Different Concentrations Of Paclitaxel Research Articles

Time- and Concentration-Dependent Adverse Effects of Paclitaxel on Non-Neuronal Cells in Rat Primary Dorsal Root Ganglia.

Paclitaxel is a chemotherapeutic agent used to treat a wide range of malignant tumors. Although it has anti-tumoral properties, paclitaxel also shows significant adverse effects on the peripheral nervous system, causing peripheral neuropathy. Paclitaxel has previously been shown to exert direct neurotoxic effects on primary DRG neurons. However, little is known about paclitaxel's effects on non-neuronal DRG cells. They provide mechanical and metabolic support and influence neuronal signaling. In the present study, paclitaxel effects on primary DRG non-neuronal cells were analyzed and their concentration or/and time dependence investigated. DRGs of Wister rats (6-8 weeks old) were isolated, and non-neuronal cell populations were separated by the density gradient centrifugation method. Different concentrations of Paclitaxel (0.01 µM-10 µM) were tested on cell viability by MTT assay, cell death by lactate dehydrogenase (LDH) assay, and propidium iodide (PI) assay, as well as cell proliferation by Bromodeoxyuridine (BrdU) assay at 24 h, 48 h, and 72 h post-treatment. Furthermore, phenotypic effects have been investigated by using immunofluorescence techniques. Paclitaxel exhibited several toxicological effects on non-neuronal cells, including a reduction in cell viability, an increase in cell death, and an inhibition of cell proliferation. These effects were concentration- and time-dependent. Cellular and nuclear changes such as shrinkage, swelling of cell bodies, nuclear condensation, chromatin fragmentation, retraction, and a loss in processes were observed. Paclitaxel showed adverse effects on primary DRG non-neuronal cells, which might have adverse functional consequences on sensory neurons of the DRG, asking for consideration in the management of peripheral neuropathy.

Palmitoylethanolamide Mitigates Paclitaxel Toxicity in Primary Dorsal Root Ganglion Neurons.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of several chemotherapeutic agents, such as Paclitaxel. The main symptoms of CIPN are pain and numbness in the hands and feet. Paclitaxel is believed to accumulate in the dorsal root ganglia and free nerve endings. Novel therapeutic agents might help to mitigate or prevent Paclitaxel toxicity on dorsal root ganglion (DRG) neurons. Thus, we used primary DRG neurons as a model to investigate the potential neuroprotective effects of the endocannabinoid-like substance, palmitoylethanolamide (PEA). DRG neurons were isolated from cervical to sacral segments of spinal nerves of Wister rats (6-8 weeks old). After isolation and purification of neuronal cell populations, different concentrations of Paclitaxel (0.01-10 µM) or PEA (0.1-10 µM) or their combination were tested on cell viability by MTT assay at 24 h, 48, and 72 h post-treatment. Furthermore, morphometric analyses of neurite length and soma size for DRG neurons were performed. Adverse Paclitaxel effects on cell viability were apparent at 72 h post-treatment whereas Paclitaxel significantly reduced the neurite length in a concentration-dependent manner nearly at all investigated time points. However, Paclitaxel significantly increased the size of neuronal cell bodies at all time windows. These phenotypic effects were significantly reduced in neurons additionally treated with PEA, indicating the neuroprotective effect of PEA. PEA alone led to a significant increase in neuron viability regardless of PEA concentrations, apparent improvements in neurite outgrowth as well as a significant decrease in soma size of neurons at different investigated time points. Taken together, PEA showed promising protective effects against Paclitaxel-related toxicity on DRG neurons.

Paclitaxel induces cognitive impairment via necroptosis, decreased synaptic plasticity and M1 polarisation of microglia

Context Paclitaxel (PTX) leads to chemotherapy brain (chemo-brain) which is characterised by cognitive impairment. It has been reported that necroptosis is associated with cognitive impairment in some neurodegenerative diseases, but it is not clear whether it is related to the development of chemo-brain. Objective To investigate the role of necroptosis and related changes in PTX-induced cognitive impairment. Materials and methods C57bl/6n mice were randomly divided into five groups: control, vehicle, and different concentrations of PTX (6, 8, 10 mg/kg). Two additional groups received pre-treatment with Gdcl3 or PBS through Intracerebroventricular (ICV) injection before PTX-treatment. Cognitive function, necroptosis, synaptic plasticity and microglia polarisation were analysed. Results PTX (10 mg/kg) induced significant cognitive impairment, accompanied by changes in synaptic plasticity, including decreased density of PSD95 (0.65-fold), BDNF (0.44-fold) and dendritic spines (0.57-fold). PTX induced necroptosis of 53.41% (RIP3) and 61.91% (MLKL) in hippocampal neurons, with high expression of RIP3 (1.58-fold) compared with the control group. MLKL (1.87-fold) exhibited the same trend, reaching a peak on the 14th day. The increased expression of iNOS (1.63-fold) and inflammatory factors such as TNF-α (1.85-fold) and IL-β (1.89-fold) compared to the control group suggests that M1 polarisation of microglia is involved in the process of cognitive impairment. Pre-treatment with Gdcl3 effectively reduced the number of microglia (0.50-fold), inhibited the release of TNF-α (0.73-fold) and IL-β (0.56-fold), and improved cognitive impairment. Conclusion We established a stable animal model of PTX-induced cognitive impairment and explored the underlying pathophysiological mechanism. These findings can guide the future treatment of chemo-brain.

LINC00337 induces tumor development and chemoresistance to paclitaxel of breast cancer by recruiting M2 tumor-associated macrophages

BackgroundM2 tumor-associated macrophages are closely related to the progression and prognosis of breast cancer (BCa), and could be regulated by long intergenic non-coding RNAs (lincRNAs). Moreover, the differential expression of lincRNAs affects tumor resistance. This study focused on the potential involvement and mechanism of LINC00337 in BCa. MethodsThe expression of LINC00337 in BCa was detected by bioinformatics analysis and RT-qPCR. Cell viability and proliferation were analyzed by cell counting kit-8 (CCK-8) and clone formation assay. BCa cells were treated with different concentrations of paclitaxel (PAX) to determine the chemotherapy resistance of LINC00337. Tumor formation assay, Western blot, ELISA and immunohistochemistry were performed to determine the relationship between LINC00337 and PAX in vivo. Macrophages were induced to M2-like polarization, and then functional experiments (CCK-8, wound healing) and molecular experiments (ELISA, RT-qPCR, Western blot) were used to verify the role of LINC00337. ResultsLINC00337 was up-regulated in BCa. High-expressed LINC00337 accelerated viability and proliferation of BCa cells, improved the resistance of BCa cells to PAX, and accelerated tumor growth. Overexpressed LINC00337 up-regulated the expressions of M2 macrophage markers and M-CSF, and reduced the level of GM-CSF. PAX significantly reduced the viability of BCa cells and down-regulated LINC00337. Furthermore, the successfully induced M2 type macrophages to promote BCa cell activity, migration and EMT protein expression, and LINC00337 enhanced the effect of M2 type macrophages. ShLINC00337 had the opposite effect to overexpressed LINC00337. ConclusionLINC00337 accelerated the malignant phenotype of BCa cells and promoted chemoresistance to paclitaxel through M2-like macrophages.

Overexpression of ARHI increases the sensitivity of cervical cancer cells to paclitaxel through inducing apoptosis and autophagy.

Cervical cancer (CC) is a common malignant tumor of the female reproductive system. This study investigated the role of aplysia ras homolog I (ARHI) in resistance to CC in vitro and in patients' tissues. Hela cells were continuously treated with different concentrations of paclitaxel (1-10nM) to construct paclitaxel-resistant cell model (Hela-TR). CC or CC-TR tissues were obtained from CC patients or CC patients who had developed paclitaxel resistance. The level of ARHI and multidrug resistance gene 1 (MDR1) in cells and tissues were detected by qRT-PCR and immunohistochemistry (IHC) staining. Cell viability, apoptosis and the number of colonies were assessed by MTT, flow cytometry and cell clone assay in Hela and Hela-TR cells after the ARHI plasmid or shARHI were transfected into cells. The autophagy and apoptosis signaling related proteins were analyzed by western blotting. The results revealed that the levels of ARHI mRNA and protein were down-regulated in CC tissues, and were further reduced in paclitaxel-resistant tissues and Hela cell model. High expression of ARHI inhibited the expression of MDR1 in Hela and Hela-TR cells. The cell viability and cell clone of Hela and Hela-TR cells were decreased by ARHI overexpression but increased by ARHI suppression. In addition, highly expressed ARHI promoted apoptosis and activated autophagy by increasing LC3-II/LC3-I through inactivating AKT/mTOR signaling pathway. In conclusion, overexpression of ARHI can increase the sensitivity of CC to paclitaxel through promoting apoptosis and autophagy in a AKT/mTOR inactivation dependent pathway.

Propofol enhanced the cell sensitivity to paclitaxel (PTX) in prostatic cancer (PC) through modulation of HOTAIR.

PTX is widely used in cancer treatments. In this paper, we explored the role and potential molecular mechanism of propofol in regulating PTX sensitivity in PC cells. Prostatic cancer cell line PC3 was treated using different concentrations of PTX (10nM, 50nM), propofol (150μM, 300μM) or transfected with overexpressed HOTAIR plasmid. HOTAIR expression was analyzed by RT-qPCR. Apoptosis of PC3 cells was observed by flow cytometry method while cell viability was evaluated by CCK-8. Moreover, apoptosis-related genes, Bcl-2 and Bax were detected by Western blot methods. E-cadherin, N-cadherin and Vimentin protein concentrations were monitored by ELISA. PTX significantly increased apoptosis of PC3 cells and reduced cell viability in a dose-dependent manner. Moreover, Protein expression of Bcl-2 was obviously inhibited while Bax protein expression level was provoked. Furthermore, E-cadherin protein concentration increased while N-cadherin and Vimentin decreased due to increasing PTX treatments. HOTAIR expression dropped due to PTX treatment while overexpression of HOTAIR induced cell viability, EMT and deterred apoptosis. Propofol ignited the PTX function while upregulation of HOTAIR partially reversed this. Propofol enhanced paclitaxel sensitivity in prostatic cancer cells through modulation of HOTAIR in vitro.

MiR-146a Enhances the Sensitivity of Breast Cancer Cells to Paclitaxel by Downregulating IRAK1.

Objective: To investigate the effect of miR-146a on the sensitivity of breast cancer cells to paclitaxel (PTX). Materials and Methods: The mRNA expressions of miR-146a in normal breast cancer cells, MCF-7, and PTX-resistant breast cancer cells, MCF-7/PTX, were detected by reverse transcriptase-polymerase chain reaction (RT-PCR). MTS was used to analyze the cytotoxicity treated with different concentrations of PTX. Overexpressed and silenced cell lines of miR-146a and interleukin-1 receptor-associated kinase 1 (IRAK1) were constructed, respectively. Cells were treated with PTX and observed the changes of cell morphology. Proliferation was detected by clone formation assay. Invasion and migration were measured by transwell. RT-PCR was applied to detect the expression of IRAK1 gene. Dual luciferase report was performed to validate the target relationship between miR-146a and IRAK1. Salvage experiments were used to further verify the relationship between miR-146a and IRAK1. Results: PTX reduces the viability of MCF-7 and MCF-7/PTX cells in a dose-dependent manner. The IC50 of PTX in MCF-7 cells was significantly lower compared with MCF-7/PTX cells (p < 0.05). Compared with MCF-7/PTX cells, the expression of miR-146a gene in MCF-7 cells was significantly increased, while the expression of IRAK1 gene was significantly reduced (p < 0.05). Cell proliferation, invasion, and migration were decreased after miR-146a overexpression or IRAK1 silencing. Whereas, miR-146a silencing and IRAK1 overexpression can increase cell proliferation, invasion, and migration ability. Salvage experiments further verify that IRAK1 can weaken the role of miR-146a. Conclusion: miR-146a can enhance the sensitivity of breast cancer cells to PTX; the mechanism may be related to the downregulation of IRAK1.

Inhibitory effects of everolimus in combination with paclitaxel on adriamycin-resistant breast cancer cell line MDA-MB-231

ObjectiveWe aimed to evaluate the therapeutic effects of paclitaxel in combination with mTOR inhibitor everolimus on adriamycin-resistant breast cancer cell line MDA-MB-231 (MDA-MB-231/ADR). Materials and methodsMDA-MB-231/ADR cells were treated with different concentrations of paclitaxel and everolimus. The IC50 values after 48 h of treatment were measured by the MTT assay. The apoptosis rate and cell cycle were detected by flow cytometry. The protein expressions of Akt, PI3K, mTOR, p-pI3K, p-AKT and p-mTOR were detected by Western blot. ResultsWhen paclitaxel at ≥1.56 μg/ml was used, the growth of MDA-MB-231/ADR cells was inhibited more significantly than that of control group (P < 0.05). After treatment with ≥6.25 μg/ml everolimus, the cell growth was also suppressed more significantly (P < 0.05). The IC50 values of everolimus and paclitaxel were 32.50 μg/ml and 7.80 μg/ml, respectively. The inhibition rate of paclitaxel plus everolimus was significantly enhanced with increasing paclitaxel concentration (P < 0.001). After treatment with 7.80 μg/ml paclitaxel, the two drugs had best synergistic inhibitory effects on proliferation. Compared with drugs alone, the combination significantly promoted apoptosis (P < 0.001). The paclitaxel + everolimus group had significantly more cells in the G0-G1 phase than those of control and individual drug groups (P < 0.001). Everolimus significantly decreased mTOR and p-mTOR expressions compared with those of control group (P < 0.001). Compared with everolimus alone, the combination reduced the expressions more significantly (P < 0.05). Paclitaxel decreased the expression levels of PI3K, p-PI3K and p-AKT. Compared with paclitaxel alone, the combination significantly promoted the reduction of PI3K, p-PI3K and p-AKT expressions (P < 0.05). ConclusionEverolimus can enhance the effect of paclitaxel on MDA-MB-231/ADR cells, inhibit cell proliferation, induce apoptosis and arrest cell cycle in the G1 phase mainly by down-regulating the expressions of key proteins in the mTOR signaling pathway.

The Effect of Paclitaxel on the Viability of U14 Cells

The research is aimed to study the effect of paclitaxel on the viability of U14 cell line of cervical cancer, and to provide new ideas for further exploring the mechanism of paclitaxel chemotherapy to cervical cancer. Then, different concentrations of paclitaxel were used to treat U14 cells of cervical cancer in logarithmic phase growth. After culturing these cells for 24h, 48h or 72h, MTT method and automatic enzyme-linked immunosorbent assay system were used to evaluate the survival rate of cultured cells with different concentrations of paclitaxel. These results showed that paclitaxel has a significant inhibitory effect on the proliferation of U14 cells in a concentration- and time-dependent manner, compared with the control group. Paclitaxel concentration of the IC50 of U14 cells was 168.8μg/ml at 24h, 22. 15 μg/ml at 48h and 8. 04μg/ml at 72 h. As the time increased, the IC50 value of U14 cells gradually decreased. The results of linear regression analysis are as follows: 24 h, y=-0.005x + 1.344, R²=0.779; 48 h, y=-0.013x +0.788, R²=0.923; 72 h, y=-0.056x + 0.950, R²=0.908. The dose-effect curve of paclitaxel on U14 cells for 48 h and the trend of linear regression fitting are better, and the growth inhibition of cells shows a clear relationship between time and dose. In conclusion, paclitaxel has an obvious inhibitory effect on the activity of cervical cancer U14 cells in mice, which provides new ideas for further exploring the mechanism of cervical cancer paclitaxel chemotherapy.

Overexpression of GATA5 Stimulates Paclitaxel to Inhibit Malignant Behaviors of Hepatocellular Carcinoma Cells.

Objective Explore the effect of GATA5 expression on Paclitaxel inhibiting growth of hepatocellular carcinoma (HCC) cells. Materials and Methods In the experimental study, HCC cell lines (HLE, Bel7402 and PLC/PRF/5) were treated with different concentrations of Paclitaxel (5-20 mg/ml) for 24 hours. HLE cells were transfected with GATA5-siRNA vector, while Bel7402 and PLC/PRF/5 cells were transfected with overexpressed GATA5 vector for 24 hours, followed by treatment of the cells with Paclitaxel (10 mg/ml) for 24 hours and subsequently 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay to detect growth of HCC cells. Soft agar cultured was used to analyze formation of colony. Apoptosis of HCC cells were detected by Flow cytometer. Migration of HCC cells was observed by trawell assays. Western blotting and laser confocal microscopy were utilized to detect expression and location of the proteins. Results Inhibiting expression of GATA5 reduced sensitivity of HLE cells to Paclitaxel, while overexpression of GATA5 increased sensitivity of Bel7402 cells and PLC/PRF/5 cells to Paclitaxel. Overexpression of GATA5 played a role in stimulating Paclitaxel to inhibit growth, colony formation and migration, as well as enhance apoptosis in HCC cells. Overexpression of GATA5 also promoted Paclitaxel to inhibit expression of reprogramming genes, such as Nanog, EpCAM, c-Myc and Sox2 in Bel7402 and PLC/PRF/5 cells. Inhibited expression of GATA5 led to enhancement of the expression of CD44 and CD133, in HLE cells. Overexpression of GATA5 was not only alone but also synergized with Paclitaxel to inhibit expression of CD44 and CD133 in Bel7402 or PLC/PRF/5 cells. ConclusionOverexpression of GATA5 played a role in enhancing Paclitaxel to inhibit the malignant behaviors of HCC cells. It was involved in suppressing expression of the reprogramming genes and stemness markers. Targeting GATA5 is an available strategy for applying paclitaxel to therapy of patients with HCC.

Paclitaxel-Loaded Nanosponges Inhibit Growth and Angiogenesis in Melanoma Cell Models.

This study investigated the effects of free paclitaxel (PTX) and PTX-loaded in pyromellitic nanosponges (PTX-PNS) in reducing in vitro and in vivo melanoma cell growth and invasivity, and in inhibiting angiogenesis. To test the response of cells to the two PTX formulations, the cell viability was evaluated by MTT assay in seven continuous cell lines, in primary melanoma cells, both in 2D and 3D cultures, and in human umbilical vein endothelial cells (HUVECs) after exposure to different concentrations of PTX or PTX-PNS. Cell motility was assessed by a scratch assay or Boyden chamber assay, evaluating cell migration in presence or absence of diverse concentrations of PTX or PTX-PNS. The effect of PTX and PTX-PNS on angiogenesis was evaluated as endothelial tube formation assay, a test able to estimate the formation of three-dimensional vessels in vitro. To assess the anticancer effect of PTX and PTX-PNS in in vivo experiments, the two drug formulations were tested in a melanoma mouse model obtained by B16-BL6 cell implantation in C57/BL6 mice. Results obtained were as follows: 1) MTT analysis revealed that cell proliferation was more affected by PTX-PNS than by PTX in all tested cell lines, in both 2D and 3D cultures; 2) the analysis of the cell migration showed that PTX-PNS acted at very lower concentrations than PTX; 3) tube formation assay showed that PTX-PNS were more effective in inhibiting tube formation than free PTX; and 4) in vivo experiments demonstrated that tumor weights, volumes, and growth were significantly reduced by PTX-PNS treatment with respect to PTX; the angiogenesis and the cell proliferation, detected in the tumor samples with CD31 and Ki-67 antibodies, respectively, indicated that, in the PTX-PNS-treated tumors, the tube formation was inhibited, and a low amount of proliferating cells was present. Taken together, our data demonstrated that our new PTX nanoformulation can respond to some important issues related to PTX treatment, lowering the anti-tumor effective doses and increasing the effectiveness in inhibiting melanoma growth in vivo.

Bi-layer blood vessel mimicking microfluidic platform for antitumor drug screening based on co-culturing 3D tumor spheroids and endothelial layers.

Two-dimensional (2D) cell culture is not ideal for traditional drug screening, because 2D culture does not accurately mimic the physiological microenvironment of tumor cells. Thus, a drug-screening system which more closely mimics the microenvironment of in vivo tumors is necessary. Here, we present a biomimicking bilayer microfluidic device that can facilitate antitumor drug screening. The microfluidic device consists of two polydimethylsiloxane (PDMS) pieces with channels which are separated by a semipermeable membrane to allow water, oxygen, and nutrition supply, while preventing cell migration. The channels embedded on the two PDMS pieces overlap each other over a long distance to ensure a larger exchange area to mimic the blood vessel-tumor model. High concentrations of endothelial cells (EC) are first seeded onto the membrane through the apical channel, and after a two-day culture, a confluent EC monolayer forms. Tumor spheroid-laden Matrigel is then seeded into the basal channel. After the Matrigel is cured, the device is ready for drug testing. Paclitaxel is used as the model drug for testing. Confocal microscopy and ImageJ are used to assess the efficacy of different concentrations of paclitaxel, and optical coherence tomography (OCT) is employed to determine the tumor volumetric change after the drug treatment. The results indicate that the proposed bilayer microfluidic device in combination with confocal and OCT optical characterization provide an efficient platform for antitumor drug testing.

Abstract 5893: Expression and role of autophagy-associated p62 (SQSTM1) in multidrug-resistant ovarian cancer

Abstract Multidrug resistance is the major cause of treatment failure in ovarian cancer. p62 (SQSTM1) is a multifunctional protein involved in multiple cellular processes including proliferation, drug sensitivity and autophagy-associated cancer cell growth. p62 is a critical indicator of autophagic flux, which is inversely associated with autophagy activity. However, the role of p62 remains controversial in drug resistance in human ovarian cancer. In this study, we examined p62 expression by immunohistochemistry in a unique ovarian cancer tissue microarray (TMA), which was constructed with paired primary, metastatic, and recurrent tumor tissues from 26 individual patients. Results showed that both the metastatic and recurrent tumor tissues expressed less p62 than the patient-matched primary tumor. A significant inverse correlation has been found between p62 expression and both the disease free survival and overall survival. In addition, multidrug resistant cancer cell lines expressed lower levels of p62 as compared with their parental drug sensitive cell lines. Importantly, cell viabilities determined by MTT assay after exposure to different concentrations of paclitaxel showed inhibition of autophagy or accumulation of p62 enhances paclitaxel sensitivity in ovarian cancer drug resistant cells. Furthermore, the wound healing assay exhibited that inhibition of autophagy significantly decreased multidrug resistant ovarian cancer cell migration in vitro. Collectively, these data highlight that autophagy pathway may be a promising therapeutic target to prevent metastasis, recurrence and to reverse drug resistance in ovarian cancer. Citation Format: Jinglu Wang, Cassandra Garbutt, Francis J Hornicek, Zhenfeng Duan. Expression and role of autophagy-associated p62 (SQSTM1) in multidrug-resistant ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5893.

Expression and role of autophagy-associated p62 (SQSTM1) in multidrug resistant ovarian cancer

ObjectiveMultidrug resistance is the major cause of treatment failure in ovarian cancer. p62 (SQSTM1) is a multifunctional protein involved in multiple cellular processes including proliferation, drug sensitivity and autophagy-associated cancer cell growth. However, the role of p62 in drug resistance remains controversial. MethodsIn this study, we examined p62 expression by immunohistochemistry in a unique ovarian cancer tissue microarray (TMA), which was constructed with paired primary, metastatic, and recurrent tumor tissues. The expression levels of p62 and autophagy related proteins were evaluated in two panels of human cancer cell lines by western blot. Cell viabilities were determined by MTT assay after exposure ovarian cancer cells to different concentrations of paclitaxel alone or in combination with autophagy inhibitors. ResultsBoth the metastatic and recurrent tumor tissues expressed less p62 than the patient-matched primary tumor. A significant inverse correlation has been found between p62 expression and both the disease-free survival and overall survival. Additionally, multidrug resistant cancer cell lines expressed lower levels of p62 as compared with their parental drug sensitive cell lines. Importantly, inhibition of autophagy enhanced paclitaxel sensitivity in drug resistant ovarian cancer cells. Furthermore, the wound healing assay exhibited that the inhibition of autophagy significantly decreased resistant ovarian cancer cell migration in vitro. ConclusionOur findings highlight the potential of p62 as a new prognostic marker for ovarian cancer patients and p62's associated autophagy pathway may be a promising therapeutic target to prevent metastasis, recurrence and to reverse drug resistance in ovarian cancer.

Baker's Yeast Sensitizes Metastatic Breast Cancer Cells to Paclitaxel In Vitro.

Our earlier studies have demonstrated that phagocytosis of baker’s yeast (Saccharomyces cerevisiae) induces apoptosis in different cancer cell lines in vitro and in vivo. This study aimed to examine how baker’s yeast sensitizes murine and human breast cancer cells (BCC) to paclitaxel in vitro. This sensitizing effect makes lower concentrations of chemotherapy more effective at killing cancer cells, thereby enhancing the capacity of treatment. Three BCC lines were used: the metastatic murine 4T1 line, the murine Ehrlich ascites carcinoma (EAC) line, and the human breast cancer MCF-7 line. Cells were cultured with different concentrations of paclitaxel in the presence or absence of baker’s yeast. Cell survival and the IC50 values were determined by MTT assay and trypan blue exclusion method. Percent of DNA damage, apoptosis, and cell proliferation were examined by flow cytometry. Yeast alone and paclitaxel alone significantly decreased 4T1 cell viability postculture (24 and 48 hours), caused DNA damage, increased apoptosis, and suppressed cell proliferation. Baker’s yeast in the presence of paclitaxel increased the sensitivity of 4T1 cells to chemotherapy and caused effects that were greater than either treatment alone. The chemosensitizing effect of yeast was also observed with murine EAC cells and human MCF-7 cells, but to a lesser extent. These data suggest that dietary baker’s yeast is an effective chemosensitizer and can enhance the apoptotic capacity of paclitaxel against breast cancer cells in vitro. Baker’s yeast may represent a novel adjuvant for chemotherapy treatment.

The sensitivity change and its mechanism of cervical cancer cells transfected with miR-200b analogues to paclitaxel

Objective To explore the effect of miR-200b on chemosensitivity of cervical cancer HeLa cells to paclitaxel and its mechanism. Methods A recombinant miR-200b mimics was transfected into cultured HeLa cells, which were divided into observation group, negative group, and blank control group. Each group was treated with different concentrations of paclitaxel. Methyl thiazolyl tetrazolium (MTT) assay was performed to evalate the proliferative ability of these treated cells to paclitaxel. AnnexinV- FITC/PI method was used to detect the apoptosis rate of Hela cells. The protein level of vascular endothelial growth factor (VEGF) was detected by Western blot. Results MTT assay showed that the OD values of three groups were totally different; and compared to the other two groups, the OD value of the transfected HeLa cells was significantly lower (P 0.05). AnnexinV-FITC/PI results demonstrated that the apoptotic index of the transfected HeLa cells was significantly higher than that in the other two groups (P<0.05). Western blot assay showed that expression of VEGF protein in three groups of relative quantity was 0.403±0.046, 0.882±0.094, and 0.901 0.102, respectively. There was significant difference among there groups (P<0.05). Conclusions miR-200b mimics can increase the sensitivity of cervical cancer HeLa cells to paclitaxel, and targeted regulation of VEGF may be the cause of increasing in drug sensitivity. Key words: Transfection; MicroRNAs; Uterine cervical neoplasms/DT; Taxoids/TU

INHIBITION OF PACLITAXEL AGAINST NEUROGLIOMA CELLS U251 GROWTH AND ITS MECHANISM.

Background:Glioma is the most common primary tumor of the central nervous system, and accounted for about 70% of primary tumors.Materials and Methods:In the study, antitumour activity and mechanism of paclitaxel was investigated. Different concentrations of paclitaxel (200, 300, 400 μmol/L) was treated in neuroglioma cellsU251.Results:Paclitaxel significantly inhibited neuroglioma cells growth, and promoted its apoptosis. Paclitaxel can block tumour cells in the G2/M phase. In addition, apoptosis-related genes caspase-3 and bax expressions were increased after paclitaxel treatment.Conclusion:Our work indicated that paclitaxel displayed strong anti-tumour activity.

Paclitaxel inhibits cell proliferation and collagen lattice contraction via TGF-β signaling pathway in human tenon's fibroblasts in vitro

As an anti-microtubule agent, paclitaxel has been widely applied clinically. However, the effects of paclitaxel on human tenon's fibroblast (HTF) proliferation and migration in vitro was still unclear. In the present study, we explored the influences of paclitaxel on HTF cell proliferation, cell viability, cell cycle phase distribution under various concentrations of paclitaxel (i.e., 0, 10−8, 10−7, 10−6mol/l) via real-time cell electronic system and flow cytometry, further determined the expression of TGF-β1 and connective tissue growth factor (CTGF) after treatment with different concentrations of paclitaxel. Moreover, extra cellular matrix production and collagen lattice contraction assay were also explored. The results indicate that paclitaxel could apparently inhibit the cell viability, induces the elevation of S and G2/M phases of HTFs, and downregulates the expression of both TGF-β1 and CTGF. Meanwhile, the levels of fibronectin extra domain A (EDA), collagen and collagen lattice contraction were apparently reduced after treatment with paclitaxel. Overall, paclitaxel could apparently inhibit the proliferation of HTFs and leads to cell cycle arrest at both S and G2/M phases, attenuates the generation of collagen and collagen lattice contraction, decreases the expressions of TGF-β1, CTGF and fibronectin EDA. The inhibitory mechanism of paclitaxel on HTFs is involved in TGF-β1 signaling pathway.

The application of Tau protein testing to gastric cancer patients treated with paclitaxel

Paclitaxel is one of the main drugs used to treat gastric cancer, but many tumors develop drug resistance, resulting in treatment failure. The levels of expression of Tau protein in breast tumors have been found to be related to paclitaxel resistance, suggesting that Tau protein expression may predict breast cancer sensitivity to paclitaxel treatment. To determine whether Tau protein expression can predict gastric cancer sensitivity to paclitaxel, we assayed Tau protein expression levels in gastric cancer specimens from 70 patients. We observed Tau protein expression in 54 of 70 (77.1%) specimens. Assays in gastric cancer cell lines showed that Tau protein expression was significantly lower in BGC823 than in MKN45 cells (P = 0.0147). MTT assays showed that different concentrations of paclitaxel inhibited the growth of MKN45 and BGC823 cells, but inhibition and apoptosis were more obvious in cells expressing low levels of Tau protein. Paclitaxel chemotherapy was effective in 34 of the 70 patients (48.6%) and was significantly correlated with low expression of Tau protein (P < 0.01). These findings indicate that Tau protein is expressed in a high percentage of gastric cancers, with paclitaxel being more effective in tumors with low Tau expression.

Molecular mechanism of chemosensitization to paclitaxel in human melanoma cells induced by targeting the EGFR signaling pathway

To study the molecular mechanism of epidermal growth factor receptor (EGFR) signaling pathway in mediating paclitaxel-resistance and improving paclitaxel sensitivity in human melanoma A375 cells. Human melanoma cell line A375 cells were treated with different concentrations of paclitaxel with or without 20 µmol/L AG1478 (EGFR inhibitor), 40 µmol/L PD98059 (extracellular signal conditioning kinase (ERK) 1/2 blockers) or 10 µmol/L LY294002 (PI3K inhibitor). MTT method was used to measure the proliferation of A375 cells. Flow cytometry was used to detect cell cycle and apoptosis in the A375 cells. The expressions of P-EGFR, P-ERK and P-AKT proteins were determined by Western blot analysis. Paclitaxel (0.001 µmol/L to 0.1 µmol/L) inhibited the growth of A375 cells (P < 0.01) and induced apoptosis (P < 0.05) in a dose- and time-dependent manner. AG1478 (20 µmol/L) increased the 0.01 µmol/L paclitaxel-induced inhibition rate from 38.5% to 62.6% at 72 h. Different doses of paclitaxel induced apoptosis in A375 cells by different ways, in which G0/G1 phase cells were decreased and mitotic phase was prolonged at 0.01 µmol/L, and cell cycle arrest at G2/M phase by 0.1 µmol/L paclitaxel. When DNA damage occurred in A375 cells exposed to paclitaxel, expression of P-EGFR, P-ERK and P-AKT proteins was increased. When EGFR signaling pathway was blocked, paclitaxel did not activate MAPK signaling pathway or PI3K/AKT signaling pathway and did not change its effect on cell cycle in vitro. When EGFR was inhibited by 20 µmol/L tyrophostin AG1478, the 0.001 and 0.01 µmol/L paclitaxel-induced early apoptosis rate in A375 cells was increased by 1.73- and 1.80-fold, respectively. When the ERK signaling was blocked by 40 µmol/L PD98059, the 0.001 and 0.01 µmol/L paclitaxel-induced early apoptosis rate in A375 cells was increased by 2.73- and 2.25-fold, respectively. When the AKT signaling was blocked by 10 µmol/L LY294002, the 0.001 and 0.01 µmol/L paclitaxel-induced early apoptosis rate in A375 cells was increased by 2.02- and 1.46-fold, respectively. Human melanoma A375 cells produce resistance to paclitaxel (0.001 to 0.1 µmol/L) by activating MAPK signaling and PI3K/AKT signaling pathways. Targeting EGFR, ERK and AKT signaling pathways significantly enhances the cytotoxic effect of paclitaxel on human melanoma cells.