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Biomedical subjects

Y C Cheng

Publications and source records attributed to Y C Cheng.

At least 19 recordsLinked to original sources
PubMed1997

Altered formation of topotecan-stabilized topoisomerase I-DNA adducts in human leukemia cells.

Topotecan (TPT) is a topoisomerase I (topo I) poison that has shown promising antineoplastic activity in solid tumors and acute leukemia. In the present study, a band depletion assay was used to evaluate the ability of TPT to stabilize topo I-DNA adducts in human leukemia cell lines and in clinical leukemia samples ex vivo. This assay showed that 50% of the cellular topo I in HL-60 human myelomonocytic leukemia cells became covalently bound to DNA at an extracellular TPT concentration of 4 micromol/L. In contrast, in 13 clinical specimens of human leukemia harvested before treatment of patients with TPT, the TPT concentration required to stabilize 50% of the cellular topo I in topo I-DNA complexes ranged from 3 to greater than 100 micromol/L (median, 30 micromol/ L). Flow microfluorimetry showed that cellular TPT accumulation varied over only a twofold range and failed to provide evidence for transport-mediated resistance in the clinical samples. These observations raise the possibility that formation of topo I-DNA adducts is diminished in many specimens of refractory/relapsed acute leukemia by a mechanism that might alter topo I sensitivity to TPT.

Antineoplastic Agents↗
PubMed1997

Dose-escalation and pharmacodynamic study of topotecan in combination with cyclophosphamide in patients with refractory cancer.

PURPOSE: Based on preclinical data that demonstrated synergy between alkylating agents and topoisomerase (topo) I poisons, we determined the maximum-tolerated dose (MTD) of topotecan, using a 5 day bolus schedule, that could be given in combination with a single, fixed dose of cyclophosphamide. Pharmacodynamics of this combination were explored by analyzing biochemical effects of treatment in peripheral-blood mononuclear cells (PBMCs). PATIENTS AND METHODS: Patients with refractory cancer were treated with cyclophosphamide 600 mg/m2 on day 1, followed by topotecan given as a 30-minute infusion for 5 consecutive days. Cycles were repeated every 3 weeks. Once the MTD was defined, granulocyte colony-stimulating factor (G-CSF) was added to the regimen in an attempt to escalate further the dose of topotecan. Plasma concentrations of topotecan were determined during the first treatment cycle by high-performance liquid chromatography. PBMCs were sampled at baseline and throughout the 5-day treatment period for analysis of topo I protein concentrations and to determine drug-induced DNA fragmentation. RESULTS: Twenty-six patients were treated with topotecan at doses that ranged from 0.5 mg/m2/d to 1.2 mg/ m2/d for a total of 74 cycles. Reversible neutropenia was dose-limiting, with mild to moderate suppression of the other blood-cell elements commonly occurring. Transfusions of RBCs and platelets were required in 24% and 7% of treatment cycles, respectively. The most prominent nonhematologic toxicities were fatigue and weight loss. Compared with previously published data in which topotecan was administered alone, cyclophosphamide did not appear to alter the pharmacokinetics of topotecan. Significant increases in topo I concentration were identified in PBMCs following the administration of cyclophosphamide on day 1 and there was a significant decrease in topo 1 during the 5-day course of treatment (P < .01, sign test). DNA fragmentation as a result of drug treatment was identified in 11 of 15 (73%) cycles analyzed. CONCLUSION: For previously treated patients, the recommended dose of topotecan in this schedule is 0.75 mg/m2/d without growth factor support and 1.0 mg/ m2/d if it is administered with G-CSF. Biochemical changes in cells induced by exposure to camptothecins can be measured in vivo and these effects may have important implication in the design of combination therapies and the optimal scheduling of this class of agents.

Adult↗
PubMed1996

Uptake and metabolism of the new anticancer compound beta-L-(-)-dioxolane-cytidine in human prostate carcinoma DU-145 cells.

Beta-L-(-)-dioxolane cytidine [(-)-OddC] is the first nucleoside analogue with the unnatural L configuration shown to have anticancer activity. The transport and metabolism of this unique compound were studied in human prostate carcinoma DU-145 cells. (-)-OddC was translocated rapidly into the cells by both equilibrative-sensitive and -insensitive nucleoside transport systems. Accumulation of (-)-OddCMP, (-)-OddCDP, and (-)-OddCTP occurred in a time- and concentration-dependent manner, with (-)-OddCDP being the major metabolite. Elimination of (-)-OddCTP was biphasic, with an initial t1/2 of 3.5 h and a second phase t1/2 of > 20 h. The incorporation of (-)-OddCTP into DNA was concentration dependent, and toxicity was directly correlated with the amount of (-)-OddCMP present in the DNA. Treatment with (-)-OddC led to the degradation of DNA into large fragments at high concentrations, but internucleosomal laddering was not observed. The rapid membrane permeation of (-)-OddC and prolonged retention of its metabolites may contribute to the potent activity of this compound against DU-145 xenografts.

Antimetabolites, Antineoplastic↗
PubMed1996

Structure--activity relationships of 1-(2-Deoxy-2-fluoro-beta-L-arabinofuranosyl)pyrimidine nucleosides as anti-hepatitis B virus agents.

Since 2'-fluoro-5-methyl-beta-L-arabinofuranosyluracil (L-FMAU) has been shown to be a potent anti-HBV agent in vitro, it was of interest to study the structure-activity relationships of related nucleosides. Thus, a series of 1-(2-deoxy-2-fluoro-beta-L-arabinofuranosyl)pyrimidine nucleosides have been synthesized and evaluated for antiviral activity against HBV in 2.2.15 cells. For this study, L-ribose was initially used as the starting material. Due to the commercial cost of L-ribose, we have developed an efficient procedure for the preparation of L-ribose derivative 6. Starting from L-xylose, 6 was obtained in an excellent total yield (70%) through the pyridinium dichromate oxidation of the 3-OH group followed by stereoselective reduction with NaBH4. It was further converted to the 1,3,5-tri-O-benzoyl-2-deoxy-2-fluoro-alpha-L-arabinofuranose (10), which was then condensed with various 5-substituted pyrimidine bases to give the nucleosides. Among the compounds synthesized, the lead compound, L-FMAU (13), exhibited the most potent anti-HBV activity (EC50 0.1 microM). None of the other uracil derivatives showed significant anti-HBV activity up to 10 microM. Among the cytosine analogues, the cytosine (27) and 5-iodocytosine (35) derivatives showed moderately potent anti-HBV activity (EC50 1.4 and 5 microM, respectively). The cytotoxicity of these nucleoside analogues has also been assessed in 2.2.15 cells as well as CEM cells. None of these compounds displayed any toxicity up to 200 microM in 2.2.15 cells. Thus, compound 13 (L-FMAU), 27, and 35 showed a selectivity of over 2000, 140, and 40, respectively.

Antiviral Agents↗
PubMed1996

Establishment and characterization of a human-papillomavirus negative, p53-mutation negative human cervical cancer cell line.

A human cervical cancer cell line, CX, was established from a patient with squamous cell carcinoma of the uterine cervix. The CX cells were epithelial in morphology with relatively large vesicular nuclei, and prominent nucleoli. Cytoplasmic organelles were generally sparse but tonofilaments were relatively abundant. The cells grew as a compact sheet with close membrane approximation interconnected by desmosome-like junctions. CX cells contained cytokeratin, but not vimentin. Elevated levels of squamous cell carcinoma antigen and carcinoembryonic antigen were detected in the cell supernatants. Population doubling time was estimated to be about 20 h. CX cells were not able to grow in soft agar and not tumorigenic in nude mice. Chromosome analysis revealed that CX cells were heterogeneous and mainly had a female diploid karyotype. Unlike cervical cancer cell lines published previously, CX cells were demonstrated to be human papillomavirus-negative, p53 mutation-negative. Based on the distinct characteristics, CX cell line may prove to be a useful tool for the study of human cervical carcinogenesis.

Antigens, Neoplasm↗
PubMed1996

Inhibition of Epstein-Barr virus replication by a novel L-nucleoside, 2'-fluoro-5-methyl-beta-L-arabinofuranosyluracil.

A novel L-nucleoside analog, 2'-fluoro-5-methyl-beta-L-arabinofuranosyluracil (L-FMAU), was found to be a potent and selective inhibitor of Epstein-Barr virus (EBV) replication. The decrease in the amount of viral production was concentration dependent with a 90% inhibitory concentration of approximately 5 muM. Upon removal of the drug from treated cells, virus production resumed in 21 days. Metabolism studies indicated that L-FMAU could be converted to its mono-,di- and triphosphate metabolites in both EBV producing and non-producing cells than in EBV non-producing cells. The mechanism of selectivity of L-FMAU against EBV producing cells. However, the amount of L-FMAU nucleotides formed was three times larger in EBV producing cells than in EBV non-producing cells. The mechanism of selectivity of L-FMAU against EBV does not appear to be due solely to the preferential phosphorylation of L-FMAU in EBV producing cells. The triphosphate of L-FMAU could not be utilized as a substrate by EBV DNA polymerase or the human DNA polymerases alpha, beta, gamma, or delta. Therefore, the incorporation of L-FMAU residues into viral DNA may not be the mechanism of antiviral activity. This compound appears to have a mechanism of action different from that of any other antiherpes virus nucleoside analogs. In addition, L-FMAU has very low cytotoxicity with 50% inhibition of cell growth occurring at a concentration of 1mM. Given the potent inhibitory activity of this compound against EBV and its inability to be incorporated into cellular DNA, L-FMAU analogs should be explored as a new class of anti-EBV agents.

Arabinofuranosyluracil↗
PubMed1996

RNA synthesis inhibitors alter the subnuclear distribution of DNA topoisomerase I.

The acute effect of RNA and DNA synthesis inhibitors on DNA topoisomerase (topo) I localization within cells was examined. Indirect immunofluorescence revealed that topo I was distributed throughout the nuclei but was concentrated in nucleoli of untreated K562 leukemia cells and A549 non-small cell lung cancer cells. Treatment with the DNA polymerase inhibitor aphidicolin did not alter this distribution. In contrast, 30-60 min after addition of the RNA synthesis inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) at concentrations that inhibited [3H]uridine incorporation into RNA by > or = 50%, topo I was visible throughout the nuclei without nucleolar accentuation. Western blotting and activity assays confirmed that the amount of topo I polypeptide and topo I activity were unaltered by the brief DRB treatment. Within 30 min of DRB removal, topo I relocalized to the nucleoli in the absence or presence of the protein synthesis inhibitor cycloheximide. Collectively, these results suggest a reversible translocation of topo I out of the nucleoli when RNA synthesis is inhibited. Treatment with the topo I poisons topotecan or camptothecin, agents that also inhibit RNA synthesis, likewise caused redistribution of topo I to nonnucleolar regions of the nucleus in a variety of cell types. In DC3F hamster lung fibroblasts, 2.5 microM topotecan or 1.25 microM camptothecin was sufficient to cause this topo I redistribution. In DC3F/C-10 cells that contain a mutant camptothecin-resistant topo I, topo I relocalization required 50-fold higher concentrations of topotecan or camptothecin but not DRB. These observations not only suggest that accumulation of topo I in the nucleolus is related to ongoing RNA synthesis but also raise the possibility of screening for some types of camptothecin resistance at the single-cell level using a rapid immunofluorescence-based assay.

Animals↗
PubMed1996

Antitumor agents. 163. Three-dimensional quantitative structure-activity relationship study of 4'-O-demethylepipodophyllotoxin analogs using the modified CoMFA/q2-GRS approach.

Analogs of 4'O-demethylepipodophyllotoxin are considered as potential anticancer agents. We have applied comparative molecular field analysis (CoMFA) and a novel CoMFA/q2-GRS technique recently developed in our group to identify the essential structural requirements for increasing the ability of these compounds to form cellular protein-DNA complex. In addition, a new method to incorporate different types of probe atoms as part of q2-GRS routine has been developed. The best final model with 101 compounds using a combination of four different sets of probe atoms and charges [C (sp3, +1), C (sp3, 0), H (+1), and O (sp3, -1)] yielded a q 2 of 0.584 and the standard error of prediction of 0.660 at 5 principal components. The steric and electrostatic contour plots of the final model were compared with the DNA phosphate backbone environment of the DNA-4'O-demethylepipodophylltoxin analog complex, which was generated using the X-ray structure of the DNA-nogalamycin complex. The comparison reveals that the CoMFA steric and electrostatic fields are compatible with stereochemical properties of the DNA backbone. The results obtained from this study shall guide our future synthetic efforts.

Antineoplastic Agents↗
PubMed1996

Antitumor agents. 164. Podophenazine, 2'',3''-dichloropodophenazine, benzopodophenazine, and their 4 beta-p-nitroaniline derivatives as novel DNA topoisomerase II inhibitors.

We report here the synthesis and biological evaluation of novel DNA topoisomerase II inhibitors, podophenazine (8), 2'',3'' "-dichloropodophenazine (9), and benzopodophenazine (10), and their 4beta-p-nitroaniline derivatives 13-15. Among these, 4'-0-demethyl-4beta-(4'''- nitroanilino)-4-desoxypodophenazine (13) and 4'-O-demethyl-2'',3''-dichloro-4beta-(4-'''-nitroanilino)-4- desoxypodophenazine (14) were found to inhibit KB cells at sub-micromolar concentrations (IC50 = 0.11 +/- 0.03 and 0.48 +/- 0.17 microM, respectively. Against KB/7d cells (a pleiotrophic multiple drug-resistant subclone selected with etoposide which has reduced level of topoisomerase II), only compound 13 out of a target series maintained activity in the sub-micromolar concentration range with a IC 50 value of 0.56 +/- 0.13 mu M. The differential toxicity ratio for 13 [IC 50 (KB/7d)/IC 50 (KB)] was approximately 5. Unlike etoposide and its congeners, compounds 13 and 14 were found to be weak inhibitors of the catalytic activity of topoisomerase II (IC100 = > 100 and > 150 microM, respectively). In vitro protein-linked DNA complex formation assay revealed that 13 and 14, respectively, induced marginal response (13 at 1 microM, 320.3 +/- 124.5 cpm; 13 at 50 microM, 308.8 +/- 139.9 cpm; 13 at 100 mu M, 446.0 +/- 153.5 cpm) and no response (14 at 1 microM, 104.9 +/- 52.6 cpm; 14 at 50 microM, 103.3 +/- 42.6 cpm; 14 at 100 microM, 101.4 +/- 35.2 cpm) compared to the enzyme control. On the basis of these results, we conclude that the mechanism of enzyme inhibition of these compounds is distinct from that of etoposide and its congeners. We are currently investigating the mechanism(s) of action of compounds 13 and 14 as well as synthesizing other derivatives in order to better characterize structure-activity relationships of this series of compounds.

Antineoplastic Agents↗
PubMed1996

Favorable interaction of beta-L(-) nucleoside analogues with clinically approved anti-HIV nucleoside analogues for the treatment of human immunodeficiency virus.

The combination of L(-)-2',3'-dideoxy-3'-thiacytidine (L(-)SddC, 3TC), L(-)-2',3'-dideoxy-5-fluorocytidine (L(-)FddC), or L(-)-2',3'-dideoxy-5-fluoro-3'-thiacytidine (L(-)(FTC) with 3'-azido-3'-deoxythymidine (AZT) synergistically inhibited replication of human immunodeficiency virus (HIV) in vitro. Similar synergistic activity was also obtained when these compounds were used in combination with 2',3'-didehyro-2',3'-dideoxythymidine (D4T). In terms of 2',3'- dideoxyinosime (ddI) and 2',3'-dideoxycytidine (ddC), only additive anti-HIV activity was observed. None of the beta-L(-) nucleoside analogues had additive toxicity in cell culture, and they could protect against the delayed mitochondrial toxicity associated with AZT, D4T, ddC, and ddI in drug-treated cells. Thus, combinations of beta-L(-) nucleoside analogues with any of the approved anti-HIV drugs could have a potentially beneficial outcome.

Antiviral Agents↗
PubMed1996

Identification of two oligodeoxyribonucleotide binding proteins on plasma membranes of human cell lines.

Two oligodeoxyribonucleotide (oligodN) binding proteins of approximately 100-110 kDa were identified in the plasma membranes of human HL-60, HepG2, H1, and KB cells by a photolabeling technique. Solubilization of cellular membranes with a nonionic detergent did not interfere with the binding of these two proteins to oligodNs, and both proteins were susceptible to serine protease action. The binding affinities of these two proteins to oligodNs were found to be similar; Scatchard plot analysis revealed the Kd for phosphodiester (PO) 21-mer oligodeoxycytidine to be 60 nM and binding sites numbered approximately 1.2 x 10(6)/cell for HepG2 cells. Both phosphorothioate (PS) and PO oligodNs could bind to these two proteins with the binding affinity for PS oligodNs being much stronger than that for PO oligodNs. The binding to oligodNs was affected by the ionic strength of the reaction. Dextran sulfate, tRNA, and double-stranded DNA inhibited the binding of oligodNs, whereas ATP, ADP, AMP, and TTP had no effect. Given their high affinity for oligodNs, these membranes proteins may play an important role in the action of oligodNs.

Base Sequence↗
PubMed1996

Camptothecin resistance involving steps subsequent to the formation of protein-linked DNA breaks in human camptothecin-resistant KB cell lines.

To identify mechanisms of camptothecin (CPT) resistance/toxicity, sublines from a human KB cell line were made resistant to CPT by continuous selection in increasing concentrations of CPT. Two CPT-resistant lines, 100 and 300, were 32- and 54-fold resistant to the growth-inhibitory properties of CPT compared to the KB line. After CPT-free culturing, partial revertant lines were established from each resistant line. These partial revertant lines, 100rev and 300rev, were 2.5- and 3.2-fold resistant to CPT compared to KB. When growth inhibition and toxicity were compared, the resistant lines alone displayed an enhanced cytostatic response to CPT. The resistant and partial revertant lines displayed no cross-resistance to etoposide or cisplatin. Comparisons of topoisomerase I (TOPI) activity, content, and protein-linked DNA break production by CPT revealed that resistant and partial revertant lines had one-half the levels as KB, with TOP1 activity that was equally sensitive to CPT in all cell lines tested. However, double-stranded DNA break induction by CPT was significantly reduced only in the resistant lines. Coincubation with 3-aminobenzamide, an inhibitor of poly(ADP-ribosyl) polymerase, potentiated CPT toxicity in the resistant lines alone, without affecting CPT: TOP1 interactions. Therefore, CPT resistance in the 100 and 300 lines was characterized by factors independent of TOP1, specific for CPT, and attenuated by poly(ADP-ribosyl) polymerase inhibition. This resistant phenotype produced fewer double-stranded DNA breaks and enhanced a cytostatic response to CPT.

Benzamides↗
PubMed1996

Effect of stavudine on human immunodeficiency virus type 1 virus load as measured by quantitative mononuclear cell culture, plasma RNA, and immune complex-dissociated antigenemia.

The antiviral effect of stavudine (2', 3'-didehydro-3'-deoxythymidine) against human immunodeficiency virus (HIV) type 1 was measured in 15 HIV-infected patients at baseline and at weeks 4, 10, 22, 34, and 52 of therapy. Patients received 0.1, 0.5, 1.0, or 2.0 mg/kg/day of stavudine. At all time points examined during the 52 weeks of therapy, the median virus titers in peripheral blood mononuclear cells were decreased 1-2 logs, and median immune complex-dissociated antigen levels were reduced 37%-67% compared with baseline values. Plasma RNA content measured by polymerase chain reaction was reduced approximately 0.5 log from baseline median values at both time points examined (weeks 10 and 52). These data demonstrate that stavudine has a substantial and durable antiviral effect.

Antigen-Antibody Complex↗
PubMed1996

Inhibition of hepatitis B virus by a novel L-nucleoside, 2'-fluoro-5-methyl-beta-L-arabinofuranosyl uracil.

2'-Fluoro-5-methyl-beta-L-arabinofuranosyl uracil (L-FMAU) was discovered to have potent antiviral activity against hepatitis B virus (HBV). L-FMAU was more potent than its D-enantiomer and produced dose-dependent inhibition of the viral DNA replication in 2.2.15 cells (human HepG2 cells with the HBV genome), with a 50% inhibitory concentration of 0.1 microM. There was no inhibitory effect on HBV transcription or protein synthesis. In the 2.2.15 cell system, L-FMAU did not show any toxicity up to 200 microM, whereas the D-enantiomer was toxic, with a 50% inhibitory concentration of 50 microM. Repeated treatments of HepG2 cells with L-FMAU at a 1 microM concentration for 9 days did not result in any decrease in the total mitochondrial DNA content, suggesting that a mode of toxicity similar to that produced by 2',3'-dideoxycytidine is unlikely. Also at concentrations as high as 200 microM, L-FMAU did not adversely affect mitochondrial function as determined by lactic acid production by L-FMAU-treated hepatoma cells. L-FMAU was metabolized in the cells to its mono-, di-, and triphosphates, A dose-dependent inhibition of HBV DNA synthesis by L-FMAU triphosphate was observed in the DNA polymerase assays with isolated HBV particles, suggesting that the mode of action of this compound could involve viral polymerase. However, L-FMAU was not incorporated into the cellular DNA. Considering the potent inhibition of the viral DNA synthesis and the nontoxicity of L-FMAU towards the host DNA synthetic machinery, this compound should be further explored for development as asn anti-HBV drug.

Antiviral Agents↗
PubMed1996

2',3'-dideoxy-beta-L-5-fluorocytidine inhibits duck hepatitis B virus reverse transcription and suppresses viral DNA synthesis in hepatocytes, both in vitro and in vivo.

beta-L-Nucleoside analogs represent a new class of potent antiviral agents with low cytotoxicity which provide new hope in the therapy of chronic hepatitis B virus (HBV) infections. We evaluated the anti-HBV activity of 2',3'-dideoxy-beta-L-5-fluorocytidine (beta-L-F-ddC), a beta-L-nucleoside analog derived from 2',3'-dideoxycytidine (ddC), in the duck HBV (DHBV) model. This compound was previously shown to inhibit HBV DNA synthesis in a stably transfected hepatoma cell line (F2215). Using a cell-free system for the expression of an enzymatically active DHBV polymerase, we could demonstrate that the triphosphate form of beta-L-F-ddC does inhibit hepadnavirus reverse transcription. In primary duck hepatocyte culture, beta-L-F-ddC showed a potent inhibitory effect on DHBV DNA synthesis which was concentration dependent. Although beta-L-F-ddC was shown to be less active than ddC against the DHBV reverse transcriptase in vitro, beta-L-F-ddC was a stronger inhibitor in hepatocytes. The oral administration of beta-L-F-ddC in experimentally infected ducklings showed that beta-L-F-ddC is a potent inhibitor of viral replication in vivo. Short-term therapy could not prevent a rebound of viral replication after the drug was withdrawn. Preventive therapy with beta-L-F-ddC could delay the onset of viremia by only 1 day compared with the time to the onset of viremia in the control group. The in vivo inhibitory effect of beta-L-F-ddC was much stronger than that of ddC and was not associated with signs of toxicity. Our data show that beta-L-F-ddC inhibits hepadnavirus reverse transcription and is a strong inhibitor of viral replication both in vitro and in vivo.

Animals↗