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Y C Cheng

Publications and source records attributed to Y C Cheng.

At least 181 records · Page 10Linked to original sources
PubMed1989

Characterization of the 3' region of the human DNA topoisomerase I gene.

Previous studies suggest that topoisomerase I (Topo I) plays a critical role in cell growth. However, the structure of the Topo I gene has not yet been determined. Two complementary DNA (cDNA) clones for the human Topo I 4.1-kilobase mRNA were isolated independently from HeLa and KB cell cDNA libraries. These clones were identical and they contained 679 base pairs of coding and 1138 base pairs of noncoding sequences. The clones had a two-base difference in the 3' noncoding region compared to the Topo I cDNA from human placenta. The structure of the 3' end of the human Topo I gene from six human tumor cell lines was examined. The Topo I cDNA recognized 16.5, 24.2, and 16.0 kilobases of genomic DNA restricted with EcoRI, HindIII and PstI, respectively. The individual genomic fragments were ordered by double digestion and hybridization with cDNA subclones. digestion and hybridization with cDNA subclones. The results indicate that the human Topo I gene contains several intervening sequences. The gene arrangement was similar in all six cell lines and no polymorphism was observed. However, each digestion contained genomic fragments that hybridized with all the subclones, suggesting that at least one Topo I pseudogene, or another Topo I gene with a different structure, was present in every cell line. As predicted, double digestions generated at 161 base pair fragment that indicates the presence of an intronless pseudogene. In contrast to the DNA topoisomerase I gene, the presumptive pseudogene(s) appears to be hypomethylated. In addition to the 4.1-kilobase Topo I mRNA, a larger 6-kilobase RNA was identified in human KB and HeLa cells which could be a processed Topo I mRNA intermediate.

Base Sequence↗
PubMed1989

Delayed cytotoxicity and selective loss of mitochondrial DNA in cells treated with the anti-human immunodeficiency virus compound 2',3'-dideoxycytidine.

The compound 2',3'-dideoxycytidine (ddC) is a potent inhibitor of human immunodeficiency virus replication in vitro and is currently in clinical trials for treatment of acquired immunodeficiency syndrome. The compound was found to exert delayed cytotoxicity against Molt-4F cells, a human T lymphoblastic cell line. At a concentration as low as 0.1 microM, the doubling time of the cells was increased after 8 days of ddC treatment. This concentration is 5-fold lower than plasma levels reached in clinical trials. The cells finally died after a 2-week exposure to 0.1 or 0.2 microM ddC. The delayed cytotoxicity was not due to a greater accumulation of 2',3'-dideoxycytidine triphosphate in cells with longer exposure to the compound. The cellular content of mitochondrial DNA was found to decrease and the rate of glycolysis was found to increase with continuous exposure of cells to ddC. The mitochondrial toxicity and cell growth inhibition were reversed when ddC was removed. The reduction in cellular content of mitochondrial DNA caused by ddC may partially explain the delayed toxicity observed in acquired immunodeficiency syndrome patients treated with the drug.

Antiviral Agents↗
PubMed1989

Effect of phosphorothioate homo-oligodeoxynucleotides on herpes simplex virus type 2-induced DNA polymerase.

Effects of phosphorothioate oligodeoxynucleotides of different chain length and base composition on herpes simplex virus (HSV) type 2 (strain 333)-induced DNA polymerase have been examined in vitro. The anti-HSV-2 DNA polymerase activity was related to the base composition of the analogs, with the order of potency: deoxycytidine greater than thymidine greater than deoxyadenosine, for compounds with equal chain length. The potency was also related to oligomer chain length, since it was observed that the longer the chain length, the more potent the inhibition exerted. Among all the compounds tested, the phosphorothioate oligodeoxycytidine 28-mer (S-(dC)28) was the most potent inhibitor of HSV-2-induced DNA polymerase. This inhibition was competitive with an activated DNA template with a Ki value of 7 nM. It was also a competitive inhibitor of the DNA polymerase-associated exonuclease activity with a Ki value of 5 nM. In contrast, this compound showed less inhibition of human DNA polymerase alpha, beta, and gamma, as well as HSV-1 (strain KOS) and Epstein-Barr virus-induced DNA polymerase. The possibility that S-oligomers can serve as primers for DNA elongation was also investigated. Poly(dG).S-(dC)28 and poly(dA).S-(T)28 are poor substrates for DNA elongation catalyzed by HSV-2 DNA polymerase. In summary, phosphorothioate oligonucleotides could be anti-template inhibitors of HSV DNA polymerase. This information may lead to the development of a new class of selective anti-HSV agents.

Animals↗
PubMed1989

Human immunodeficiency virus reverse transcriptase-associated RNase H activity.

Biochemical characteristics of the RNase H activity associated with immunoaffinity purified human immunodeficiency virus (HIV) reverse transcriptase (RT) were examined. Glycerol gradient centrifugation of HIV RT resulted in a single peak of RNase H, associated with RT activity, with an apparent molecular weight of 110,000. HIV RNase H exhibited a marked substrate preference for poly(dC).[3H]poly(rG) compared to poly(dT).[3H]poly(rA). It did not hydrolyze single-stranded RNA or the DNA component of DNA.RNA hybrids. Products of the HIV RT-associated RNase H reaction consisted primarily of monomers, dimers, and trimers with 3' OH groups. This reaction was Mg2+ dependent, with greater than 90% of maximum activity at MgCl2 concentrations between 4 and 12 mM. The optimum KCl concentration for HIV RT catalyzed polymerization with a poly(rA).(dT)10 template. The optimum pH for HIV RNase H activity was between 8.0 and 8.5, in contrast to an optimum pH of 7.5 to 8.0 for HIV RT activity. The association of RNase H activity with the p66 component of HIV RT was demonstrated by activity gel analysis. These results indicate that HIV RT has an integral RNase H activity; however, some of its properties are different from those of RNase H associated with other retroviral RT's, and optimal assay conditions are different than those for HIV RT catalyzed DNA polymerization.

Centrifugation, Density Gradient↗
PubMed1989

Interaction of several nucleoside triphosphate analogues and 10-hydroxycamptothecin with human DNA topoisomerases.

DNA topoisomerase I (Topo I) can exist in several different molecular weight forms in human leukemic cells. The Mr 98,000 form of Topo I was inhibited by several nucleoside triphosphates and their analogues at a 500 microM concentration in the order: dideoxy-GTP greater than 2-bromo-dATP greater than dideoxy-ATP greater than dideoxy-CTP greater than 2-fluoro-dATP greater than 2-chloro-dATP. The same concentration of these nucleoside triphosphates also inhibited the Mr 32,000 and the Mr 35,000 Topo I forms in the order: 2-bromo-dATP greater than dideoxy-GTP greater than 2-fluoro-dATP greater than dideoxy-ATP; however, dideoxy-CTP and 2-chloro-dATP did not inhibit these forms. ATP inhibited both the large and the small molecular weight forms of Topo I at a concentration of 8 mM. DNA topoisomerase II (Topo II) isolated from human leukemic cells requires ATP for its activity. Of the nucleoside triphosphates examined, only dATP could substitute for ATP. In the presence of 500 microM ATP, equimolar concentrations of 2-bromo-dATP, dideoxy-ATP, 2-chloro-dATP, 2-fluoro-dATP, and dideoxy-GTP nucleotide analogues inhibited the unknotting activity of the Topo II enzyme. When the nucleotide analogue concentration was decreased to 250 microM, only 2-bromo-dATP still had a significant inhibitory effect on Topo II. With the exception of 2-bromo-dATP, the analogues studied appeared to inhibit the nicking step of both the Topo I and Topo II enzyme activity. These results differ from previously described mechanisms of inhibition by camptothecin of Topo I and etoposide of Topo II. These enzymatic studies suggest the inhibition of Topo I and Topo II activities could contribute to the cytotoxicity of the respective nucleoside analogues in cell culture, particularly when high concentrations of these nucleoside analogues accumulate as triphosphates inside the cells.

Adenosine Triphosphate↗
PubMed1989

Phenotypic instability of drug sensitivity in a human colon carcinoma cell line.

Colon cancer is one of the tumors most refractory to treatment by chemotherapy, and this may be due to inherent phenotypic instability of such tumor cells with respect to the biochemical determinants of drug sensitivity. To test this hypothesis, a clonal human colon carcinoma cell line, clone A, was passaged in culture in the absence of selection conditions or mutagens. During this time, sensitivity to several drugs was examined, and was found to decrease 4-fold during 30 weeks of culture. Five randomly selected subclones, having never been exposed to drug or mutagen, displayed a range of sensitivities to etoposide (50% inhibitory concentrations ranging from 1.5 to 4.9 microM) and to vincristine (9-fold range), but all had the same sensitivity to methotrexate. With time these sensitivities also changed, and subsequent subclones were chosen from the lines with highest and lowest drug sensitivity. Again a wide range of phenotypes was observed. Sensitivity to vincristine ranged 14-fold and to doxorubicin 3-fold. Several biochemical determinants of drug sensitivity had a broad range of expression between cell lines. Cellular accumulation of [3H]vincristine, as well as expression of multidrug resistance protein P170 and glutathione transferase activity all varied significantly between subclonal lines. This suggests that some human colon tumors may be phenotypically unstable with respect to drug sensitivity, and this could contribute to clinical resistance to chemotherapeutic compounds.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1989

Inhibition of human immunodeficiency virus reverse transcriptase by 2',3'-dideoxynucleoside triphosphates: template dependence, and combination with phosphonoformate.

The 2',3'-dideoxynucleoside triphosphates (ddNTPs) are potent substrate analog inhibitors of human immunodeficiency virus (HIV) reverse transcriptase and have clinical utility in the treatment of acquired immunodeficiency syndrome. Several issues regarding the interaction of these compounds with HIV reverse transcriptase were examined. The potency of unsubstituted ddNTPs and the 3'-azido analog of dTTP (AZTTP) was influenced by the choice of template. Both compounds were more potent with the complementary homopolymer templates than with gapped duplex DNA, although the Km for the competing dNTP was similar with different templates. The Ki for AZTTP was greater than for the unsubstituted ddNTPs with either a homopolymer or a gapped duplex DNA template. HIV reverse transcriptase incorporated ddCMP and AZTMP into primed phage m13 DNA at sites specified for insertion of dCMP and dTMP, respectively. ddCTP was more efficiently utilized as a substrate than was AZTTP. Primer elongation due to base misincorporation was observed in the absence of one dNTP. The combined effect of ddNTPs and the pyrophosphate analog phosphonoformate (PFA) on HIV reverse transcriptase was also examined, and inhibition by PFA in combination with ddTTP or AZTTP was mutually exclusive.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Enhancement of herpes simplex virus type 2 (HSV-2) DNA synthesis in infected cells that constitutively express the BglII-N region of the HSV-2 genome.

The BglII-N fragment of the herpes simplex virus type-2 (HSV-2) genome encodes one of two known transforming regions of this DNA virus. In this study, we report the derivation of HeLa S3 cells (2DC4) that stably express the HSV-2 BglII-N region, including the small subunit of HSV-2 ribonucleotide reductase (RR). Superinfection of the 2DC4 cells with wild-type HSV-2 resulted in the efficient induction of HSV-2-encoded ICP10, DNA polymerase, and thymidine kinase. The amount of HSV-2 DNA synthesis in 8-hr HSV-2-infected 2DC4 cells was enhanced 2.6 +/- 0.6-fold relative to infected control cells. Furthermore, the replication kinetics of HSV-2 DNA in 2DC4 cells were accelerated relative to HeLa S3 cells; HSV-2 DNA synthesis was detectable as early as 3 hr postinfection in 2DC4 cells as compared to 6 hr postinfection in HeLa S3 cells. These results suggest that the BglII-N region of HSV-2 encodes function(s) that activate the viral DNA synthesis apparatus and that this activation could relate to the transforming ability of this DNA region.

Base Sequence↗
PubMed1989

Human topoisomerase 1 messenger RNA is not destabilized by the herpes simplex virus type 2 virion-associated shut-off function.

A cDNA for human topoisomerase I (Topo 1) was used to identify a 4.1 kb polyadenylated Topo 1 mRNA in methotrexate-resistant human KB cells that are permissive for herpes simplex virus type 2 (HSV-2) infection. Using these cells, no effect of the HSV-2-associated early shut-off function on levels of Topo-1 mRNA was observed up to 6 hours postinfection, whereas the actin mRNA level was 22% cellular transcripts are susceptible. The level of several host-cell polyadenylated RNAs detected as cDNA clones (class 3 transcripts) were unchanged 8 hours after HSV-2 infection, and other cellular transcripts (class 2) actually accumulated at postinfection.

Cell Transformation, Viral↗
PubMed1989

Antitumor agents. 100. Inhibition of human DNA topoisomerase II by cytotoxic ether and ester derivatives of podophyllotoxin and alpha-peltatin.

A principal mechanism of action of the clinical antitumor drugs etoposide (1) and teniposide (2) is the inhibition of catalytic activity of type II DNA topoisomerase and concurrent enzyme-mediated production of lethal DNA strand breaks. Substitution of the glycosidic moiety of 1 or 2 by ester and ethers, as well as the esterification and etherification of alpha-peltatin (4) including its glucosidic ethylidene and thenylidene cyclic acetals (25 and 26), has afforded compounds of much less activity than that of 1. The in vitro cytotoxicity (KB) appears to have no correlation with the inhibitory activity of the human DNA topoisomerase II.

Antineoplastic Agents, Phytogenic↗
PubMed1989

Anti-herpes simplex virus activity of 5-substituted 2-pyrimidinone nucleosides.

Several 5-substituted 2-pyrimidinone 2'-deoxyribonucleoside (PdR) analogs were examined for their anti-herpes simplex virus (HSV) activity in cell culture. The order of potency of their antiviral activities against HSV type 1 (HSV-1) and HSV-2 was iodo PdR approximately ethynyl PdR approximately propynyl PdR. The antiviral action of iodo PdR is dependent on the ability of HSV to induce virus-specified thymidine kinase in infected cells. Several HSV-1 variants with altered thymidine kinase changed their sensitivity to iodo PdR, whereas HSV-1 variants with altered DNA polymerase were as sensitive as the parental virus to iodo PdR. Continuous presence of iodo PdR for more than one virus replication cycle was required for optimal antiviral activity. Iodo PdR (100 microM) had no activity against Epstein-Barr virus DNA replication in P3HR-1 cells. With an oral, an intraperitoneal, or a subcutaneous route of injection, iodo PdR administered twice a day for 2.5 days could prevent the death of mice infected with HSV-2. This in vivo activity is unlikely to be related to the potential conversion of iodo PdR to iododeoxyuridine, since iodo PdR is not a substrate of xanthine oxidase.

Animals↗
PubMed1989

Podophyllotoxin analogs: effects on DNA topoisomerase II, tubulin polymerization, human tumor KB cells, and their VP-16-resistant variants.

Several derivatives of podophyllotoxin with modifications at the C-4 position of ring C, in addition to demethylation at the C-4' position of ring E, were examined for inhibitory activity against DNA topoisomerase II and tubulin polymerization, generation of protein-linked DNA breaks, and cytotoxicity against KB cells and VP-16-resistant KB variants. Substitution of podophyllotoxin with a group in the beta configuration at the C-4 position of ring C resulted in compounds with greater inhibitory activity against DNA topoisomerase II and lower inhibitory activity against tubulin polymerization than those with an alpha configuration. These active analogs exhibited the same mechanism of DNA topoisomerase II inhibition as the epipodophyllotoxin derivative VP-16, which causes protein-linked DNA breaks in vitro as well as in cells. Two analogs selectively inhibited DNA topoisomerases II to a greater extent than tubulin polymerization. These analogs were cytotoxic towards KB cells in addition to VP-16-resistant KB cell lines, which indicated limited cross-resistance with VP-16 in VP-16-resistant KB variants.

DNA Damage↗
PubMed1989

[Pulmonary barotrauma during anesthesia].

There were three cases of pulmonary barotrauma during anesthesia. The causes of barotrauma were: 1) Undue length of the tube pressed by machine's wheel which connect the ventilator to the anesthesia machine. 2) Inadvertent connection of the breathing tube to the inspiratory side of the machine when using the Bain system. 3) Inadvertent placement of expiratory valve. All resulted in obstruction to air outflow. The condition further aggravated by repeated flushing of the oxygen flush valve, leading to rapid increase in intraluminal pressure and rupture of alveolar. The condition can be rapidly recognized by palpation of the neck, auscultation of breathing sound, and finally, with a portable chest X-ray. When any problem exists in the breathing system of anesthesia machines, disconnecting the patient from the machine is mandatory. The patient can be ventilated with an Ambu bag while checking the system thus lessening the incidence of barotrauma.

Adult↗
PubMed1989

[Effects of the specific gravity on spinal anesthesia with 0.5% tetracaine].

In a double-blind study, 0.5% tetracaine 10 mg in hyperbaric or isobaric solution was administered intrathecally at random to 102 adult patients. Except the variable under study, identical technique was used for every patient enrolled. Other influential factors were also under careful control. The extent and pattern of anesthesia were compared. The time for maximal spread was short, the mean times was 7.74 min and 7.8 min for hyper-and isobaric groups respectively, there was no differences statistically. (P less than 0.05). The maximum cephalad spread was T5 (mean, range T12-T3) for the hyperbaric solution and T7 (mean, range T10-T3) for the isobaric solution. The difference was 2 dermatomes and was statistically significant (P less than 0.01). Forty three patients (86%) in the hyperbaric group and 16 patients (30%) in the isobaric group had a sensory blockade higher than T6 level. The hyperbaric group has a much higher percentage of high blockade. The isobaric group has a more predictory lower blockade, but occasionally blockade up to T3 did occur. The 2-segment regression times were also statistically significant. That of hyperbaric group was 88.9 min (mean, range 60-150 min) and 180.5 min (mean, 75-270 min) in isobaric group with a P value less than 0.01. Statistically significant differences in regard to maximum motor blockade and its onset times were found between the two groups. Maximum motor blockade assessed by modified Bromage scale was degree 2 (mean, range 1-3) for the hyperbaric group and degree 3 (mean, range 2-3) for the isobaric group.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1988

Combined modalities of resistance in etoposide-resistant human KB cell lines.

The alkaloid derivative 4'-demethylepipodophyllotoxin 9-(4,6-O-ethylidene)-beta-D-glucopyranoside (etoposide, VP-16) is believed to exert cytotoxicity by causing double-stranded DNA breaks through interruption of the breaking-resealing reaction of topoisomerase II (topo II). Thus it was conceivable that cells could become resistant to VP-16 by a decrease in topo II enzyme level, since this would lead to fewer DNA breaks. As well, given the structure of VP-16, it was also possible that a pleiotropic mechanism of resistance could decrease sensitivity to this drug. To study these possibilities, a series of VP-16-resistant human KB cell lines was established by stepwise selection. The concentrations of VP-16 required to inhibit cell proliferation by 50% in the parent line and KB/1c, KB/7d, KB/20a, and KB/40a lines were, respectively, 0.16, 4.7, 24, 31, and 47 microM. These cell lines expressed cross-resistance to 4'-(9-acridinylamino)methanesulfon-m-anisidide, doxorubicin, vincristine, and methotrexate, although the pattern of relative drug sensitivity was quite different from that of pleiotropic resistant cell lines reported elsewhere. The resistance to vincristine and methotrexate did not increase above the level of the KB/1c cells, and resistance to VP-16, doxorubicin, and especially vincristine was unstable in VP-16-resistant cells cultured in the absence of drug. Although the drug resistance marker Mr 180,000 glycoprotein could not be detected in any of our cell lines, cellular accumulation of [3H]VP-16 was reduced 50-75% in the resistant lines compared with parent KB. With increasing VP-16 resistance, the level of topo II protein, detected by antibody staining, decreased at each step of selection, concomitant with a general decrease in topo II unknotting activity. Sensitivity of the topo II unknotting assay to inhibition by VP-16 was the same for the parent and all resistant cell lines. The level of topo I activity and enzyme increased slightly in the resistant cells. Thus, these cell lines are resistant to VP-16 by virtue of at least two mechanisms: (a) reduced levels of topo II, which confers cross-resistance to other compounds which are topo II-dependent cytotoxic agents; and (b) reduced accumulation of drug, which is likely also responsible for vincristine and methotrexate resistance. However, the possible existence of other mechanisms of resistance cannot be ruled out.

DNA Damage↗