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At least 163 records · Page 9Linked to original sources
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↗
PubMed2002

ent-Kaur-16-en-19-oic acid, a KB cells cytotoxic diterpenoid from Elaeoselinum foetidum.

Toxic and cytotoxic activities of the toxic plant Elaeoselinum foetidum (Apiaceae) were evaluated using the brine shrimp toxicity (BST) and KB cell cytotoxicity assays. The active chloroform extract was subjected to a bioactivity-directed fractionation, monitored by the BST assay, that led to the isolation of the diterpenoid ent-kaur-16-en-19-oic acid. This compound was potent against the brine shrimp (LC(50) = 4.8 microg/mL) and KB cells (IC(50) = 1.6 microg/mL).

Animals↗
PubMed1995

Structural and functional analysis of the human KB cell folate receptor gene P4 promoter: cooperation of three clustered Sp1-binding sites with initiator region for basal promoter activity.

The human folate receptors (hFRs) are important in the cellular accumulation of folates and antifolates. We described the structure of the human KB cell FR (hFR-KB) gene and identified two discrete promoter regions (P1 and P4) upstream from exons 1 and 4, respectively (Elwood et al., 1993). To further understand the molecular basis of hFR expression, we have now analyzed the basal transcription of the P4 promoter localized upstream of a major transcription start site. The sequence upstream from exon 4 contains several potential transcriptional factor-binding sites and a consensus initiator region sequence at the transcription start site but does not contain canonical TATA or CAAT boxes. While deletion of a 5' flanking sequence from nt -1023 to nt -605 of P4 promoter region decreases the luciferase reporter gene expression in KB cells to 54-70% of control construct, the removal of the sequence between nt -292 and nt -46 markedly decreases the activity to 3%. DNase I footprints and competitive mobility shift and supershift mobility assays indicate that Sp1 or Sp1-related nuclear protein(s) bind to three clustered GC-rich regions within the sequence between nt -292 and nt -46 of the hFR-KB P4 promoter. Both in vitro and in vivo analyses of the expression of promoter constructs containing site-specific mutation(s) of these three Sp1-binding sites and initiator sequence demonstrate that each of three Sp1 sites and the initiator sequence are required for optimum promoter activity and that they interact cooperatively in this P4 promoter of the hFR-KB gene.

Base Sequence↗
PubMed1994

Multiple pathways for activation of E2A expression in human KB cells by the 243R E1A protein of adenovirus type 5.

Adenovirus type 5 (Ad5) mutant dl520, which produces only the smaller 243 residue (243R) E1A protein, induced efficient production of the viral E2A 72-kDa DNA binding protein (DBP) in human KB cells, but not in human WI38, 143, or HeLa cells. In transient expression assays, the 243R E1A protein induced transcription from the E2 early promoter in KB but not in HeLa cells; there was no transcription from the E3 promoter in either cell line. In KB cells, truncation of the E2 promoter from -285 to -97 basepairs dramatically reduced transactivation by the 243R E1A product but not by wt E1A, suggesting that the 243R protein acts through factors binding in this region. Multiple deletions in both exon 1 and exon 2 of the 243R E1A protein failed to disrupt its ability to induce DBP expression. The possible redundant pathways for this induction are discussed. The multiplicity of these pathways and the fact that they are all inactivated in the WI38 and 143 lines are surprising.

Adenoviridae↗
PubMed2002

Interaction of Porphyromonas gingivalis with KB cells: comparison of different clinical isolates.

The ability of different Porphyromonas gingivalis strains (15 clinical isolates and ATCC 33277) to attach to and invade KB cells, in relation to other properties such as release of interleukin (IL)-6 and IL-8, cytotoxicity, proteolytic activity and types of fimbriae genes present, was examined. A hierarchical cluster analysis based on adherence and internalization resulted in four groups. Eight of the 15 clinical isolates belonged to a cluster group whose adherence and internalization were about 10% those of the ATCC strain. A negative correlation between lysine-specific protease activity and adherence was found. In all cases the released concentrations of IL-6 and IL-8 were very low. Only one strain was found to be cytotoxic to KB cells. Principal components analysis demonstrated correlations between adherence, internalization and autoaggregation. Most strains had fimA type I and II, type I being associated with elastase-like activity. The ability of P. gingivalis to invade epithelial cells may be a key factor for maintaining periodontal disease.

Bacterial Adhesion↗
PubMed1989

5-Fluorouracil inhibits dihydrofolate reductase precursor mRNA processing and/or nuclear mRNA stability in methotrexate-resistant KB cells.

This laboratory previously reported that 5-fluorouracil (FUra) increases dihydrofolate reductase (DHFR) precursor mRNA (pre-mRNA) levels relative to DHFR mRNA levels in a methotrexate-resistant KB cell line; these data suggested that incorporation of FUra into RNA may, in part, lead to cell death through the inhibition of mRNA processing (Will, C. L., and Dolnick, B.J. (1987) J. Biol. Chem. 262, 5433-5436). Utilizing a methotrexate-resistant KB cell line designated 1BT, we now report the kinetic basis for altered levels of DHFR RNA observed in FUra-treated cells. Long-term exposure to FUra had no effect on the steady-state level of DHFR pre-mRNA containing intron V or I. However, steady-state levels of total DHFR mRNA decreased 2.0-fold on a per cell basis in cells exposed to 1.0 microM FUra. No significant change in the half-life of total DHFR mRNA or pre-mRNA was observed in cells exposed to FUra (t1/2 = approximately 11.5 h and 50 min, respectively). Nuclear/cytoplasmic RNA labeling experiments demonstrated that the rate of nuclear DHFR RNA conversion to cytoplasmic DHFR mRNA decreased approximately 1.8-fold in FUra-treated cells. These results provide further evidence the FUra may inhibit processing of mRNA precursors and/or affect the stability of nuclear DHFR mRNA.

Blotting, Northern↗
PubMed2006

The apoptosis effect of hispolon from Phellinus linteus (Berkeley & Curtis) Teng on human epidermoid KB cells.

Phellinus linteus (Berkeley & Curtis) Teng, a well-known fungus of the genus Phellinus in the family of Hymenochaetaceae, is being increasingly used to treat a wide variety of disease processes such as oral ulcer, gastroenteric disorder, inflammation, lymphatic disease, and various cancers. However, the mechanism underlying its anti-oral cancer effect is poorly understood. In the present study, we prepared the ethanol extract of Phellinus linteus as a crude drug, and then obtained the active component hispolon by bioassay-guided isolation. Hispolon showed a dose-dependent inhibition of human epidermoid KB cell proliferation with IC50 of 4.62+/-0.16 microg/ml. Furthermore, it was revealed that hispolon could induce human epidermoid KB cell apoptosis with the characteristic of a DNA ladder, and with a significant increase of sub-G1. This process was accompanied by the collapse of mitochondrial membrane potential, the release of cytochrome c and the activation of Caspase-3. These results demonstrated that hispolon induced the death of KB cells through a mitochondria mediated apoptotic pathway. We propose that Phellinus linteus and its effective components could be used as an anti-oral cancer drug for future studies.

Apoptosis↗
PubMed1985

Isolation and genetic characterization of human KB cell lines resistant to multiple drugs.

Human KB cell lines resistant to high levels of colchicine were isolated by several successive single-step selections. Most of these selection steps resulted in cross-resistance to vincristine, vinblastine, adriamycin, actinomycin D, and puromycin; however, at the highest levels of colchicine resistance, increased cross-resistance to other drugs was not observed. There was no major change in protein synthesis or alteration in protein phosphorylation or [14C]glucosamine labeling patterns accompanying the development of multiple drug resistance as measured by analysis of metabolically labeled proteins on SDS gels. Cell-cell hybridization experiments showed that the colchicine-resistant and multiple drug-resistant phenotypes were incompletely dominant. In addition, colchicine resistance was found to segregate independently from resistance to other drugs in one somatic cell hybrid, suggesting that complex genetic loci are involved in the development of the multiple drug-resistant phenotype. These mutants should be useful for the study of the clinically important problem of multiple drug resistance in human cancer.

Antineoplastic Agents↗
PubMed1972

Characterization of messenger ribonucleoprotein and messenger RNA from KB cells.

Messenger ribonucleoprotein and mRNA from KB-cells were isolated under conditions designed to minimize nonspecific RNA-protein interaction and to minimize degradation by contaminating ribonucleases. A large fraction, 60-70%, of the messenger ribonucleoprotein from polysomes dissociated in vitro by either EDTA or puromycin sedimented faster than the large ribosome subunit. Messenger ribonucleoprotein particles with sedimentation coefficients up to 200 S were observed. Released mRNA was also large, with maximal molecular weights around 5 x 10(6).

Cell Fractionation↗
PubMed1994

Delivery of liposomes into cultured KB cells via folate receptor-mediated endocytosis.

Folic acid was covalently conjugated to 66-nm liposomes via spacers of various lengths in an attempt to target the liposomes to KB cells expressing folate receptors. Spacers of short and intermediate lengths were unable to mediate association of folate-conjugated liposomes with receptor-bearing cells, however, use of a 250 A polyethyleneglycol spacer (PEG, M(r) approximately 3350) permitted avid uptake of the liposomes at approximately 2.5 x 10(5) sites/cell. The binding of folate-PEG liposomes to KB cells could be competitively inhibited by excess free folate or by antiserum against the folate receptor, demonstrating the interaction is mediated by the cell surface folate-binding protein. Following binding, cell-associated folate-PEG liposomes were internalized by folate-receptor-mediated endocytosis at 37 degrees C but not at 4 degrees C. These folate-PEG liposomes show potential for delivering large quantities of low molecular weight compounds nondestructively into folate receptor-bearing cells.

Biological Transport↗
PubMed1998

Enhancement of drug accumulation by andrastin A produced by Penicillium sp. FO-3929 in vincristine-resistant KB cells.

In the course of our screening for compounds that reverse multidrug resistance, we found that the cytotoxicity of vincristine was enhanced 1.5-20-fold depending on the concentration of andrastin A in vincristine-resistant KB cells (VJ-300). Andrastin A alone had no effect on the growth of drug sensitive KB cells and VJ-300 cells. On the other hand, andrastin A (25 and 50 micrograms/ml) significantly enhanced accumulation of [3H]vincristine in VJ-300 cells. Andrastin A (50 micrograms/ml) completely inhibited the binding of [3H]azidopine to the P-glycoprotein in VJ-300 cells. The result suggests that andrastin A directly interacts with P-glycoprotein and inhibits the efflux of antitumor agents in drug resistant cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1994

Alpha-interferon induces depletion of intracellular iron content and upregulation of functional transferrin receptors on human epidermoid cancer KB cells.

We have demonstrated that interferon-alpha (IFN alpha) upregulates the epidermal growth factor receptor (EGF-R) on human epidermoid carcinoma cells. Here we report that IFN alpha induces growth inhibition and upregulation of transferrin receptor (TRF-R) on epidermoid cancer KB cells. IFN alpha does not alter TRF-R affinity for its ligand and induces a two-fold increase of TRF binding sites. IFN alpha does not modify receptor internalization and cycling. Intracellular iron levels are known to regulate TRF-R expression: we have, therefore, evaluated whether changes in the iron content could be determined by IFN alpha. Iron levels are transiently increased after addition of fresh growth medium in untreated controls but not in KB cells exposed for 48 h to IFN alpha. Iron depletion is however completely reversed 24 h later when maximal TRF-R upregulation occurs in IFN alpha-treated cells. We suggest that IFN alpha-induced iron depletion elicits a homeostatic cellular response through upregulation of TRF-R.

Cell Cycle↗
PubMed1976

Characterization of KB cell alkaline phosphatase. Evidence of similarity to placental alkaline phosphatase.

The alkaline phosphatase from KB cells was purified, characterized, and compared to placental alkaline phosphatase, which it resembles immunologically. Two nonidentical nonomeric subunits of the KB phosphatase were found. The two subunits, which have apparent molecular weights of 64,000 and 72,000, can be separated on polyacrylamide gels containing sodium dodecyl sulfate. The Mr = 64,000 KB subunit appears to be identical in protein structure to the monomer of placental alkaline phosphatase. The Mr = 72,000 KB subunit, while differing in the NH2-terminal amino acid, appears also to be very similar to the placental alkaline phosphatase monomer. Both KB phosphatase subunits bind (32P)phosphate, and bind to Sepharose-bound anti-placental alkaline phosphatase. Native KB phosphatase is identical to the placental isozyme in isoelectric point, pH optimum, and inhibition by amino acids, and has a very similar peptide map. The data presented support the hypothesis that the Mr = 64,000 KB phosphatase subunit may the the same gene product as the monomer of placental alkaline phosphatase. This paper strengthens the evidence that the gene for this fetal protein, normally repressed in all cells but placenta, is derepressed in the KB cell line. In addition, this paper presents the first structural evidence that there are two different subunit proteins comprising the placental-like alkaline phosphatase from a human tumor cell line.

Alkaline Phosphatase↗
PubMed1999

Electrochemical treatment of human KB cells in vitro.

Electrochemical treatment (ECT) of cancer is a promising new method by which direct current is delivered into tumor tissue to induce tumor regression. The purpose of this study is to evaluate the effectiveness of ECT on human cancer cells and to investigate the factors that affect ECT. The biological mechanisms of ECT in cancer treatment were also explored. Using human KB cells, ECT was found to delay cell growth by using 0.3 coulombs (C)/ml (1.5 C in 5 ml of culture medium; 3 V, 400 microA for 62.5 min). From the results of a colony-forming assay, it was clearly demonstrated that increasing the ECT dose decreases tumor cell survival. A cytotoxicity study, in which a methylene blue assay was used, determined that, for 2.5 x 10(5) cells in culture, the 1D50 was 0.68 C/ml. For a fixed dose of 0.6 C/ml (3 C in 5 ml), using higher current and shorter treatment time resulted in better cell survival. Time, therefore, is an important factor. When cell concentration was altered, the survival was higher for increased cell concentrations. A thymidine incorporation assay indicated that the amount of [3H]thymidine incorporated into DNA decreased as the ECT dose increased. After treatment with 1 C/ml (5 C in 5 ml; 3 V, 400 microA for 208.4 min), pH at the anode decreased to 4.53 and at the cathode increased to 10.46. These results indicate that ECT is effective for killing human KB cells in vitro and that the toxicity effect is related to charge, current, and treatment time. The effect of pH alteration on cells is one of the mechanisms of ECT.

Cell Division↗
PubMedDate not supplied

In vitro effect of hyperthermia on chemoenhancement and uptake of cisplatin in human pharyngeal carcinoma KB cells.

The purpose of this study was to assess the efficacy of hyperthermia (42 or 44 degrees C) as a modifier of cis-diamminedichloroplatinum (II) (CDDP) cytotoxicity and platinum incorporation in human pharyngeal carcinoma KB cells. To maximize the interactive effect, we examined the time sequence of high (above 43 degrees C) or low (below 43 degrees C) hyperthermia and CDDP. Within the dose range of CDDP studied, there was a marked synergism between the effects of heating at 44 degrees C and subsequent CDDP exposure for 5 h. Pretreatment at 44 degrees C for 30 min or at 42 degrees C for 4 h enhanced CDDP cytotoxicity more than posttreatment at 44 degrees C for 30 min or at 42 degrees C for 4 h. However, the chemoenhancement ratio of pretreatment at 44 degrees C for 30 min was higher then that of pretreatment at 42 degrees C for 4 h, although the thermal isotoxic dose decreased the cell count to 60% under both conditions. There was a significant increase in CDDP uptake after hyperthermia at 44 degrees C. These results indicate that high hyperthermia effectively enhances subsequent CDDP cytotoxicity in human pharyngeal carcinoma KB cells.

Antineoplastic Agents↗
PubMed1985

Effects of 5-fluorouracil on dihydrofolate reductase and dihydrofolate reductase mRNA from methotrexate-resistant KB cells.

Growth of methotrexate-resistant dihydrofolate reductase gene-amplified KB cells in the presence of 5-fluorouracil results in an increase in dihydrofolate reductase mRNA. This increase can be solely attributed to a species of RNA of approximately 3.5 kilobase pairs in size. Although dihydrofolate reductase enzyme activity increases per cell with increasing 5-fluorouracil, there is a decrease of enzyme activity per mg of protein (Dolnick, B. J., and Pink, J. J. (1983) J. Biol. Chem. 258, 13299-13306). The rate of in vivo enzyme synthesis, as assayed by immunoprecipitation and supported by gel electrophoresis, does not decrease and may in fact increase with increasing 5-fluorouracil. Translation of purified dihydrofolate reductase mRNA in vitro shows that the rate of translation is unaffected by 5-fluorouracil incorporation into mRNA. The inhibition of dihydrofolate reductase by a monospecific polyclonal antiserum is reduced with extracts from 5-fluorouracil-treated cells. Inhibition of dihydrofolate reductase by methotrexate is significantly reduced in extracts from 5-fluorouracil-treated cells compared to control extracts. Tight binding of [3H]methotrexate is also different in extracts from 5-fluorouracil-treated cells. This data supports the hypothesis of translational miscoding during protein synthesis as a major mechanism of 5-fluorouracil-mediated cytotoxicity and suggests a new mechanism of 5-fluorouracil-methotrexate antagonism.

Animals↗
PubMed1984

Down-regulation of the epidermal growth factor receptor in KB cells is due to receptor internalization and subsequent degradation in lysosomes.

Using a monoclonal antibody to the human epidermal growth factor (EGF) receptor (EGF-R1), we have followed the metabolism of the receptor and the pathway of its internalization in KB cells after the addition of EGF. Measurement of surface binding of 125I-labeled EGF showed that about 80% of EGF binding activity disappeared from the plasma membrane after a 10-min exposure to EGF at 37 degrees C. Immunoprecipitation of the receptor from [35S]methionine-labeled cell extracts with EGF-R1 showed that EGF caused the receptor to be degraded with a half-life of 40 min. Immunofluorescence using EGF-R1 showed an EGF-dependent redistribution of the EGF receptor. In cells not exposed to EGF, almost all of the receptor was diffusely distributed on the cell surface. After EGF addition, the receptor was rapidly internalized, first appearing in small punctate organelles characteristic of receptosomes and then in larger perinuclear lysosome-like structures. By 120 min almost all of the immunoreactive EGF receptor had disappeared from the cells. Immunocytochemistry at the electron microscopic level confirmed these light microscopic findings. The diffusely distributed receptor on the cell surface first clustered into clathrin-coated pits in the presence of EGF, next was internalized into receptosomes, appeared transiently in transreticular Golgi elements, and finally was seen in lysosomes. This EGF-dependent down-regulation and degradation of the EGF receptor in KB cells provides a striking example of ligand-dependent clustering and internalization of a receptor, followed by degradation in lysosomes of both ligand and receptor.

Antibodies, Monoclonal↗
PubMed1987

Purified membrane and soluble folate binding proteins from cultured KB cells have similar amino acid compositions and molecular weights but differ in fatty acid acylation.

A membrane-associated folate binding protein (FBP) and a soluble FBP, which is released into the culture medium, have been purified from human KB cells using affinity chromatography. By NaDodSO4/PAGE, both proteins have an apparent Mr of approximately 42,000. However, in the presence of Triton X-100, the soluble FBP eluted from a Sephadex G-150 column with an apparent Mr of approximately 40,000 (similar to NaDodSO4/PAGE) but the membrane-associated FBP eluted with an apparent Mr of approximately 160,000, indicating that this species contains a hydrophobic domain that interacts with the detergent micelles. The amino acid compositions of both forms of FBP were similar, especially with respect to the apolar amino acids. In addition, the 18 amino acids at the amino termini of both proteins were identical. The membrane FBP, following delipidation with chloroform/methanol, contained 7.1 mol of fatty acid per mol of protein, of which 4.7 mol was amide-linked and 2.4 mol was ester-linked. The soluble FBP contained only 0.05 mol of fatty acid per mol of protein. These studies indicate that the membrane FBP of KB cells contains covalently bound fatty acids that may serve to anchor the protein in the cell membrane.

Acylation↗