by measuring the foot pad thickness at 24 h after the inoculation into the foot pad. DTH was suppressed by anterior chamber inoculation with more than 10(3) plaque-forming units (PFU) of the VR-3, UW-268, 186, and SL strains, and with 10(5) PFU of the F strain, but not by the MS strain. Until now, HSV-2 was believed not to induce ACAID, however, our results indicate that some HSV-2 strains induce ACAID.ACAID induction by HSV-1 and HSV-2 may depend on both the viral strain and the amount of ocular viral antigen.

Animals↗

PubMed1996

Meisenheimer rearrangement of azetopyridoindoles. VIII. Synthesis and antiviral activities of 12-carbaeudistomin analogs.

Eudistomins, isolated from the colonial tunicate Eudistoma olivaceum, have been a synthetic target due to their strong antiviral activity against Herpes simplex virus (HSV-1) and activities against certain types of tumors in vivo. In order to examine the structure-activity relationship of eudistomins, 12-carbaeudistomin analogs were synthesized and their activities against influenza A and B virus, HSV-1, HSV-2 and human cytomegalovirus were investigated. Among them, racemic 6-methoxy-12-carbaeudistomin showed similar activity to (-)-debromoeudistomin K, synthesized as a control compound.

Animals↗

PubMed1995

A new mutation in exon 12 of the gp91-phox gene leading to cytochrome b-positive X-linked chronic granulomatous disease.

We have previously reported a patient with cytochrome b-positive X-linked chronic granulomatous disease. Although the O2- production of neutrophils from the patient was completely defective, we presented data suggesting that the patient's cytochrome b was present at a normal level and possibly had normal spectroscopic features. Thus, to look for a mutation in the cytochrome b heavy chain (gp91-phox) gene, DNA analysis of gp91-phox cDNA derived from this patient was performed. As a result, we found that five nucleotides (1521 through 1525) within exon 12 were deleted, and a new sequence of eight nucleotides was inserted. This mutation converted Gln507-Lys508-Thr509 into His-Ile-Trp-Ala. Mismatched polymerase chain reaction showed that the mother has both wild and mutated alleles, confirming that this case was transmitted in an X-linked fashion. This mutation is different from those previously reported by others. The translocation of p47-phox and p67-phox to the membrane fraction occurred, indicating the complete formation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex. We conclude that this case suggests that the structure encoded on exon 12 of gp91-phox is important for electron transfer.

Adult↗

PubMed1995

Analysis of the relationship between cellular thymidine kinase activity and virulence of thymidine kinase-negative herpes simplex virus types 1 and 2.

The virulence of thymidine kinase-negative herpes simplex virus type 1 (HSV-1; VRTK- strain) and type 2 (HSV-2; UWTK- strain) was studied in comparison with that of their parental strains (VR-3 and UW-268, respectively) in an encephalitis model of adult (4-week-old) and newborn (3-day-old) mice. Viral thymidine kinase (TK) activity was essential for the maximum expression of virulence of HSV-1, because the 50% lethal dose (LD50) of VRTK- was 60 times higher than that of VR-3 in the brains of newborn mice expressing high levels of cellular TK activity. However, the UWTK- strain showed that replication of the UWTK- strain was completely supported by cellular TK activity. This difference in the role of viral and cellular TKs for virus growth between HSV-1 and HSV-2 was¿ confirmed with the one-step growth of virus strains in L-M and L-M(TK-) cells.

Amino Acid Sequence↗

PubMed1995

Identification of varicella-zoster virus strains by PCR analysis of three repeat elements and a PstI-site-less region.

We established a method of identifying varicella-zoster virus (VZV) strains, especially those of the Oka vaccine, in patients with clinical VZV infections. The DNAs of 30 clinically isolated strains and 4 laboratory strains including the Oka vaccine strain and its parent VZV strain, were analyzed by PCR with four sets of primers for the four variable regions, R2, R4, R5, and a region without a PstI site (PS). R4 was unstable in four laboratory VZV strains and was excluded from the study. The other regions were stable in several passages in cell culture. The number of copies in R2 and R5 were distributed from 2 to 13 and from 1 to 3, respectively, in the strains analyzed. The vaccine strain had seven copies in R2 and two copies in R5, and it was PS negative. Among 30 clinical isolates, 3, 23, and 11 strains had the same characteristics as the vaccine strain in R2, R5, and PS, respectively. Therefore, by this method, 97.2% of the isolates were distinguished from the Oka vaccine strain. This strategy will be useful in diagnosing VZV infections induced by the vaccine strain.

Adult↗

PubMed1993

Kinetic studies of the predicted substrate-binding site of varicella-zoster virus thymidine kinase.

To investigate the mechanism of kinetic action and substrate recognition of varicella-zoster virus (VZV) thymidine kinase (TK), we designed and isolated a site-directed mutant VZV TK which has double amino acid substitutions, 136threonine to leucine and 137isoleucine to leucine (SDM TK). This mutant was designed to alter the substrate-binding site of the VZV TK to duplicate that of the herpes simplex virus type 2 enzyme. Kinetic studies of the activity of wild-type TK indicated that the binding order of ATP and thymidine is random and that wild-type VZV TK possessed high thymidylate kinase (TM-K) activity. The sensitivity of VZV TK to bisubstrate analogues, dinucleotides of adenosine and thymidine, showed that the optimum distance between the ATP- and substrate-binding sites is two phosphoryl groups greater than with the natural substrate for TK activity. SDM TK lost deoxycytidine kinase activity and had reduced TK and TM-K activities. Inhibition studies on both WT and SDM TK by 5-halogenovinyluridine analogues and their 5' monophosphate derivatives revealed that amino acids at positions 136 and 137 are involved in substrate binding, probably through a role in the formation of the binding pocket for bulky substrates.

Amino Acid Sequence↗

PubMed1993

Antiviral activity of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil against thymidine kinase negative strains of varicella-zoster virus.

Mechanism of antiviral activity of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU) against the YSR strain of varicella-zoster virus (VZV), which is a mutant derived from the wild YS strain and is completely deficient in viral thymidine kinase (TK), was searched in comparison with antiviral activity of other thymidine analogues, guanosine analogue and thymidylate synthase (TS) inhibitor in human embryo lung fibroblast cells. Thymidine analogues, such as BV-araU,5-iododeoxyuridine (IUDR), 1-beta-D-arabinofuranosylthymine (araT), and guanosine analogue, such as 9-(2-hydroxyethoxymethyl)guanine (ACV), showed higher antiviral activity to the YS strain than to the YSR strain. Though, BV-araU also had the antiviral activity of a microgram level against the YSR strain. In contrast to these results, TS inhibitor, 5-fluorodeoxyuridine (FUDR), had higher antiviral activity to the YSR strain than to the YS strain. Highly synergistic antiviral activities of FUDR to the YS strain and the YSR strain were observed in combination with IUDR, araT, or ACV. However, weakly synergistic or additive inhibition to the YSR strain was shown in combination of BV-araU and FUDR, in spite of highly synergistic effect of this combination to the YS strain. The viral and cellular TS activity was partially inhibited by BV-araU monophosphate, but not by BV-araU. These results indicate that BV-araU is converted into BV-araU monophosphate by cellular TK, and the inhibition of TS activity by BV-araU monophosphate in the YSR strain-infected cells results in the suppression of viral replication.

Antiviral Agents↗

PubMed1993

Mechanism of inhibition of DNA synthesis by 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil.

The mechanism of the inhibitory action of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl) uracil triphosphate (BV-araUTP) on DNA synthesis by Escherichia coli DNA polymerase I Klenow fragment was studied. Acting as a chain terminator, BV-araUTP inhibited DNA synthesis by Klenow fragment more effectively than 2',3'-dideoxythymidine triphosphate (ddTTP). However, the incorporation sites of BV-araU monophosphate were restricted at consecutive dTMP sequence whereas ddTMP was incorporated at every dTMP site.

Antiviral Agents↗

PubMed1993

[Antiviral compounds: current approaches for development].

Recent dynamic advances of molecular biology have been discovering the specific virus functions and replication strategies that differ greatly from their host cells. These studies made it possible to design the antiviral agents which block virus replication specifically. In this paper, we review the recent approach for development of anti-HIV agents and the problems of anti-herpes compounds applied in clinical treatment.

Acyclovir↗

PubMed1992

Analysis of toxic and mutagenic activities of antiherpesvirus nucleosides against HeLa cells and herpes simplex virus type 1.

The toxic and mutagenic activities of five antiherpesvirus agents to HeLa cells and herpes simplex virus type 1 (HSV-1) were investigated. 5-Iodo-2'-deoxyuridine (IDU) and 9-beta-D-arabinofuranosyl-adenine (araA) showed very potent inhibitory effects on cell growth and the cloning efficiency of HeLa cells, whereas 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU), E-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) and 9-(2-hydroxyethoxymethyl)guanine (ACV) showed less inhibitory effect. 50% inhibitory doses of BV-araU and BVDU for cell growth were 657 and 253 micrograms/ml, respectively. Although the growth inhibitory activity of BVDU was very weak, as above, the mutagenic activity of this drug to the cells, estimated by induction of colchicine-resistant mutants, was observed to be 4 micrograms/ml, which was a markedly smaller dose than the inhibitory dose for cell growth, and the highest frequency of mutation of the cells was shown at 100 micrograms/ml of BVDU. This activity was more potent than that of IDU. No mutagenic activity of BV-araU, araA and ACV to cells was observed within the concentration range of 1-800 micrograms/ml. IDU showed high mutagenic activity to HSV-1 growing in human embryo lung fibroblasts, and IDU-resistant mutants were induced at a high frequency. BVDU also induced a small amount of BVDU-resistant mutant virus, although this drug induced many mutant cells. No mutagenic activity of BV-araU, araA and ACV to HSV-1 was observed.

Acyclovir↗

PubMed1992

Random mutagenesis of the thymidine kinase gene of varicella-zoster virus.

To understand the relationship between the primary structure and function of varicella-zoster virus thymidine kinase (VZV TK; EC 2.7.1.21), we established rapid screening and phenotypic selection of mutant VZV TK genes in TK-deficient Escherichia coli C600 by using a constitutive pKK223-3 expression plasmid. In this screening system, mutant TK genes generated by random mutagenesis were identified by the sensitivity of E. coli-expressing VZV TKs to 5-bromo-2'-deoxyuridine and 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl) uracil. Twenty-four mutant clones with amino acid substitutions were isolated, and their nucleotide sequence and enzymatic activities were determined. Of the 24 clones, 20 had single amino acid substitutions, 2 clones had double amino acid substitutions, and 1 clone had triple amino acid substitutions. In 17 cases of single amino acid substitution, six mutations led to lost enzyme activity, and four of these six mutations centered in the ATP-binding site. The other 11 mutations resulted in reduction of both TK and thymidylate kinase activities or only thymidylate kinase activity and were located in scattered positions in the VZV TK gene, although 5 mutations showed a tendency to cluster in the region between positions 251 and 260.

Amino Acid Sequence↗

PubMed1992

Effects of acyclovir, oxetanocin-G, and carbocyclic oxetanocin-G in combinations on the replications of herpes simplex virus type 1 and type 2 in Vero cells.

9-(2-hydroxyethoxymethyl)guanine (acyclovir, ACV) and novel nucleosides, 9-(2-deoxy-2-hydroxymethyl-beta-D-erythro-oxetanocyl)guanine (oxetanocin-G, OXT-G) and (+)-9-[(1R, 2R, 3S)-2, 3-bis(hydroxymethyl)cyclobutyl]guanine (carbocyclic oxetanocin-G, carbocyclic OXT-G) possessed substantial antiviral activities against herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2). ACV inhibited only viral thymidine kinase positive (TK+) herpes viruses, although the latter two compounds inhibited the replications of the TK deficient (TK-) mutants of HSV-1 and HSV-2 as well as the TK+ parent strains in vitro. The TK- mutants of HSV-1 and HSV-2 (HSV-1 TK- and HSV-2 TK-) were as susceptible to OXT-G as the TK parent strains. However, the TK- mutants were less susceptible to carbocyclic OXT-G than the TK+ parent strains. We demonstrated synergistic inhibition of the replications of HSV-1 and HSV-2 by ACV and OXT-G in combination, additive inhibition of HSV-1 and HSV-2 by ACV and carbocyclic OXT-G in combination, synergistic inhibition of HSV-1 by OXT-G and carbocyclic OXT-G in combination, and additive inhibition of HSV-2 by these two compounds. We investigated the metabolism of ACV and OXT-G in HSV-1 TK(+)-, HSV-1 TK(-)- and mock-infected Vero cells by thin layer chromatography. ACV-triphosphate increased more in HSV-1 TK(+)-infected Vero cells than in HSV-1 TK(-)- and mock-infected Vero cells. The metabolism of OXT-G had almost the same pattern in HSV-1 TK(+)-, HSV-1 TK(-)- and mock-infected Vero cells. These results suggest that ACV is phosphorylated by virus-induced TK, and OXT-G is phosphorylated by cellular nucleoside and nucleotide kinases.

Acyclovir↗

PubMed1992

Identification of a cell membrane receptor for interferon induction by poly rI:rC.

PR-RK, a cell line derived from rabbit kidney cells (RK-13), was insensitive to the cytotoxic effect and interferon (IFN) inducing activity of the copolymer of riboinosinic and ribocytidylic acid (poly rI:rC). However, PR-RK was sensitive to the cytotoxic effect of the copolymer of riboadenylic and ribouridylic acid (poly rA:rU). Comparison of PR-RK cells and RK-13 cells by cytofluorometric analysis revealed that the binding of poly rI:rC was considerably reduced on PR-RK cells. These results suggested that the receptor for poly rI:rC might be different from the receptor for poly rA:rU, and this difference could provide a basis for the identification of the dsRNA receptor on cell surface. Western blot analysis of the components of cell membrane fraction prepared from RK-13 cells was performed by using a monoclonal antibody, which binds to cell membrane of RK-13 cells but not to PR-RK cells, and which blocks IFN induction by poly rI:rC in RK-13 cells. The 60K protein was identified as one of the poly rI:rC receptor protein.

Animals↗

PubMed1991

Analysis of mutations in the thymidine kinase genes of drug-resistant varicella-zoster virus populations using the polymerase chain reaction.

We have applied the polymerase chain reaction (PCR) technique to analyse mutations in the thymidine kinase (TK) gene of varicella-zoster virus (VZV) associated with resistance to the 5-bromovinyl (BVaraU) and 5-propynyl (PYaraU) analogues of arabinofuranosyl deoxyuridine. The results from this study allow three clear conclusions to be drawn. Firstly, the technique clearly shows that populations of VZV derived from plaque purification were truly clonal only when the plaques were initiated from cell-free virus (representing a tiny fraction of infectious virus) and plaques initiated by infected cells contained a mixture of variants. Secondly, despite the background mutations caused by errors of the Taq DNA polymerase, mutations relevant to drug resistance can easily be distinguished. The BVaraU-resistant mutant, 7-1, contained an aspartic acid to asparagine mutation at residue 18 and a single base deletion (position 65298 of the VZV DNA sequence), resulting in a frameshift and premature termination of the polypeptide chain, was found in the BVaraU-resistant mutant YSR. PYaraU-resistant virus populations contained viruses with one or more of three independent mutations, i.e. single base substitutions resulting in mutations from leucine to proline at residue 92, histidine to arginine at residue 97 and a deletion of 20bp (residues 65,135 to 65,154). Finally, the technique has uncovered novel sites in the virus TK associated with drug resistance. We conclude that in vitro amplification using the PCR combined with cloning and sequencing is a relatively rapid method for identifying mutations in small virus populations even when they are not homogeneous.

Amino Acid Sequence↗