Human KB Cells Research Articles

Feroniellides A and B, apotirucallane triterpenes with novel cyclic acetals from Feroniella lucida

The isolation and structure elucidation of two new triterpenes named feroniellides A ( 1) and B ( 2) from Feroniella lucida are described. Feroniellide A has a novel dioxabicyclic [3.2.1]octane moiety, and feroniellide B is the C-3 epimer of the known triterpenoid 3. Their overall structures and relative configurations were established by combined spectral data analysis. The cytotoxicity of 1 and 2 was also evaluated against human KB and HeLa carcinoma cells.

Folate-Modified Cholesterol-Bearing Pullulan, a New Cancer-Targeted Nanoparticle Drug Carrier: Synthesis and Applications

Folate-modified cholesterol-bearing pullulan (FA-CHP) was synthesized by the reaction of folic acid [.beta]-2-aminoethylamide and 4-nitrophenyl chloroformate-activated cholesterol-bearing pullulan. The substitution ratio was estimated as 10% per glucoside units. Several combinations of FA-CHP, unmodified cholesterol-bearing pullulan (CHP) and doxorubicin (DOX) mixtures were tested for cancer-selective cytotoxicity in vitro. A combination of FA-CHP/CHP/DOX of 1:4:0.02 (weight ratio) had aggressive and selective effects on human epidermoid cancer KB cells, known to express a high level of folate receptor, while the same nanoparticle mixture had a weak effect on the growth of normal human fetal lung fibroblast TIG-1-20 cells.

C35 Terpenoids from the Bark of Calocedrus macrolepis var. formosana with Activity against Human Cancer Cell Lines

Ferrugicadinol (1) and ferrugieudesmol (2), two new compounds with a unique C35 terpene skeleton, were isolated from the bark of Calocedrus macrolepis var. formosana. Their structures were elucidated principally from spectroscopic evidence. The two new C35 terpenes showed cytotoxicity against human oral epidermoid carcinoma KB cells with IC50 values ranging from 11 to 14 microM.

Lgf-YL-9 induces apoptosis in human epidermoid carcinoma KB cells and multidrug resistant KBv200 cells via reactive oxygen species-independent mitochondrial pathway

Pyrazolon derivatives were reported to have cytotoxicity to some tumour cells. In the present study, we investigated the effect of Lgf-YL-9 on cytotoxicity and cell apoptosis in human epidermoid carcinoma drug-sensitive parental KB cells and multidrug resistant (MDR) KBv200 cells. Lgf-YL-9 exhibited potent cytotoxicity not only to KB cells but also to KBv200 cells, and the IC 50 were 3.81 and 3.45 μg/mL in KB cells and KBv200 cells, respectively. Importantly, Lgf-YL-9 effectively inhibited tumour growth of KB cell xenografts in nude mice. Lgf-YL-9-induced cell apoptosis was confirmed by chromatin condensation, DNA fragmentation, Annexin-V and propidium iodide (PI) double-staining assay and poly(ADP–ribose) polymerase (PARP) cleavage. Furthermore, Lgf-YL-9-mediated apoptosis in KB cells and KBv200 cells was accompanied by the loss of mitochondrial membrane potential (Δ Ψ m), the release of cytochrome c, and the activation of caspases-3, -7, and -9, but not by intercalating to DNA. Although Lgf-YL-9-induced apoptosis was associated with the decrease of Δ Ψ m, reactive oxygen species (ROS) reduction was interestingly observed in both cell lines. The data suggest that Lgf-YL-9 has similar cytotoxicity to drug-sensitive parental KB cells and MDR KBv200 cells. Lgf-YL-9-induced apoptosis is involved in a new ROS-independent mitochondrial dysfunction pathway, but not in intercalating to DNA.

Synthesis and Characterization of Phosphorylcholine-Substituted Chitosans Soluble in Physiological pH Conditions

A polymer analogous synthesis involving the reductive amination of phosphorylcholine (PC)-glyceraldehyde with primary amines of deacetylated chitosan (M(w) approximately 57000 g mol(-1)) was used to prepare phosphorylcholine-substituted chitosans (PC-CH) with a degree of substitution (DS) ranging from approximately 11 to approximately 53 mol % PC-substituted glucosamine residues. The PC-CH derivatives were characterized by (1)H NMR spectroscopy, FTIR spectroscopy, and multiangle laser light scattering gel permeation chromatography (MALLS-GPC). The pK(a) of the PC-substituted amine groups (pK(a) approximately 7.20) was determined by (1)H NMR titration. The PC-CH samples (1.0 g L(-1)) were shown to be nontoxic using an MTT assay performed with human KB cells. Aqueous solutions of PC-CH samples (4.0 g L(-1)) of DS >or= 22 mol % PC-substituted glucosamine residues remained clear, independently of pH (4.0 < pH < 11.0). The remarkable water solubility and nontoxicity displayed by the new PC-CH samples open up new opportunities in the design of chitosan-based biomaterials and nanoparticles.

Synthesis and activity of a folate peptide camptothecin prodrug

A folate receptor targeted camptothecin prodrug was synthesized using a hydrophilic peptide spacer linked to folate via a releasable disulfide carbonate linker. The conjugate was found to possess high affinity for folate receptor-expressing cells and inhibited cell proliferation in human KB cells with an IC 50 of 10 nM. Activity of the prodrug was completely blocked by excess folic acid, demonstrating receptor-mediated uptake.

Neoplaether, a new cytotoxic and antifungal endophyte metabolite fromNeoplaconema napellumIFB-E016

A new diphenyl ether, named neoplaether, together with five known compounds monomethylsulochrin, physcion, helvolic acid, ergosterol and ergosterol peroxide, was isolated from the culture of Neoplaconema napellum IFB-E016, an endophytic fungus residing in the healthy leaves of Hopea hainanensis. The structure of neoplaether was elucidated by a correlative interpretation of its infrared spectrometry, mass spectroscopy and nuclear magnetic resonance spectra, and confirmed by its single-crystal X-ray diffraction analysis. Neoplaether exhibited significant cytotoxic activity against the human nasopharyngeal epidermoid tumor KB cell line, with an IC(50) value of 13.0 microg mL(-1), comparable to that of 5-fluorouracil (2.5 microg mL(-1)) co-assayed as a positive reference. In addition, it showed antifungal activity against Candida albicans, with a minimal inhibitory concentration value of 6.2 microg mL(-1) (amphotericin as a positive control had a value of 1.5 microg mL(-1)).

Effects of VIP and VIP–DAP on Proliferation and Lipid Peroxidation Metabolism in Human KB Cells

In the present study, we have utilized the transglutaminase (TGase) enzyme to modify the primary structure of VIP with diaminopropane (DAP) at the level of the Gln16. We have investigated the conformational stability of VIP and VIP-DAP in solution by limited proteolysis experiments. The VIP-DAP appears to be more resistant to the proteolytic attack of trypsin, thus indicating that the derivatization in position 16 is able to stabilize the structure of the peptide. However, we have studied their role in cell cycle modulation and antioxidant activity in the oropharyngeal epidermoid carcinoma KB cells.

A dual-ligand approach for enhancing targeting selectivity of therapeutic nanocarriers

Conjugation of ligands to nano-scale drug carriers targeting over-expressed cell surface receptors is a promising approach for delivery of therapeutic agents to tumor cells. However, most commonly utilized ligands are directed at receptors expressed not only on target cells but also on other cells in the body, leading to unintended uptake in these off-target cells. In this study, a novel, dual-ligand approach is reported, which targets tumor cells while sparing off-target cells by exploiting the fact that tumor cells typically over-express multiple types of surface receptors. This approach was tested in the human KB cell line, which over-expresses both folate receptor (FR) and the epidermal growth factor receptor (EGFR). Liposomal nanocarriers loaded with doxorubicin and bearing controlled numbers of both folic acid and a monoclonal antibody against the EGFR were designed. Cytotoxicity was used to determine targeting selectivity of the designed carriers in vitro by utilizing KB cells expressing both FR and EGFR and off-target control cells in which one or both receptors were blocked. The data demonstrates that nanocarriers can be designed to achieve toxicity only when all targeted receptors are available, providing an approach to improve selectivity over current single-ligand approaches.

Application of multicomponent reactions to antimalarial drug discovery. Part 2: New antiplasmodial and antitrypanosomal 4-aminoquinoline γ- and δ-lactams via a ‘catch and release’ protocol

A parallel synthesis of a new series of 4-aminoquinoline γ- and δ-lactams synthesized via the Ugi 3-component 4-centre multicomponent reaction is described. The basicity of the quinoline nitrogen was exploited in the purification of compounds via a ‘catch and release’ protocol. Yields ranging from 60% to 77% and purities as high as 96% were obtained. Compound 29, the most active against a chloroquine-resistant W2 strain of Plasmodium falciparum with an IC 50 of 0.096 μM, also inhibited recombinant falcipain-2 in vitro (IC 50 = 17.6 μM). Compound 17 inhibited the growth of Trypanosoma brucei with an ED 50 of 1.44 μM whilst exhibiting a favourable therapeutic index of 409 against a human KB cell line.

Synthesis and in Vitro/in Vivo Evaluation of Novel 99mTc(CO)3-Folates

Novel organometallic 99mTc(I)-folate derivatives have been synthesized and evaluated in vitro and in vivo in order to assess the influence of the overall charge of the radioconjugates and the spacer entity on the affinity and pharmacokinetic profile. Folic acid has been functionalized at the gamma-carboxylate group of the glutamate moiety with (i) a hydrophilic diethoxyethyl spacer bearing a picolylamine monoacetic acid chelate, (ii) a hexyl spacer bearing an iminodiacetic acid chelate, and (iii) a hexyl spacer with a bis(pyridylmethyl)amine chelating system. Coordination of the 99mTc(CO)3-core resulted in neutral complex 21, anionic complex 22, and cationic complex 23 in excellent yields (>90%) at ligand concentrations of 10(-4) M. Complexes 21-23 were HPLC purified for in vitro and in vivo experiments. In the case of 23, separation from the unlabeled folate analogue was incomplete, leading to low specific activity and, hence, significantly inferior in vivo uptake in folate-receptor-positive (FR-positive) organs and tissues (tumors and kidneys). Time dependent in vivo studies were performed in female, athymic nude mice bearing subcutaneous FR-positive human KB cell xenografts at 1, 4, and 24 h post injection (p.i.) of the radiotracers. Tumor uptake ranged between 1.9-2.7% ID/g, 4 h p.i. and 1.6-2.2% ID/g, 24 h p.i. for 21 and 22, and 0.9% ID/g, 4 h p.i. and 1.1% ID/g, 24 h p.i. for 23. Blood clearance was fast for all derivatives (< or =0.2% ID/g 1 h p.i.). Significant fractions of radioactivity were found in nontargeted and FR-negative organs and tissues (particularly in the liver and the intestines/intestinal contents) at early time points p.i. Coadministration of folic acid reduced radioactivity in FR-positive tissues and organs to background levels. In conclusion, overall charge and the nature of the spacer entity seemed to have a relatively minor influence on receptor affinity and the in vivo pharmacokinetic profile of the tested radiofolates.

Feroniellins A–C, novel cytotoxic furanocoumarins with highly oxygenated C 10 moieties from Feroniella lucida

Three new furanocoumarins, named feroniellins A ( 1), B ( 2), and C ( 3), were isolated from the roots of Feroniella lucida. Compounds 1– 3 are novel structures having an oxolane, oxane, and oxepane moiety. Their overall structures and configurations were determined by spectral and chemical methods. The cytotoxicities of 1– 3 were evaluated against human KB and HeLa carcinoma cells.

Synthesis of Variously Substituted 3-Phenoxymethyl Quinoxalin-2-Ones and Quinoxalines Capable to Potentiate In Vitro the Antiproliferative Activity of Anticancer Drugs in Multi-Drug Resistant Cell Lines

Two series of 1,6-dimethyl-3-phenoxymethylquinoxalin-2-ones and 1-benzyl-3-phenoxymethyl-7-trifluoromethylquinoxalin-2-ones, and a series of 2-benzyloxy-3-phenoxymethyl-7-trifluoromethylquinoxaline were synthesized. Their capability to restore/potentiate the antiproliferative activity of clinically useful drugs, such as doxorubicin (Doxo), vincristine (VCR) and etoposide (VP16), in drug-resistant human nasopharyngeal carcinoma KB cells (KB(WT), KB(MDR), KB(7D)and KB(V20C)) was evaluated. In vitro data show that many quinoxalin-2-ones and quinoxalines potentiate the antiproliferative activity of Doxo and VCR in tumor-derived MDR cell lines. In this series, 17a turned out to be the most potent quinoxaline derivative in potentiating the antiproliferative activity of doxorubicin and vincristine against KB(MDR) and KB(V20C) resistant cell lines, respectively.

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 μg/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.

Inhibition of growth and increase of alkaline phosphatase activity in cultured human oral cancer cells by all- trans retinoic acid

In this study, the effects of all- trans retinoic acid (ATRA) on human oral cancer cells with regard to cell growth, the cell cycle, and alkaline phosphatase (ALP) activity were evaluated. Human oral cancer KB cells were treated with various concentrations of ATRA, and cell growth was then determined using the MTT viability assay. The cell-cycle distribution and ALP activity were analysed using a flow cytometer and chemical analyser, respectively. The KB cells were inhibited by ATRA at concentrations of 1–16 μM (1 μM, P < 0.05; 2 μM, P < 0.01; 4, 8 and 16 μM, P < 0.001) in a dose-dependent manner. ATRA arrested KB cells in the G0/G1 phase. The ALP activity in KB cells was increased by ATRA. This is one of the first studies to focus on the expression of ALP in human head-and-neck carcinoma cells treated with retinoids. These findings suggest that the anti-tumour effects of ATRA on human oral cancer are associated with G0/G1 phase arrest and an increase in ALP activity.

Effect of fibrinogen on the adherence of Porphyromonas gingivalis to human oral epithelial cells

To study the role of fibrinogen molecule in the pathogenesis of periodontal diseases. An in vitro cell culture model was used. Methyl-(3)H Thymidine radiolabeled Porphyromonas gingivalis (Pg) ATCC 33277 were examined for their ability to adhere to and invade the confluent monolayers of human oral epithelial KB cells with or without exogenous human fibrinogens by scintillation spectrometry. The addition of exogenous fibrinogens made more amount of and higher ratios of adhesive and invasive Pg, in contrast to the group without exogenous fibrinogen (P < 0.001). At different concentrations of exogenous fibrinogen, the amount and ratios of adhesive and invasive Pg varied significantly (P < or = 0.007). The higher concentrations of exogenous fibrinogen was added, the greater amount and ratios of adhesive and invasive Pg were found. Fibrinogen promotes the adherence of Pg to human oral epithelial cells and may play an important role in the pathogenesis of periodontal diseases.

Chaetoglobosin U, a Cytochalasan Alkaloid from Endophytic Chaetomium globosum IFB-E019

A new cytotoxic cytochalasan-based alkaloid named chaetoglobosin U (1), along with four known analogues, chaetoglobosins C (2), F (3), and E (5) and penochalasin A (4), has been characterized from the EtOAc extract of a solid culture of Chaetomium globosum IFB-E019, an endophytic fungus residing inside the stem of healthy Imperata cylindrica. The structure of chaetoglobosin U was determined through correlative analyses of its UV, IR, CD, MS, and 1D ((1)H and(13)C NMR and DEPT) and 2D NMR (COSY, NOESY, HMQC, and HMBC) data. Chaetoglobosin U (1) exhibited cytotoxic activity against the human nasopharyngeal epidermoid tumor KB cell line with an IC(50) value of 16.0 microM, comparable to that (14.0 microM) of 5-fluorouracil co-assayed as a positive reference. The known analogues 2-5 were moderately active to the cell line, with IC(50) values of 34.0, 52.0, 48.0, and 40.0 microM, respectively.

A new and rapid bioassay for the detection of gliotoxin and related epipolythiodioxopiperazines produced by fungi

Gliotoxin is an immunosuppressive cytotoxin produced by numerous environmental or pathogenic fungal species. For this reason, it is one of the mycotoxins which must be systematically searched for in samples for biological control. In this study, a new, rapid and sensitive method for detecting gliotoxin has been developed. This bioassay is based on the induction of morphological changes in cultured cells (human KB cell line) by gliotoxin. Interpretation of the assay can be carried out after 1 h of incubation, either by direct microscopic observation, or with an automated microplate-reader at 630 nm. The limit of detection is 18–20 ng of gliotoxin in the well, depending on the used observation method. A high degree of specificity of the detection is brought about by the ability of the reducing reactant dithiothreitol to inhibit the biological activities of epipolythiodioxopiperazines (ETPs), such as gliotoxin, by reducing their polysulfide bridge. The bioassay allows a rapid primary screening of samples and a semi-quantitative evaluation of the gliotoxin concentration in extracts. The method has been used to study the gliotoxin production by different fungal strains, allowing to highlight 3 strains of Aspergillus fumigatus producing gliotoxin in various extracts.

Involvement of PPARγ in Oxidative Stress-Mediated Prostaglandin E2 Production in SZ95 Human Sebaceous Gland Cells

Peroxisome proliferator-activated receptor gamma (PPARgamma) is thought to play a role in sebaceous gland cell function. We previously demonstrated in human epidermoid carcinoma KB cells that UVB irradiation activates PPARgamma via the generation of multiple oxidized glycerophosphocholine species with PPARgamma ligand activity. UVB-induced cyclooxygenase 2 (COX-2) expression was also shown to be PPARgamma-dependent. We therefore reasoned that PPARgamma activation and PPARgamma-dependent COX-2 expression may occur as a general consequence of oxidative stress. The present studies were designed to examine the effects of the oxidant tert-butylhydroperoxide (TBH) on PPARgamma activation and COX-2 expression in SZ95 sebocytes. We first verified that functional PPARgamma is expressed and activated by UVB irradiation in these cells. We next demonstrated that TBH increased PPARgamma reporter activity in SZ95 sebocytes. Increased COX-2 protein, mRNA expression, and prostaglandin E(2) (PGE(2)) production was observed after TBH or PPARgamma agonist treatment. The ability of PPARgamma agonists and TBH to induce COX-2 expression and PGE(2) production was blocked by pretreatment with the specific PPARgamma antagonist GW9662. Finally, TBH and PPARgamma agonists failed to elicit a PGE(2) response in SZ95 sebocytes stably expressing a dominant-negative PPARgamma. This study illustrates the importance of the PPARgamma system in regulating cellular responses to oxidative stress.

Folate-mediated targeting of polymeric conjugates of gemcitabine

The synthesis of two new macromolecular prodrugs for active tumor targeting was set up. Gemcitabine (2′-deoxy-2′,2′-difluorocytidine) was conjugated to α,β-poly( N-2-hydroxyethyl)- dl-aspartamide (PHEA) through succinyl or diglycolyl hydrolysable spacers. The targeting agent folic acid was attached to the macromolecular backbone through the aminocaproic spacer. The two conjugates [PHEA-(5′-succinylgemcitabine)-1′-carboxypentyl-folamide and PHEA-(5′-diglycolyl-gemcitabine)-1′-carboxypentyl-folamide], were purified and extensively characterised by spectroscopic (UV, IR and NMR) and chromatographic analyses to determine the correct chemical structure, the purity degree and the reaction yield. In vitro studies demonstrated that the drug release depends on the spacer arm (diglycolyil or succinyl) and incubation pH. After 30 h incubation at pH 7.4, mimicking the plasma and extracellular compartments, the gemcitabine release from the succinyl and diglycolyl derivatives was 28 and 31%, respectively. After 30 h incubation at pH 5.5, mimicking the lisosomial compartment, the drug released from both bioconjugates was lower than 13%. In plasma, the polymer conjugation increased the drug stability and provided for a sustained drug release. In vitro citotoxicity studies performed using human nasopharyngeal epidermal carcinoma KB cells demonstrated that PHEA-(5′-succinylgemcitabine)-1′-carboxypentyl-folamide displays an higher dose dependent cytotoxic effect with respect to PHEA-(5′-diglycolyl-gemcitabine)-1′-carboxypentyl-folamide.