Re: Synergistic anticancer effects of ganciclovir/thymidine kinase and 5-fluorocytosine/cytosine deaminase gene therapies.
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Animals↗
Biomedical subjects
F Y Wu
Publications and source records attributed to F Y Wu.
Vitamin K3 induces cell cycle arrest and cell death by inhibiting Cdc25 phosphatase.
Our early reports have indicated that vitamin K3 (VK3) exerts antitumour activity by inhibiting Cdk1 activity and overexpressing the c-myc gene to induce an apoptotic cell death. In the present study, we investigated the effect of VK3 on Cdc25 phosphatase, a Cdk1 activator and c-Myc-downstream protein. Increased protein level but decreased activity of Cdc25A phosphatase was found in cervical carcinoma SiHa cells treated with VK3 for 1 h and allowed to recover for 8, 24, 30 or 45 h. The binding of VK3 to Cdc25 phosphatase was proven by incubating [methyl-3H]-VK3 with the 27 kDa-catalytic domain of Cdc25A phosphatase at 35 degrees C for 2 h. We found that VK3 inhibited cyclin E expression at late G1 phase and cyclin A at G1/S transition of the aphidicolin-synchronised SiHa cells, but had no effect on Cdk2 and Cdk4. The inhibition of cyclins E and A expression was associated with cell cycle progression delay in the S phase. These results indicate that binding of VK3 to the catalytic domain of Cdc25 phosphatase results in the formation of inactive, hyperphosphorylated Cdk1 that subsequently induces cell cycle arrest, leading to cell death. These findings suggest a possible therapeutic strategy, with VK3 serving as a potential antagonist to tumours expressing high levels of proteins containing cysteine such as oncogenic Cdc25A phosphatase.
Apoptosis↗
Suppression of tumorigenicity in cervical carcinoma HeLa cells by an episomal form of adeno-associated virus.
To compare the oncosuppressive activity of integrated and episomal adeno-associated virus (AAV), two Epstein-Barr virus (EBV) derived episomal form vectors, p205 and p220, were used to generate plasmids containing episomal AAV. HeLa cells transfected with p205, p220, and the plasmids with forward (pA205-1, pA220-1) and reverse (pA205-1, pA220-2) orientation of inserted AAV DNA were designated H205, H220, HA205-1, HA220-1, HA205-1, and Ha220-2, respectively. The respective average copy numbers of pA205 and pA220 per cell are 4.9-6.9 and 4.3. The AAV-transfected HeLa cells displayed growth inhibition when compared to parental and the vector-transfected HeLa cells. Nude mice assay showed that the tumor size from HA205-1 and HA205-2 cells were 11.3 and 22.6%, and those from HA220-1 and HA220-2 cells were 54.8 and 57.3%, respectively, when compared to parental HeLa cells. HA205-1 cells containing forward AAV insert exerted more oncosuppressive effect than HA205-2 cells containing reverse AAV insert. Our data indicate that the episomal AAV can exert oncosuppressive activity as compared to the integrated AAV in HeLa cells.
Animals↗
Alien hand syndrome: report of two cases.
Alien hand syndrome (AHS) refers to the occurrence of apparently purposeful movements in the hand which are independent of volitional control. Two subtypes of AHS have been proposed: frontal AHS, with grasp reflex and compulsive manipulation of tools by the dominant hand, and callosal AHS, which occurs in the nondominant hand and is characterized mainly by intermanual conflict. Here, we report two cases of frontal-type alien hand syndrome with symptoms of reflexive grasping, impulsive groping, and apraxia (in case 1), and compulsive manipulation of tools (in case 2). Brain computed tomography revealed a left anterior cerebral artery (ACA) territory infarct and multiple small infarcts of both hemispheres in patient 1 and a left ACA infarct in patient 2. The involuntary movements were bothersome to these patients in their daily activities. Both patients attended conventional physical and occupational therapies, and patient 2 received additional biofeedback training. Follow-up studies showed the spontaneous grasping behavior was still present in patient 1 and AHS had subsided in patient 2. We also describe a potentially effective technique involving biofeedback for patients with alien hand syndrome.
Aged↗
Menadione (vitamin K3) enhances the mitogenic signal of epidermal growth factor via extracellular signal-regulated kinases.
The effects of vitamin K3 (VK3) on DNA synthesis, cell proliferation and mitogen-activated protein kinase pathway were investigated in G0-arrested NIH 3T3 fibroblasts. VK3 (5 microM) alone stimulates DNA synthesis by 40% and moderately increases the mitogenic effects of EGF, which is preceded by a rapid phosphorylation of the extracellular signal-regulated kinases (ERKs). At 20 microM, VK3 had an antiproliferative effect. VK3 alone (5 and 50 microM) or in concert with EGF increases the activity of ERK2 (by 2.5 and 5 fold, respectively). Our studies demonstrate that the activation of ERKs by VK3 alone, or VK3 plus EGF can promote either stimulatory or inhibitory effects on the mitogenic signal.
3T3 Cells↗
The interaction between human cytomegalovirus immediate-early gene 2 (IE2) protein and heterogeneous ribonucleoprotein A1.
Human cytomegalovirus (HCMV) is an ubiquitous pathogen which causes significant illness in immunocompromised individuals. The immediate-early gene 2 (IE2) protein of HCMV plays an important role in the regulation of virus replication. Previous studies have shown that the IE2 protein is able to interact with several cellular proteins, but many of the IE2 interacting partners remain unidentified. By utilizing the yeast two-hybrid system, the heterogeneous ribonucleoprotein A1 (hnRNP A1) was identified as an IE2 interacting protein. The interaction was confirmed via the in vitro binding of bacterial expressed glutathione S-transferase (GST) IE2 fusion protein with the in vitro translated hnRNP A1. The mutational analysis showed that both the N-terminal half (1-290 residues) and C-terminal half (291-579 residues) of IE2 protein can interact with the hnRNP A1, indicating that more than one region of IE2 protein are involved in the binding with hnRNP A1.
Base Sequence↗
Sister chromatid exchanges in the peripheral lymphocytes of newborns with Down syndrome after in vitro exposure to blue or green light.
This study determines whether irradiation by blue or green light has an adverse effect on the DNA of Down syndrome (DS) cells by examining the sister chromatid exchange (SCE) frequency of peripheral lymphocytes obtained from five neonates with DS and five karyotypically normal neonates (control). Lymphocytes in G0 of the cell cycle were irradiated with blue or green fluorescent light for 1, 2, 4 or 6 h, and then cultured using a conventional method. Our results revealed that the induction of SCEs per cell in both groups increased in a dose-dependent manner, although more SCEs were respectively induced by the blue light. In addition, after 6 h of blue light irradiation, the net-induced SCEs in the DS groups were higher than those in the control groups.
Cells, Cultured↗
A pharmacokinetic study with the high-dose anticancer agent menadione in rabbits.
The aim of this investigation was to assess the pharmacokinetic properties of high-dose menadione (VK3), as an anticancer agent, in plasma and red blood cells (RBCs) in rabbits. An extremely high dose of 75 mg menadiol sodium diphosphate (Synkayvite) was intravenously injected. HPLC analysis was applied to measure the major metabolite, menadione, VK3. The kinetic properties of VK3 in both plasma and red blood cells showed a short elimination half-life, high clearance, and large volume of distribution in plasma and RBCs. The mean elimination t1/2 values of menadione in plasma and in RBCs were 27.17 +/- 10.49 min and 35.22 +/- 11.82 min, respectively. The plasma clearance (CL/F) of VK3 was 0.822 +/- 0.254 L min-1. The systemic clearance in RBCs was 0.407 +/- 0.152 L min-1. The apparent volume of distribution (Vd/F) in plasma was 30.833 +/- 12.835 L and that in RBCs 20.488 +/- 9.401 L. The plasma AUC was 32.453 +/- 9.785 micrograms min mL-1 and that of RBCs 67.219 +/- 24.449 micrograms min mL-1. Menadiol was rapidly biotransformed to menadione in blood. The formation rate constant (kf) of menadione in plasma was 0.589 +/- 0.246 min-1, and that of RBCs 1.520 +/- 1.345 min-1. Through this study the estimated menadione dosage needed to maintain a plasma level of 1 microgram mL-1 for anticancer purposes was 19.7 mg kg-1 every hour.
Animals↗
Differential suppression of the tumorigenicity of HeLa and SiHa cells by adeno-associated virus.
Adeno-associated virus (AAV) is well known for suppression of oncogenesis in rodents, but its inhibitory effects on human carcinoma are less well understood. We report the differential ability of AAV to inhibit the tumorigenicity of two human cervical carcinoma cell lines. The wild-type AAV-2 DNA carried by a pSV2Neo vector was transfected into HeLa cells, which contain 50 copies of human papillomavirus type 18 (HPV-18), and SiHa cells, which contain 1-2 copies of HPV-16. About 1-3 copies of AAV genome were introduced per cell. AAV transfection moderately reduced the growth rate and anchorage-independent activity of the cells. In nude mice, the size of tumours arising from SiHa cells was reduced by 87%, in contrast to no reduction in tumour size arising from HeLa cells. This suggests that the differential suppression exerted by AAV may be due to differences in HPV copy number. To define the region that is responsible for the oncosuppression, mutation analyses were conducted. The results of nude mice assays showed that both the replication gene and inverted terminal repeats of AAV were important for the inhibition. This study may provide a model system for further studies on the underlying mechanism of AAV oncosuppressive activity.
Animals↗
Determination of anticancer drug vitamin K3 in plasma by high-performance liquid chromatography.
Synthetic vitamin K3 (VK3, 2-methyl-1,4-naphthoquinone, or menadione) has been found to exhibit antitumor activity against various human cancer cells at relative high dose. Parallel to our study on the mechanism of VK3 action and for future clinical trials in Taiwan, we developed a simple, sensitive and accurate high-performance liquid chromatographic method for the determination of VK3 in biological fluids. VK3 was extracted from the plasma samples with n-hexane. The chromatographic separation employed an ODS analytical column (5 microns, 250 x 4.6 mm I.D.) with a mobile phase of methanol-water (70:30, v/v) and UV detection at 265 nm. On completely drying of the extraction solution, n-hexane, by a stream of nitrogen, menadione was lost to a great extent. Methanol (70%, 200 microliters) was added to the extraction solvent after extraction and centrifugation to prevent the loss of menadione. The absolute recovery was 82.4 +/- 7.69% (n = 7). The within-day and between-day calibration curves of VK3 in plasma in the ranges of interest (0.01-10.00 micrograms/ml; 0.01-5.00 micrograms/ml) showed good linearity (r > 0.999) and acceptable precision. The limit of quantitation of VK3 was 10 ng/ml in plasma. This method has been successfully applied to a pilot pharmacokinetic study of VK3 in rabbits receiving an intravenous high-dose bolus injection of 75 mg menadiol sodium diphosphate (Synkayvite). The pharmacokinetic properties of menadione could be described adequately by an open two-compartment model. The mean half-life of menadiol (transformation to menadione) was 2.60 +/- 0.12 min.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Cloning and characterization of the endogenous retroviral-tRNA(Glu) multigene family from human genomes of different racial backgrounds.
An 8.3-kb human endogenous retroviral-tRNA(Glu) (HERV-E)-encoding cDNA clone and a 1.5-kb genomic clone were isolated from a Chinese-derived cervical cancer cell line, CC7T, and their sequences determined. The former is a full-length endogenous retroviral cDNA containing corresponding u5-gag-pol-env-u3-r regions. The latter is a partial retroviral DNA segment, covering the gag and pol genes. Analysis of normal human DNA by Southern blot hybridization with three specific HERV-E molecular DNA probes revealed complex restriction-fragment length polymorphisms (RFLP), implying that the human genome contains diverse proviral structures and dispersed integration sites. The complex patterns were virtually identical between DNAs from African-Americans, Asians and Caucasians, with only a few minor variations. The data suggest that these proviral sequences were mostly incorporated into the human genome before racial divergence and, hence, may serve as markers for distinct chromosomal sites.
Amino Acid Sequence↗
Chromosomal translocations in two feline T-cell lymphomas.
Two feline malignant lymphoma cell lines, FT-1 and FT-G, established from cats naturally infected with the feline leukemia virus were analyzed for chromosomal aberrations. Both FT-1 and FT-G cells had a modal number of 38 which is the normal diploid (2n) chromosome number of the domestic cat. G-banding-analysis showed that FT-1 had a translocation involving the short arms of chromosome A2 and D3--t (A2;D3) (p-;p+), and FT-G had a translocation involving the short arms of chromosomes A2 and B2--t (A2;B2) (p-;p+). Our data suggest that the chromosomal translocations were closely associated with the tumorigenesis in malignant lymphoma in cats.
Animals↗
Bovine fast-twitch myosin light chain 1: cloning and mRNA amount in muscle of cattle treated with clenbuterol.
The cDNA clone encoding the fast-twitch isoform of myosin light chain 1 (MLC-1f) was isolated from bovine longissimus dorsi muscle and sequenced in M13 and pUC8. An 0.8-kb subclone, produced by digestion of the cDNA with EcoRI, contained the portion of the molecule common to MLC-1f and MLC-3f. The cDNA in pUC8 contained an additional 81 bp upstream of the EcoR I digestion site, which was unique to MLC-1f. The cDNA clone was used to measure MLC-1f mRNA in longissimus dorsi muscle of cattle chronically administered the beta-adrenergic agonist clenbuterol. Treatment with clenbuterol for 50 days increased succinic dehydrogenase negative (type IIB) and positive (types I and IIA) myofiber cross-sectional areas by 25%. After the 50-day treatment period, the amount of MLC-1f mRNA was 90% greater in longissimus dorsi muscle of treated animals than in the initial group. This effect was lost when clenbuterol treatment was withdrawn for a 78-day period, during which time muscle growth in the treated animals stopped completely. We conclude that we have cloned the bovine cDNA for MLC-1f, which has provided additional evidence that beta-adrenergic agonists increase myofibrillar gene expression.
Amino Acid Sequence↗
Neural network analysis of the P300 event-related potential in multiple sclerosis.
Neural network analysis is sensitive to subtle changes in patterns of data. We hypothesized that a disease process which can cause impairment of cortical function such as multiple sclerosis (MS) would affect the P300 cognitive evoked potential (P300) in a manner detectable by a feedforward backpropagation neural network. Such a network was trained using a learning data set consisting of 101 P300 wave forms (from 26 MS patients and 26 normal controls). The network was then used to classify a randomly selected test data set of 20 studies (2 studies each of 5 MS patients and 5 controls) to which it had not been previously exposed, with an average accuracy (MS = abnormal, control = normal) of 81% for a single midline electrode, increasing to 90% using 3 midline electrodes in a jury system. Neural network analysis can be of help in distinguishing normal (control) P300 from abnormal (MS) P300.
Acoustic Stimulation↗
Neural network approach in multichannel auditory event-related potential analysis.
Even though there are presently no clearly defined criteria for the assessment of P300 event-related potential (ERP) abnormality, it is strongly indicated through statistical analysis that such criteria exist for classifying control subjects and patients with diseases resulting in neuropsychological impairment such as multiple sclerosis (MS). We have demonstrated the feasibility of artificial neural network (ANN) methods in classifying ERP waveforms measured at a single channel (Cz) from control subjects and MS patients. In this paper, we report the results of multichannel ERP analysis and a modified network analysis methodology to enhance automation of the classification rule extraction process. The proposed methodology significantly reduces the work of statistical analysis. It also helps to standardize the criteria of P300 ERP assessment and facilitate the computer-aided analysis on neuropsychological functions.
Algorithms↗
Vitamin K3 inhibits growth of human hepatoma HepG2 cells by decreasing activities of both p34cdc2 kinase and phosphatase.
Synthetic vitamin K3 is known to exhibit in vitro and in vivo antitumor activity against rodent and human cancer cells. To determine vitamin K3's effect on the cell cycle, we treated both synchronized and asynchronized human hepatoma HepG2 cells with 100 microM vitamin K3 for 1 h and followed cells to recover for various times, then monitored cell cycle progression by flow cytometry and by phosphorylation status of p34cdc2 kinase. Vitamin K3 induced a delay at the S/G2 phase of the cell cycle. This was accompanied by constant hyperphosphorylation status and decreased activity of p34cdc2 kinase. Protein-tyrosine phosphatase activity also decreased 2- to 3-fold after vitamin K3 treatment. Our results suggest that vitamin K3 inhibits the growth of HepG2 cells via a cell cycle progression delay and altered phosphorylation patterns and activities of both p34cdc2 kinase and protein-tyrosine phosphatase.
CDC2 Protein Kinase↗
A neural network design for event-related potential diagnosis.
Many diseases resulting in neuropsychological impairment show abnormalities by EEG (electroencephalogram) tests. However, EEG analysis is complicated by a wide spectrum of normal patterns. Cerebral evoked potentials are stimulus-induced, averaged EEG potentials that have been found useful in patients with dementing illness. A recent report using the P300 auditory evoked potential shows that simple latency and waveform criteria result in classification accuracy of 65% for multiple sclerosis (MS) patients versus 91% for control subjects. Analysis on the same data set was performed using an artificial neural network (ANN) and a nearest neighbor (NN) classifier. An ANN classifier demonstrated a classification accuracy of 75% versus 87% on MS and control subject groups. Thus prediction accuracy was improved on average, compared with that obtained by NN classifiers or P300 statistical analysis. The classification strategy discovered by a trained ANN was analyzed by a weight pattern analysis method and compared with the P300 latency criteria.
Diagnosis, Computer-Assisted↗
Regional cerebral blood flow estimation by neural network-based parametric regression analysis.
An artificial neural network (ANN) model was proposed for real-time estimation of regional cerebral blood flow (rCBF), by given head and expired air curves obtained through 133Xe inhalation. The network was constructed according to a regression model described by a linear differential equation. Experimental results compare well with those obtained by conventional curve fitting strategies, but the parameter estimation process is much simplified. A systematic procedure in developing ANN for parametric regression analysis was introduced; networks are constructed according to the selected regression model so that the obtained weights of a trained network directly represent parameters of the regression model which best fits the observed data set. Such a design-oriented methodology extends the classification-based applications of ANN to parametric regression analysis, and therefore may have more generalized applications besides rCBF estimation.
Algorithms↗