Concentrations Of Taxol Research Articles

Bioactive coatings for coronary stents: Modulation of cell proliferation by controlled release of anti-proliferative drugs

Drug eluting coronary stents coated with a bioactive and biostable polymer system (THBA70), based on acrylate copolymers bearing a salicylic acid derivative as side substituent, were evaluated. The microstructural architecture of the copolymer THBA70 comprises a random copolymer of a methacrylate derivative from triflusal (4-trifluoromethyl salicylic acid) with 45 mol% of THEMA (2-methacryloyloxyethyl [2-(acetyloxy)-4-(trifluoromethyl)] benzoate), which makes the system anti-thrombogenic with good adhesion to the surface of metallic stents. The bioactive coating prevented thrombosis, an adverse effect associated with the implantation of drug-eluting stents. The in vitro drug delivery of drug-eluting stents under dynamic conditions indicated excellent controlled drug release. The THBA70 films loaded with relatively low concentrations of taxol or simvastatin in contact with fibroblasts produced inhibition of cell proliferation with a dose-dependent bioactivity.

Optimization of the basal medium for improving production and secretion of taxanes from suspension cell culture of Taxus baccata L

Background and purpose of the studyTaxol is one of the most effective anticancer drugs that isolated from Taxus sp. due to the slow growth of Taxus trees and low concentration of Taxol in the tissues, the biotechnological approaches especially plant cell culture have been considered to produce Taxol in commercial scale.MethodsWe investigated the effects of basal medium type used in culture media on production of Taxol and other taxane compounds from cell suspension culture of T. baccata L. Briefly, five commonly basal media including Gamborg, Murashige and Skoog, Woody Plant, Schenk and Hildebrandt, and Driver and Kuniyuki medium were used for preparing separate suspension culture media. The intra- and extra-cellular yields of taxanes were analyzed by using HPLC after 21 days period of culturing.ResultsThe yields of taxanes were significantly different for the cultures prepared by different basal media. Moreover, the effects of basal medium on the yield of products differed for varius taxane compounds. Maximum yields of Baccatin III (10.03 mgl-1) and 10-deacetyl baccatin III (4.2 mgl-1) were achieved from the DKW basal media, but the yield of Taxol was maximum (16.58 mgl-1) in the WPM basal media. Furthermore, the secretion of taxanes from the cells into medium was also considerably affected by the type of basal medium. The maximum extra-cellular yield of Taxol (7.81 mgl-1), Baccatin III (5.0 mgl-1), and 10-deacetyl baccatin III (1.45 mgl-1) were also obtained by using DKW basal medium that were significantly higher than those obtained from other culture media.

The inhibition of tumor growth and metastasis by self-assembled nanofibers of taxol

Molecular hydrogels have big potential for local delivery and sustained release of therapeutic agents. In this paper, we reported on a molecular hydrogel mainly formed by the widely used anti-cancer drug of taxol. The hydrogel was formed by an ester bond hydrolysis process from a taxol derivative (Taxol-SA-GSSG, 1) and could be administrated into solid tumors to dramatically hinder their growths and prevent their metastasis. Besides the improved anti-cancer effect compared to the clinically used intravenous (i.v.) injection of Taxol®, the concentration of taxol in blood was low due to the local administration of taxol hydrogels, which greatly enhanced the dosage tolerance of mice to taxol and might reduce side effects of taxol during chemotherapy. Our observations suggested that the hydrogel mainly composed of taxol would have great potential for its practical applications.

Abstract LB-84: The role of survivin and survivin variants in the mitotic arrest-induced cell death

Abstract Eukaryotic cells rely on a mitotic checkpoint in order to preserve their proper ploidy. This mechanism stops mitosis progression before anaphase until each kinetochore is adequately bound to mitotic microtubules, and tension is generated which splits up chromatids towards opposite poles of the mitotic spindle. The resulting paused state of the cell cycle is known as mitotic arrest, and has been long known to induce cell death. However, the mechanisms driving mitotic arrest-mediated cell death have not yet been elucidated. It has been hypothesized that an anti-apoptotic/pro-apoptotic protein interplay might define the mitotic arrest outcome. We are interested in defining the role of survivin and its splicing variants, survivin Δex3 and survivin 2b, in the mitotic arrest-induced cell death. Survivin is a multifaceted member of the IAP family of proteins with functions in the chromosome passenger complex that has been shown to be over expressed in nearly every human tumor type. Noteworthy, Survivin Δex3 retains IAP activity but Survivin 2b has been shown to act as a pro-apoptotic factor. In this study, we defined the mitotic index (MI) of HCT116 cells in response to different concentrations of taxol and vinblastine. The results show that vinblastine induces a wide MI oscillation in contrast to taxol, which produces a homogeneous behavior in the cell population. This difference in population behavior might be explained by the different mechanisms of action of taxol and vinblastine. From the maximum taxol-induced MI point on, the percentage of sub-2n cells increased at the expense of mitotic cells, as assessed by flow cytometry. These results suggest that, in our model, cells die by exposure to taxol in a mitotic arrest-dependent manner. We also determined, by RT-PCR, that survivin variants are endogenously expressed in HCT116 cells. Furthermore, in immunoblot experiments, we found that survivin protein level decreases throughout taxol treatment in contrast to Bub1 levels, a member of the spindle assembly checkpoint. These results suggest that the decreased survivin level might be related to mitotic arrest-dependent cell death. The analysis of the intracellular localization of survivin variants throughout the mitotic arrest showed that survivin is localized in mitotic chromosomes throughout mitosis (before anaphase) independently of the time of treatment with taxol and vinblastine. Noteworthy, we also found that survivin Δex3 localized to the cytosol, and its level decreases throughout mitosis while survivin 2b protein was diffusely localized in the cytosol and mitotic chromosomes, and its level and localization remained unchanged. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-84. doi:1538-7445.AM2012-LB-84

Abstract 5677: A comparative study of the interaction of the maytansinoid, DM1-Me, vinblastine, and taxol with human cells

Abstract The current interest in antimitotic compounds that interact with microtubules is caused, in part, by their clinical importance in oncology. Several microtubule-binding compounds, including Vinca alkaloids, taxanes, maytansinoids, and auristatins, are in clinical use, some additionally being evaluated as antibody conjugates. To better understand the factors that determine the potency of antibody-maytansinoid conjugates in killing cells, we examined the interaction of the maytansinoid, DM1-Me, with MCF7 cells, in comparison with vinblastine and taxol. DM1-Me was more potent in arresting mitosis than vinblastine or taxol (5- and 13-fold, respectively). We compared the levels of accumulation of these three compounds by MCF7 cells following a 24 hour exposure at the mitotic IC50 concentration and found that, at this concentration, cells accumulated comparable amounts of vinblastine and DM1-Me, but required more than a 10-fold greater concentration of taxol to achieve a similar effect. Additionally, cells exposed to each compound for 2 hours prior to being washed into drug free medium, retained most of the accumulated DM1-Me or taxol for at least 24 h, while most vinblastine was released into the medium within the first 2 h. DM1-Me and taxol had similar affinities for MCF7 cells, while that of vinblastine was 4-6-fold lower. These properties of DM1-Me, vinblastine and taxol suggest that maytansinoids are a better choice for antibody conjugates than vinblastine or taxol, a conclusion which is consistent with the experimental data and clinical experience with antibody conjugates made with compounds of these three classes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5677. doi:1538-7445.AM2012-5677

Abstract 1492: Eavesdropping of crosstalk between adipose stem cells (ASC) and ovarian cancer cells

Abstract Background Adipose tissue contains a population of multipotent adipose stromal cells (ASC) which migrate into tumors and form supportive tumor stroma elements. ASCs release various growth factors, cytokines and metabolites that influence on physiochemical nature of the local environment. Ovarian cancer cells have a predilection for metastasis in omentum, the primary repository for visceral adipose. The effects of omental derived adipose stem cells (O-ASC) and subcutaneous tissue derived adipose stem cells (SC-ASC) on ovarian cancer biology including ovarian cancer proliferation, gene expression and response to chemotherapy were investigated. Methods and Results Luciferase labelled ovarian carcinoma cells (OVCA 429) were co-cultured with O-ASC, SC-ASC and control fibroblast line Wi38. Significantly higher levels of luciferase activity was detected when ovarian carcinoma cells were co-cultured with O-ASC than with SC-ASC and Wi38 (3470.4 RLU vs 1253.2 RLU, p=<0.005). Moreover, co-culture of 429 cells with ASCs in presence of different concentrations of Taxol revealed that O-ASC influence tumor cell chemosensitivity. Additionally, O-ASC promoted formation of significantly more ovarian carcinoma tumorispheres which were more closer associated to each other than those created in sphere medium only. To identify differentially expressed genes in O-ASC and SC-ASC, we performed microarray experiments (NimbleGen platform) which were analyzed using Ingenuity Pathway Analysis (IPA) software. The highest expressed genes were connected with growth, proliferation and development of cells and most of the proteins coded by these genes were localized in the extracellular space, plasma membrane or exosomes (CALR, UNC5B in O-ASC or COL1A2, FN1 in SC-ASC). We also performed ELISA assays focused on release of growth factors which revealed that O-ASC and SC-ASC secrete more VEGF (1678 pg/ml) and FGF (29 pg/ml) compared to control cells Wi38 (114 pg/ml and 2.7 pg/ml respectively). Chemokine SDF-1α was also secreted in high level by O-ASC S1 (1537 pg/ml) but SC-ASC released only 187 pg/ml relative to Wi38 (706 pg/ml) which indicates that these two types of adipose stem cells can create different surrounding environment. Summary O-ASC cells have unique tumor promoting effects and create specific environmental niche by releasing growth factors, cytokines and other factors which can stimulate ovarian cancer cells to grow faster and clump together. ASC may also influence protection of ovarian cancer from toxicity of chemotherapeutics. The knowledge about microenvironmental crosstalk between normal stem cells and tumor cells can contribute in the development of new treatment to prevent metastasis of ovarian cancer and make them vulnerable to chemotherapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1492. doi:1538-7445.AM2012-1492

Overexpression of Tyro3 receptor tyrosine kinase leads to the acquisition of taxol resistance in ovarian cancer cells.

The majority of patients with ovarian cancer are diagnosed at the advanced stages (III/IV) and their 5-year-survival rate is relatively low. One of the major causes of the poor prognosis of ovarian cancer is the development of resistance to first-line chemotherapy, including platinum and taxol. Therefore, improvements in current understanding of chemoresistance is required for the successful treatment of ovarian cancer. In the present study, taxol-resistant ovarian cancer cells, SKOV3/TR, were established by exposing parental SKOV3 cells to increasing concentrations of taxol. . Briefly, cells were treated with 1.5 nM (for 4 weeks), 3 nM (for 4 weeks), 6 nM (for 5 weeks), 12 nM (for 5 weeks) and 24 nM taxol (for 8 weeks) over 6 months. The SKOV3/TR cells were found to be smaller in size and rounder in shape compared with their parental cells. Cell viability and colony formation assays demonstrated an increase in the population doubling time of the SKOV3/TR cells, indicating a reduction in the proliferative capacity of these cells. Reverse transcription-polymerase chain reaction and western blot analysis revealed that, among the TAM receptor tyrosine kinases (RTKs), the mRNA and protein expression levels of Tyro3 RTK were increased, while those of Axl and Mer RTK were decreased in the SKOV3/TR cells. In addition, restoration of the level of Tyro3 by transfecting Tyro3-specific small interfering RNA into the SKOV3/TR cells reduced the proliferative capacity of the cells, indicating that upregulation of the expression of Tyro3 in SKOV3/TR cells may promote survival in the presence of taxol, which eventually resulted in the acquisition of resistance upon taxol treatment. The present study subsequently found that, in the SKOV3/TR cells, the level of intracellular reactive oxygen species (ROS) was elevated, and antioxidant treatment with N-acetyl cysteine (NAC) exerted more profound antiproliferative effects compared with the parental cells. The western blot analysis demonstrated that treatment of the SKOV3/TR cells with NAC reduced the protein expression of Tyro3, and the inhibitory effect of NAC on the phosphorylation of Akt was increased, which may have had a positive effect on the proliferation of the SKOV3/TR cells. The levels of phosphorylation and protein expression of signal transducers and activators of transcription 3 (STAT3) were not affected by NAC treatment, indicating that the phosphorylation of Akt, but not expression or phosphorylation of STAT3, was associated with the increased intracellular ROS level in the SKOV3/TR cells. Taken together, the results of the present study demonstrated that the acquired taxol resistance of ovarian cancer cells was associated with ROS-dependent upregulation in the expression of Tyro3 RTK and the subsequent activation of Akt.

Intratumoral injection of taxol in vivo suppresses A549 tumor showing cytoplasmic vacuolization

Based on our recent in vitro studies, this report was designed to explore the mechanism by which high concentration of taxol (70 µM) induced paraptosis-like cell death in human lung carcinoma (A549) cells, and to evaluate the therapeutic efficacy of taxol using A549 tumor-bearing mice in vivo. Exposure of cells to taxol induced time-dependent cytotoxicity and cytoplasmic vacuolization without the involvement of Bax, Bak, Mcl-1, Bcl-XL, and caspase-3. Although taxol treatment induced activating transcription factor 6 (ATF6) cleavage indicative of endoplasmic reticulum (ER) stress, silencing ATF6 by shATF6 did not prevent taxol-induced both cytotoxcity and cytoplasmic vacuolization, suggesting that taxol-induced cytoplasmic vacuolization and cell death were not due to ER stress. Moreover, taxol-treated cells did not show DNA fragmentation and loss of mitochondrial membrane potential, the typical characteristics of apoptosis. In addition, taxol-induced cytoplasmic vacuolization did not show the cellular lysis, the characteristics of oncosis, and positive of β-galactosidase, the characteristic of senescence, indicating that taxol induced paraptosis-like cell death is neither oncosis nor senescence. Moreover, our in vivo data showed that intratumoral injection of taxol (50 mg/kg) in A549 tumor xenograft mice on day 1 and day 19 potently suppressed tumor growth showing significant ER vacuolization without toxicity. In conclusion, high concentration of taxol exhibits a significant anticancer activity by inducing paraptosis-like cell death in vitro and in vivo, without significant toxicity, suggesting a promising therapeutic strategy for apoptosis-resistance cancer by inducing ER vacuolization.

In vitro Studies on anticancer activity of fungal taxol against human breast cancer cell line MCF-7 cells

ObjectiveTo prove the anticancer activity of fungal taxol obtained from Pestalotiopsis pauciseta VM1 endophytic fungus of Tabebuia pentaphylla on human breast cancer cell line MCF-7 by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay.record ethnobotanical information from a hill-dwelling aboriginal tribe of Odisha. MethodsDifferent concentrations of fungal taxol ranging from 100 μg to 700 μg were tested against the MCF-7 breast cancer cell line showed significant decrease in the concentration of 350 μg. ResultsThis cell viability of control cells was consistently 85–90%, The cell shrinkage increased progressively. ConclusionsThus, the fungal taxol isolated from Pestalotiopsis pauciseta VM1, exhibited a very high degree of in vitro cytotoxic activity against MCF-7 breast cancer cell line.

Isolation of taxol, an anticancer drug produced by the endophytic fungus, Phoma betae

Phoma betae, an endophytic fungus, was isolated from the healthy leaves of Ginkgo biloba . The fungus was screened for the production of taxol on a modified liquid medium for the first time. The fungal species were identified by their characteristic culture morphology and molecular analysis. The presence of taxol was confirmed by spectroscopic: ultraviolet (UV), infrared (IR), nuclear magnetic resonance (NMR), liquid chromatography–mass spectrometry (LC-MS); and chromatographic: thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) methods of analysis. The taxol production was quantified by HPLC analysis. The maximum amount of taxol production was recorded as 795 μg/L. The production rate was 15,900-fold more than that found in the culture broth of earlier reported fungus, Taxomyces andreanae . The extracted fungal taxol demonstrate a strong cytotoxic activity in the in vitro culture of tested human cancer cells by apoptotic assay. This indicates that the increase in taxol concentration induced an increased cell death. A polymerase chain reaction (PCR) based screening for taxadiene synthase (ts), a unique gene in the formation of the taxane skeleton was confirmed as a molecular blueprint for taxol biosynthesis. These results designate that the fungus, P. betae is an excellent candidate for taxol supply and can serve as a potential species for genetic engineering to enhance the production of taxol to a higher level. Key words : Taxol production, Phoma betae, analytical methods, cytotoxicity assay, taxadiene synthase.

In vitro studies on anticancer activity of fungal taxol against human breast cancer cell line MCF-7 cells

The present study was taken to prove the anticancer activity of fungal taxol obtained from Pestalotiopsis pauciseta VM1 endophytic fungus of Tabebuia pentaphylla on human breast cancer cell line MCF-7 by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Different concentrations of fungal taxol ranging from 100μg to 700μg were tested against the MCF-7 breast cancer cell line showed significant decrease in the concentration of 350μg. Therefore, the IC50 value of the fungal taxol was calculated as 350μg/mL.

Impact of taxol on dermal papilla cells — A proteomics and bioinformatics analysis

Dermal papilla (DP) cells play a regulatory role in hair growth, and also play a role in alopecia (hair loss). However, effects of taxol, which is a widely used chemotherapy drug, on DP cells remain unclear, despite that theoretically taxol can impact on DP cells to contribute to taxol-induced alopecia. To better understand pathophysiology of taxol-induced damage in DP cells, morphological and biochemical analyses were performed to check whether taxol can cause apoptosis in cultured DP cells or not. If it can, proteomics and bioinformatics analyses were then performed to investigate the protein networks which are impacted by the taxol treatment. Our data showed that taxol can cause apoptotic damage in DP cells in a concentration-dependant manner, as demonstrated by various apoptotic markers. Proteomic analysis on DP cells treated with the lowest apoptosis-inducible concentration of taxol revealed that taxol can affect expression of proteins involved in Ca2+-regulated biological processes, vesicles transport, protein folding, reductive detoxification, and biomolecules metabolism. Furthermore, bioinformatics analysis indicated that taxol can impact on multiple biological networks. Taken together, this biochemical, proteomics, and bioinformatics data may give an insight into pathophysiology of taxol-induced damage in DP cells and shed light on mechanisms underlying taxol-induced alopecia.

Sensitization of breast cancer cells to taxol by inhibition of taxol resistance gene 1

The purpose of this study was to determine the role of taxol resistance gene 1 (TXR1) in a taxol-resistant breast cancer cell line. Expression levels of TXR1 and thrombospondin-1 (TSP1) were detected by RT-PCR and Western blot analysis. In MCF-7 cells transfected with TXR1 plasmids, the expression of a number of drug resistance genes was monitored, as well as cell proliferation. MCF-7 cells were also transiently transfected with a chemically synthesized small interfering RNA (siRNA) targeting TXR1. Taxol concentrations varying from 0.6 to 6 μg/ml were shown to inhibit the proliferation of MCF-7 cells in a time- and dose-dependent manner by arresting cells in the G2/M phase. Additionally, 0.06 μg/ml taxol was used to establish a taxol-resistant MCF-7 cell line, MCF-7/T. When TXR1 was exogenously expressed, a taxol-resistant phenotype was further induced and the expression levels of BCRP, GSTP1 and MVP increased. Transient transfection of TXR1-targeting siRNAs was shown to restore the chemosensitivity of MCF-7 cells. These results suggest that an increased expression of TXR1 is a novel mechanism for reversing the taxol resistance of breast cancer cells.

The ability of the mouse lymphoma TK assay to detect aneugens

There is some evidence that the mouse lymphoma TK assay (MLA) can detect aneugens, and this is accepted in the current International Conference on Harmonisation guidance for testing pharmaceuticals. However, whether or not it can be used as a reliable screen for aneugenicity has been the subject of debate. Consequently, aneugens with diverse mechanisms of action were tested in the MLA using 24-h exposure. No evidence of increased mutant frequency was seen with noscapine, diazepam or colchicine and increases were seen with taxol, carbendazim, econazole and chloral hydrate only at high levels of toxicity (for all but one taxol concentration survival reduced to ≤10% of control). None of these agents would be unequivocally classified as positive using currently accepted criteria. The largest increases in mutant number were seen with taxol and carbendazim; therefore, trifluorothymidine (TFT)-resistant clones resulting from treatment with them were cultured and analysed for chromosome 11 copy number using fluorescent in situ hybridisation (FISH) and loss of heterozygosity (LOH). High concentrations of these aneugens induced LOH at all loci examined indicating only one chromosome 11 was present but, perhaps surprisingly, all were found to have two copies of chromosome 11 using FISH. This would be consistent with loss of the tk(+) chromosome 11b with concomitant duplication of chromosome 11a, which has been proposed as a likely mechanism for induction of TFT-resistant clones. However, it was also surprising that analysis of centromere size showed that almost all the clones had both small and large centromeres, i.e. suggesting the presence of both chromosomes 11a and 11b. In conclusion, it appears that the TFT-resistant mutants resulting from treatment with toxic concentrations of some aneugens such as taxol and carbendazim have undergone complex genetic changes. However, these data show that the MLA cannot be used as a routine screen to detect aneugens.

Regulation of Retinoschisin Secretion in Weri-Rb1 Cells by the F-Actin and Microtubule Cytoskeleton

Retinoschisin is encoded by the gene responsible for X-linked retinoschisis (XLRS), an early onset macular degeneration that results in a splitting of the inner layers of the retina and severe loss in vision. Retinoschisin is predominantly expressed and secreted from photoreceptor cells as a homo-oligomer protein; it then associates with the surface of retinal cells and maintains the retina cellular architecture. Many missense mutations in the XLRS1 gene are known to cause intracellular retention of retinoschisin, indicating that the secretion process of the protein is a critical step for its normal function in the retina. However, the molecular mechanisms underlying retinoschisin's secretion remain to be fully elucidated. In this study, we investigated the role of the F-actin cytoskeleton in the secretion of retinoschisin by treating Weri-Rb1 cells, which are known to secrete retinoschisin, with cytochalasin D, jasplakinolide, Y-27632, and dibutyryl cGMP. Our results show that cytochalasin D and jasplakinolide inhibit retinoschisin secretion, whereas Y-27632 and dibutyryl cGMP enhance secretion causing F-actin alterations. We also demonstrate that high concentrations of taxol, which hyperpolymerizes microtubules, inhibit retinoschisin secretion. Our data suggest that retinoschisin secretion is regulated by the F-actin cytoskeleton, that cGMP or inhibition of ROCK alters F-actin structure enhancing the secretion, and that the microtubule cytoskeleton is also involved in this process.

The small GTPase Cdc42 promotes membrane protrusion during polar body emission via ARP2-nucleated actin polymerization

Polar body emission is a specialized cell division throughout the animal kingdom, serving to reduce chromosome ploidy while preserving the egg cytoplasm. Critical to polar body emission are the asymmetric positioning of the meiotic spindle prior to anaphase, with one pole attached to the oocyte cortex, and the simultaneous membrane protrusion during subsequent cytokinesis. We have shown that, during Xenopus oocyte maturation, the small GTPase Cdc42 promotes membrane protrusion while a classical RhoA contractile ring forms and constricts at the base of the protrusion. We report here that treating oocytes with low concentrations of nocodazole diminished the size of metaphase I spindles and prevented polar body emission, and yet an active Cdc42 cap of correspondingly diminished size still developed, on time, atop of the spindle pole. Conversely, treating oocytes with low concentrations of taxol resulted in a spindle with multiple poles attached to the cortex, but still each of these poles were associated with activated cortical Cdc42 at the appropriate time. Therefore, the asymmetric positioning of the meiotic spindle with one pole anchored to the cortex is a prerequisite for Cdc42 activation. Furthermore, we demonstrated that the Cdc42-regulated F-actin nucleator ARP2/3 complex was similarly localized at the cortex of the protruding polar body membrane, suggesting that Cdc42 promotes membrane protrusion through an F-actin meshwork mechanism. Finally, we demonstrated that Cdc42 and RhoA formed similarly complementary activity zones during egg activation and that inhibition of Cdc42 prevented second polar body emission. Therefore, Cdc42 activation likely promotes membrane protrusion during polar body emission in widespread systems.

Abstract 2130: Overcoming chemotherapy resistance by targeting LDHA mediated glycolysis

Abstract Taxol is one of the most effective chemotherapeutic agents for the treatment of patients with breast cancer. Despite impressive clinical responses initially, the majority of patients eventually develop resistance to Taxol. Lactate dehydrogenase-A (LDH-A) is one of the predominant isoforms of LDH expressed in breast tissue, which controls the conversion of pyruvate to lactate and plays an important role in glucose metabolism. In this study we investigated the role of LDH-A in mediating Taxol resistance in human breast cancer cells. Taxol-resistant subclones, derived from the cancer cell line MDA-MB-435, sustained continuous growth in high concentrations of Taxol while the Taxol-sensitive cells could not. The increased expression and activity of LDH-A were detected in Taxol-resistant cells when compared with their parental cells. The downregulation of LDH-A by siRNA significantly increased the sensitivity of Taxol-resistant cells to Taxol. A higher sensitivity to the specific LDH inhibitor, oxamate, was found in the Taxol-resistant cells. Furthermore, treating cells with the combination of Taxol and oxamate showed a synergistical inhibitory effect on Taxol-resistant breast cancer cells by promoting apoptosis in these cells. LDH-A plays an important role in Taxol resistance and inhibition of LDH-A re-sensitizes Taxol-resistant cells to Taxol. This supports that Warburg effect is a property of Taxol resistant cancer cells and may play an important role in the development of Taxol resistance. To our knowledge, this is the first report showing that the increased expression of LDH-A plays an important role in Taxol resistance of human breast cancer cells. This study provides valuable information for the future development and use of targeted therapies, such as oxamate, for the treatment of patients with Taxol-resistant breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2130. doi:10.1158/1538-7445.AM2011-2130

Development of sulfhydrylated antibody functionalized microcantilever immunosensor for taxol

An immobilization method using a sulfhydrylated monoclonal antibody (mAb) on the gold surface of a microcantilever immunosensor was developed for label-free detection of taxol. The sulfhydrylated anti-taxol mAb was synthesized. Then it was immobilized on the gold surface of the microcantilever. The deflection of the microcantilever corresponding to different taxol concentrations was real time monitored by optical lever technique. The activity of the anti-taxol mAb before and after sulfhydrylation, and the binding of the sulfhydrylated anti-taxol mAb on the gold surface of the microcantilever were evaluated by non-competitive enzyme-linked immunosorbent assay (ELISA). The limit of detection is 1 ng/mL and the method was used for detection of taxol in human plasma sample.

Sequential administration of camptothecin sensitizes human colon cancer HCT116 cells to paclitaxel via p21Cip1/WAF1

Colorectal cancer is the third leading cause of cancer-related death in Western countries. Chemotherapeutic agents with different mechanisms of action have shown an increase in cure rates. In the present study, we investigated the effect of a combination of low concentration of paclitaxel (taxol, 5 nM) and topoisomerase 1 inhibitor camptothecin (CPT) on HCT116 colon cancer cells. Although the viability of cells treated with taxol alone was similar to that of control cells, sequential treatment with taxol and CPT exhibited high cytotoxicity. However, the opposite sequence of treatment did not exert cytotoxic effects on HCT116 cells. This enhanced cytotoxicity of the sequential combination therapy was the result of mitotic arrest, which increased the level of p21Cip1/WAF1 through the p38 mitogen-activated protein kinase (MAPK) pathway. Knockdown by p21Cip1/WAF1 siRNA or treatment with a p38 inhibitor reduced the viability of cells sequentially exposed to taxol and CPT. Taken together, a low taxol concentration in combination with CPT induced mitotic arrest in HCT116 cells, leading to synergistic cell death through enhanced expression of p21Cip1/WAF1 and p38 MAPK pathway. Therefore, taxol could play a role as a sensitizer of CPT in colon cancer cells.

In vitro screening of taxol, an anticancer drug produced by the fungus, Colletotrichum capsici

AbstractThe fungus Colletotrichum capsici was isolated from the diseased fruits of Chilli plant, Capsicum annuum. The isolated test fungus was identified by its morphological and molecular characteristic features. For the first time, the fungus was screened for the production of taxol on modified liquid medium. The presence of taxol was confirmed by the spectroscopic and chromatographic methods of analyses. The amount of taxol produced by this fungus was quantified by HPLC. The maximum amount of fungal taxol production was recorded as 687 μg/L. The production rate was 13 740‐fold higher than that, previously reported for the fungus Taxomyces andreanae. The extracted fungal taxol showed a strong cytotoxic activity in an in vitro culture of human cancer cells indicating that the increase in taxol concentration induces increased cell death. A PCR‐based screening for taxadiene synthase (ts), a unique gene in the formation of the taxane skeleton, confirmed the molecular blueprint for taxol biosynthesis. The results show that the fungus C. capsici is an excellent candidate for an alternate source of taxol supply and can serve as a potential species for genetic engineering to enhance the production of taxol to a higher level.