Overcome Taxol Resistance Research Articles

Activation of the γ-secretase/NICD-PXR/Notch pathway induces Taxol resistance in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is currently the only subtype lacking efficient targeted therapies. Taxol is the primary chemotherapeutic agent for TNBC. However, Taxol resistance often develops in the treatment of TNBC patients, which importantly contributes to high mortality and poor prognosis in TNBC patients. Recent preclinical studies have shown that the inhibition of Notch pathway by γ-secretase inhibitors can slow down the progression of TNBC. Our studies in bioinformatic analysis of breast cancer patients and TNBC/Taxol cells in vitro showed that there was high correlation between the activation of Notch pathway and Taxol resistance in TNBC. Increased γ-secretase activity (by the overexpression of catalytic core PSEN-1) significantly reduced Taxol sensitivity of TNBC cells, and enhanced biological characteristics of malignancy in vitro, and tumour growth in vivo. Mechanistically, increased γ-secretase activity led to the accumulation of NICD in the nucleus, promoting the interaction between NICD and PXR to activate PXR, which triggered the transcription of PXR downstream associated drug resistance genes. Furthermore, we showed that pharmacological inhibition of γ-secretase with γ-secretase inhibitors (Nirogacestat and DAPT) can reverse Taxol resistance in vivo and in vitro. Our results for the first time demonstrate that the activation of γ −secretase/NCD-PXR/Notch pathway is one of important mechanisms to cause Taxol resistance in TNBC, and the blockades of this pathway may represent a new therapeutic strategy for overcoming Taxol resistance in TNBC.

Thiostrepton induces spindle abnormalities and enhances Taxol cytotoxicity in MDA-MB-231 cells

BackgroundThiostrepton (TST) is a known inhibitor of the transcription factor Forkhead box M1 (FoxM1) and inducer of heat shock response (HSR) and autophagy. TST thus may be one potential candidate of anticancer drugs for combination chemotherapy.Methods and ResultsImmunofluorescence staining of mitotic spindles and flow cytometry analysis revealed that TST induces mitotic spindle abnormalities, mitotic arrest, and apoptotic cell death in the MDA-MB-231 triple-negative breast cancer cell line. Interestingly, overexpression or depletion of FoxM1 in MDA-MB-231 cells did not affect TST induction of spindle abnormalities; however, TST-induced spindle defects were enhanced by inhibition of HSP70 or autophagy. Moreover, TST exhibited low affinity for tubulin and only slightly inhibited in vitro tubulin polymerization, but it severely impeded tubulin polymerization and destabilized microtubules in arrested mitotic MDA-MB-231 cells. Additionally, TST significantly enhanced Taxol cytotoxicity. TST also caused cytotoxicity and spindle abnormalities in a Taxol-resistant cell line, MDA-MB-231-T4R.ConclusionsThese results suggest that, in addition to inhibiting FoxM1, TST may induce proteotoxicity and autophagy to disrupt cellular tubulin polymerization, and this mechanism might account for its antimitotic effects, enhancement of Taxol anticancer effects, and ability to overcome Taxol resistance in MDA-MB-231 cells. These data further imply that TST may be useful to improve the therapeutic efficacy of Taxol.

Discovery of the cereblon-recruiting tubulin PROTACs effective in overcoming Taxol resistance in vitro and in vivo

Overexpression of β3-tubulin is a common occurrence in human tumors and is associated with resistance to microtubule-targeting agents. PROTAC strategy has demonstrated significant potential in overcoming drug resistance. Herein, we report the discovery of W13 as the first PROTAC against tubulin, which was created by connecting a CRBN ligand to the widely recognized microtubule-destabilizing agent CA-4. Notably, it retains the inhibitory activity of the parental CA-4 and further exhibits substantial degradation of α/β/β3-tubulin in both A549 and A549/Taxol cell lines. The degradation of tubulin was subsequently verified to be mediated by the ubiquitin-proteasome system. Importantly, tumor xenograft research clearly showed W13's promising antitumor activity against human lung cancer. Taken together, the discovery of W13 demonstrated the practicality and feasibility of PROTAC targeting tubulin, hence establishing a potential therapeutic approach for treating NSCLC caused by the overexpression of β3-tubulin.

Structure-Based Drug Design and Synthesis of Novel N-Aryl-2,4-bithiazole-2-amine CYP1B1-Selective Inhibitors in Overcoming Taxol Resistance in A549 Cells

As a promising therapeutic target for cancer, CYP1B1 is overexpressed in Taxol-resistant A549 cells; however, its role in drug resistance still remains unclear. Bioinformatic analysis data indicated that CYP1B1 was closely correlated with AKT/ERK1/2 and focal adhesion pathways, thereby playing an important role in Taxol resistance and cancer migration/invasion. Along similar lines, the AhR agonist 7,12-dimethylbenz[a]anthracene (DMBA) enhanced Taxol resistance and promoted migration/invasion of A549 and H460 cells likely stemming from CYP1B1 upregulation. Moreover, 83 novel N-aryl-2,4-bithiazole-2-amine CYP1B1-selective inhibitors were designed and synthesized to verify the role of CYP1B1 in Taxol-resistant A549 cells. Impressively, the most potent and selective one, namely, 77, remarkably inhibited AKT/ERK1/2 and FAK/SRC pathways and thereby reversed Taxol resistance as well as inhibited both migration and invasion of A549/Taxol cells. Collectively, this study not only displayed the role of CYP1B1 in Taxol resistance and cancer migration/invasion but also helped unlock the CYP1B1-oriented anticancer discovery.

Mdivi-1 induces spindle abnormalities and augments taxol cytotoxicity in MDA-MB-231 cells

Taxol is a first-line chemotherapeutic for numerous cancers, including the highly refractory triple-negative breast cancer (TNBC). However, it is often associated with toxic side effects and chemoresistance in breast cancer patients, which greatly limits the clinical utility of the drug. Hence, compounds that act in concert with taxol to promote cytotoxicity may be useful to improve the efficacy of taxol-based chemotherapy. In this study, we demonstrated that mdivi-1, a putative inhibitor of mitochondrial fission protein Drp1, enhances the anticancer effects of taxol and overcomes taxol resistance in a TNBC cell line (MDA-MB-231). Not only did mdivi-1 induce mitotic spindle abnormalities and mitotic arrest when used alone, but it also enhanced taxol-induced antimitotic effects when applied in combination. In addition, mdivi-1 induced pronounced spindle abnormalities and cytotoxicity in a taxol-resistant cell line, indicating that it can overcome taxol resistance. Notably, the antimitotic effects of mdivi-1 were not accompanied by prominent morphological or functional alterations in mitochondria and were Drp1-independent. Instead, mdivi-1 exhibited affinity to tubulin at μM level, inhibited tubulin polymerization, and immediately disrupted spindle assembly when cells entered mitosis. Together, our results show that mdivi-1 associates with tubulin and impedes tubulin polymerization, actions which may underlie its antimitotic activity and its ability to enhance taxol cytotoxicity and overcome taxol resistance in MDA-MB-231 cells. Furthermore, our data imply a possibility that mdivi-1 could be useful to improve the therapeutic efficacy of taxol in breast cancer.

TLR4/IL-6/IRF1 signaling regulates androgen receptor expression: A potential therapeutic target to overcome taxol resistance in ovarian cancer

Ovarian cancer is poorly treatable due, at least in part, to induced drug resistance to taxol- and cisplatin-based chemotherapy. Recent studies showed that ectopic overexpression of toll-like receptor 4 (TLR4) in ovarian cancer cells leads to upregulation of the androgen receptor (AR) and transactivation of taxol resistance genes, thereby causing chemoresistance. In the present study, we examined the signaling pathways involving TLR4 and interleukin 6 (IL-6) that enhance AR expression. Based on transcriptomic analysis, we show that IL-6 functions as a hub gene among the upregulated genes in taxol-treated TLR4-overexpressing ovarian cancer cells. Both the TLR4 activator taxol and IL-6 can induce AKT phosphorylation, whereas TLR4 knockdown or inhibition of the IL-6 signal transducer GP130 abrogates AKT activation. Furthermore, expression of AR and IL-6 is downregulated in TLR4-knockdown, taxol-resistant cells. In addition, TLR4 knockdown inhibits GP130 and IL-6 receptor alpha (IL6Rα) activities, indicating that TLR4 plays a critical role in IL-6 signaling. On the other hand, nuclear translocation of AR is induced by IL-6 treatment, whereas knockdown of endogenous IL-6 reduces AR and TLR4 expression in taxol-resistant ovarian cancer cells. These results indicate that TLR4 and IL-6 play a crucial role in AR gene regulation and function. We also identify interferon regulatory factor 1 (IRF1) as a downstream target of IL-6 signaling and as a regulator of AR expression. Moreover, analysis of clinical samples indicates that high IL-6 expression correlates with poor progression-free survival in ovarian cancer patients treated with taxol. Overall, our findings indicate that the TLR4/IL-6/IRF1 signaling axis represents a potential therapeutic target to overcome AR-based taxol resistance in ovarian cancer.

The nature of the modification at position 37 of tRNAPhe correlates with acquired taxol resistance.

Acquired drug resistance is a major obstacle in cancer therapy. Recent studies revealed that reprogramming of tRNA modifications modulates cancer survival in response to chemotherapy. However, dynamic changes in tRNA modification were not elucidated. In this study, comparative analysis of the human cancer cell lines and their taxol resistant strains based on tRNA mapping was performed by using UHPLC–MS/MS. It was observed for the first time in all three cell lines that 4-demethylwyosine (imG-14) substitutes for hydroxywybutosine (OHyW) due to tRNA-wybutosine synthesizing enzyme-2 (TYW2) downregulation and becomes the predominant modification at the 37th position of tRNAphe in the taxol-resistant strains. Further analysis indicated that the increase in imG-14 levels is caused by downregulation of TYW2. The time courses of the increase in imG-14 and downregulation of TYW2 are consistent with each other as well as consistent with the time course of the development of taxol-resistance. Knockdown of TYW2 in HeLa cells caused both an accumulation of imG-14 and reduction in taxol potency. Taken together, low expression of TYW2 enzyme promotes the cancer survival and resistance to taxol therapy, implying a novel mechanism for taxol resistance. Reduction of imG-14 deposition offers an underlying rationale to overcome taxol resistance in cancer chemotherapy.

Upregulation of miR-129-5p increases the sensitivity to Taxol through inhibiting HMGB1-mediated cell autophagy in breast cancer MCF-7 cells.

Although Taxol has improved the survival of cancer patients as a first-line chemotherapeutic agent, an increasing number of patients develop resistance to Taxol after prolonged treatment. The potential mechanisms of cancer cell resistance to Taxol are not completely clear. It has been reported that microRNAs (miRNAs) are involved in regulating the sensitivity of cancer cells to various chemotherapeutic agents. In this study, we aimed to explore the role of miR-129-5p in regulating the sensitivity of breast cancer cells to Taxol. Cell apoptosis and autophagy, and the sensitivity of MCF-7 cells to Taxol were assessed with a series of in vitro assays. Our results showed that the inhibition of autophagy increased the Taxol-induced apoptosis and the sensitivity of MCF-7 cells to Taxol. Up-regulation of miR-129-5p also inhibited autophagy and induced apoptosis. Furthermore, miR-129-5p overexpression increased the sensitivity of MCF-7 cells to Taxol. High mobility group box 1 (HMGB1), a target gene of miR-129-5p and a regulator of autophagy, was negatively regulated by miR-129-5p. We found that interference of HMGB1 enhanced the chemosensitivity of Taxol by inhibiting autophagy and inducing apoptosis in MCF-7 cells. Taken together, our findings suggested that miR-129-5p increased the chemosensitivity of MCF-7 cells to Taxol through suppressing autophagy and enhancing apoptosis by inhibiting HMGB1. Using miR-129-5p/HMGB1/autophagy-based therapeutic strategies may be a potential treatment for overcoming Taxol resistance in breast cancer.

Highlights of This Issue

The rarity of RET oncofusions in lung cancer impairs the pace of clinical studies in this subtype. Huang and colleagues established a transgenic KIF5B–RET fusion lung cancer mouse model that can develop invasive lung adenocarcinoma with fibrotic desmoplasia. The authors also identified RETV804L gatekeeper and vandetanib-resistant RETG810A mutations in cultured cells with increasing concentrations of RET inhibitors. Ponatinib was identified as the most potent RET inhibitor, displayed hyperactivity to RETG810A, and caused lung tumor regression in the animals. These studies established preclinical models that can rapidly evaluate efficacy of RET inhibitors, discover resistance mechanisms, and identify approaches to thwart resistance.Therapeutically targeting intracellular kinases is biologically attractive but fundamentally challenging. Interpreting drug behavior in cellular and animal models is confounded by redundant enzymatic activities, common substrates, and cell-specific signaling networks. To better understand clinical characteristics, cyclin-dependent kinase drugs (abemaciclib, dinaciclib, palbociclib, ribociclib) were profiled using cellular (untransformed, tumorigenic), biophysical, and structural approaches. CDK–drug co-crystal structures coupled with biochemical analyses reveal a unique hinge architecture that enables kinome selectivity and the extent of CDK family engagement is derived from other ATP binding pocket interactions. Together, these findings provide insight into drug pharmacology and a framework for rational drug design.A liposome-based platform for the co-delivery of the 3rd generation P-gp inhibitor tariquidar and paclitaxel was developed to overcome Taxol resistance in ovarian cancer cells. Using several functional assays and imaging approaches, Zhang, Sriraman, and colleagues showed that the dual drug-loaded liposomes inhibited cell viability, blocked proliferation, and caused G2–M arrest in vitro. In a xenograft model of Taxol resistance ovarian cancer, the liposomes significantly impeded tumor growth through the induction of apoptosis and reduction of proliferation. This novel nanomedicine has potential for clinical testing in ovarian cancer patients with P-gp–overexpressing tumors.The expression of small heterodimer partner (SHP, NROB2) is downregulated in metastatic HCC and is negatively associated with patient survival in several types of aggressive and highly metastatic human cancers. In this study, Yang and colleagues performed small molecule microarrays and discovered a novel activator of SHP, namely DSHN. DSHN transcriptionally activated Shp mRNA, but also stabilized SHP protein by preventing its ubiquitination and degradation. In vivo and in vitro evidence indicated that DSHN inhibited HCC metastasis via SHP-mediated repression of Ccl2 signaling. This study highlights the potential use of SHP as a therapeutically tractable gene in preclinical and translational studies.

MicroRNA-16 sensitizes breast cancer cells to paclitaxel through suppression of IKBKB expression.

Paclitaxel (Taxol) is an effective chemotherapeutic agent for treating breast cancer patients. However, chemoresistance is a major obstacle in cancer treatment. Here, we showed that overexpression of miR-16 promoted Taxol-induced cytotoxicity and apoptosis in breast cancer cells. Furthermore, IκB kinase β (IKBKB) was identified as a direct target of miR-16. Up-regulation of IKBKB suppressed Taxol-induced apoptosis and led to an increased resistance to Taxol, and restoring IKBKB expression in miR-16-overexpressing breast cancer cells recovered Taxol resistance. Moreover, miR-16 was highly expressed in Taxol-sensitive breast cancer tissues compared with Taxol-resistant tissues, and there was an inverse correlation between miR-16 expression and IKBKB expression in breast cancer tissues. The expression levels of miR-16 were negatively associated with T stages, whereas the expression of IKBKB was positively correlated with T stages, lymph node metastasis and clinical stages. Taken together, our data demonstrates that miR-16 sensitizes breast cancer cells to Taxol through the suppression of IKBKB expression, and targeting miR-16/IKBKB axis will be a promising strategy for overcoming Taxol resistance in breast cancer.

MP55-01 CYTOPLASMIC FOXO1 NEGATIVELY REGULATES ERK TO OVERCOME TAXOL RESISTANCE IN PROSTATE CANCER

You have accessJournal of UrologyProstate Cancer: Basic Research III1 Apr 2015MP55-01 CYTOPLASMIC FOXO1 NEGATIVELY REGULATES ERK TO OVERCOME TAXOL RESISTANCE IN PROSTATE CANCER Chunwu Pan, Jian An, Yunqian Pan, Liguo Wang, Jun Zhang, Jun Qi, and Haojie Huang Chunwu PanChunwu Pan More articles by this author , Jian AnJian An More articles by this author , Yunqian PanYunqian Pan More articles by this author , Liguo WangLiguo Wang More articles by this author , Jun ZhangJun Zhang More articles by this author , Jun QiJun Qi More articles by this author , and Haojie HuangHaojie Huang More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.2044AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES O-class forkhead factor 1 (FoxO1) acts as a tumor suppressor in the nucleus through its transcriptional function. FOXO1 can be phosphorylated by activated protein kinase B (AKT) and exported to cytoplasm, thereby losing its tumor suppressor function. The PI3K/AKT signaling is frequently hyperactivated in human cancers, and inhibitors of PI3K/AKT are currently tested in various preclinical and clinical settings. Intriguingly, inhibition of PI3K/AKT invariably triggers activation of extracellular signal-regulated kinase (ERK) signaling, which is an important mechanism of chemoresistance. Here we identify a transcription-independent tumor suppressor function of FOXO1 in the cytoplasm, finding of which has significant applications in overcoming taxol chemoresistance through inhibiting ERK signaling in prostate cancer. METHODS We employed unbiased tandem affinity purification (TAP) and mass spectrometry to identify interacting proteins of FOXO1. We used GST-Pulldown and co-immuniprecipitation (co-IP) to define the interaction domains in both IQGAP1 and FOXO1, Western-blot to detect the protein expression level, immunocytochemistry (ICC) to detect FOXO1 cellular localization, immunohistochemistry (IHC) to analysize FOXO1 and phosphor-ERK level in human prostate cancer using tissue microarray (TMA), and MTS assay to determine cells proliferation. RESULTS We identified the scaffold protein IQGAP1 as a binding partner of FOXO1 (Fig A). We demonstrated that serine-319 phosphorylation on FOXO1 is important for FOXO1 binding to IQGAP1 (Fig B). Cytoplasmic FOXO1 inhibits IQGAP1-augmented phosphorylation of ERK1/2 by blocking IQGAP1 interaction with RAF, MEK and ERK proteins (Fig C-E). We also found that there is an inverse correlation between FOXO1 and phosphorylated ERK1/2 in human prostate cancer tissues (Fig F, G). We further demonstrated that low expression of FOXO1 and increased expression of ERK1/2 phosphorylation significantly associates with prostate cancer progression (Fig H). Finaly, we demonstrated that cytoplasmic FOXO1 inhibits PI3K/AKT inhibitor or taxol-induced ERK activation and thereby overcoming taxol-resistance in prostate cancer (Fig I-L). CONCLUSIONS Cytoplasmic FOXO1 overcomes taxol resistance in prostate cancer by negatively regulating ERK activation. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e672-e673 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Chunwu Pan More articles by this author Jian An More articles by this author Yunqian Pan More articles by this author Liguo Wang More articles by this author Jun Zhang More articles by this author Jun Qi More articles by this author Haojie Huang More articles by this author Expand All Advertisement Advertisement PDF DownloadLoading ...

MP55-02 PROSTATE ADENOCARCINOMAS COMPARED WITH NON-TUMOR PROSTATE TISSUE. EXPRESSION OF COLLAGEN, ELASTIC SYSTEM FIBERS, SMOOTH MUSCLE CELLS AND VESSELS, ACCORDING TO THE GLEASON SCORE

METHODS: We employed unbiased tandem affinity purification (TAP) and mass spectrometry to identify interacting proteins of FOXO1. We used GST-Pulldown and co-immuniprecipitation (co-IP) to define the interaction domains in both IQGAP1 and FOXO1, Western-blot to detect the protein expression level, immunocytochemistry (ICC) to detect FOXO1 cellular localization, immunohistochemistry (IHC) to analysize FOXO1 and phosphor-ERK level in human prostate cancer using tissue microarray (TMA), and MTS assay to determine cells proliferation. RESULTS: We identified the scaffold protein IQGAP1 as a binding partner of FOXO1 (Fig A). We demonstrated that serine-319 phosphorylation on FOXO1 is important for FOXO1 binding to IQGAP1 (Fig B). Cytoplasmic FOXO1 inhibits IQGAP1-augmented phosphorylation of ERK1/2 by blocking IQGAP1 interaction with RAF, MEK and ERK proteins (Fig C-E). We also found that there is an inverse correlation between FOXO1 and phosphorylated ERK1/2 in human prostate cancer tissues (Fig F, G). We further demonstrated that low expression of FOXO1 and increased expression of ERK1/2 phosphorylation significantly associates with prostate cancer progression (Fig H). Finaly, we demonstrated that cytoplasmic FOXO1 inhibits PI3K/AKT inhibitor or taxol-induced ERK activation and thereby overcoming taxol-resistance in prostate cancer (Fig I-L). CONCLUSIONS: Cytoplasmic FOXO1 overcomes taxol resistance in prostate cancer by negatively regulating ERK activation.

Synergistic cytotoxicity of cisplatin and Taxol in overcoming Taxol resistance through the inhibition of LDHA in oral squamous cell carcinoma.

The development of chemoresistance in patients represents a major challenge in cancer treatment. Lactate dehydrogenase-A (LDHA) is one of the principle isoforms of LDH that is expressed in breast tissue, controlling the conversion of pyruvate to lactate and also playing a significant role in the metabolism of glucose. The aim of this study was to identify whether LDHA was involved in oral cancer cell resistance to Taxol and whether the downregulation of LDHA, as a result of cisplatin treatment, may overcome Taxol resistance in human oral squamous cells. The OECM-1 oral epidermal carcinoma cell line was used, which has been widely used as a model of oral cancer in previous studies. The role of LDHA in Taxol and cisplatin resistance were investigated and the synergistic cytotoxicity of cisplatin and/or Taxol in oral squamous cells was analyzed. Cell viability was analyzed by MTT assay, LDHA expression was analyzed by western blot analysis and siRNA tranfection was performed to knock down LDHA expression. The present study results showed that decreased levels of LDHA were responsible for the resistance of oral cancer cells to cisplatin (CDDP). CDDP treatments downregulated LDHA expression, and lower levels of LDHA were detected in the CDDP-resistant oral cancer cells compared with the CDDP-sensitive cells. By contrast, the Taxol-resistant cancer cells showed elevated LDHA expression levels. In addition, small interfering RNA-knockdown of LDHA sensitized the cells to Taxol, but desensitized them to CDDP treatment, while exogenous expression of LDHA sensitized the cells to CDDP, but desensitized them to Taxol. The present study also revealed the synergistic cytotoxicity of CDDP and Taxol for killing oral cancer cells through the inhibition of LDHA. This study highlights LDHA as a novel therapeutic target for overcoming Taxol resistance in oral cancer patients using the combined treatments of Taxol and CDDP.

Silencing of glutaminase 1 resensitizes Taxol-resistant breast cancer cells to Taxol

Taxol is a front‑line chemotherapeutic agent for the treatment of patients with multiple types of tumor. However, resistance to Taxol remains one of the principal causes of cancer‑associated mortality. Glutamine, which is metabolized via a glutaminase (GLS)‑dependent process, termed glutaminolysis, is important in cell growth and metabolism. The present study reported a novel mechanism underlying Taxol resistance in breast cancer cells. By investigating the glutamine metabolism of breast cancer cells in response to treatment with Taxol invitro, it was observed that Taxol induced the uptake of glutamine and the expression of GLS1. Notably, Taxol‑resistant cancer cells exhibited upregulation in the metabolism of glutamine and expression of GLS1. In addition, overexpression of GLS1 rendered cancer cells resistant to Taxol, indicating that GLS1 may be the therapeutic target for overcoming Taxol resistance in clinical therapeutics. The results also demonstrated that knock‑down of GLS1 using small interfering RNA, resensitized the Taxol‑resistant breast cancer cells to Taxol.

Inhibition of IL-6/STAT3 axis and targeting Axl and Tyro3 receptor tyrosine kinases by apigenin circumvent taxol resistance in ovarian cancer cells.

Ovarian cancer is the number one cause of death from gynaecological malignancy. Platinum-based and taxol-based chemotherapy has been used as a standard therapy, but intrinsic and acquired resistance to chemotherapy is a major obstacle to treat the disease. In the present study, we found that in the chemoresistant ovarian cancer SKOV3/TR cells, interleukin-6 (IL-6), IL-6 receptor and signal transducers and activators of transcription 3 (STAT3) expression as well as STAT3 phosphorylation were upregulated compared to those in parental cells. Silencing of IL-6 using IL-6 siRNA was found to suppress IL-6 production, STAT3 and phosphoSTAT3 levels, which eventually reduced proliferation and clonogenicity of taxol-resistant SKOV3/TR cells. In addition, stattic, a STAT3 inhibitor, was found to result in decrease of cell viability and clonogenicity of these cells, indicating that the elevated IL-6 and STAT3, phosphoSTAT3 levels are associated with the development of taxol resistance. Next, we found anti-proliferative effect of apigenin on both SKOV3 and SKOV3/TR cells. RT-PCR and western blot results showed that apigenin significantly reduced the expression of Axl and Tyro3 receptor tyrosine kinases (RTKs) at mRNA and protein level, which account for its cytotoxic activity. We further found that apigenin decreased Akt phosphorylation and the level of B-cell lymphoma-extra large (Bcl-xl or BCL2-like 1 isoform 1), an inhibitor of apoptosis. On the contrary to these results, apigenin had no effect on IL-6 production, STAT3 and phosphoSTAT3 protein levels, suggesting that apigenin exerts its anti-proliferative activity via downregulation of Axl and Tyro3 expression, Akt phosphorylation and Bcl-xl expression, but not modulation of IL-6/STAT3 axis. Taken together, our data suggest that inhibition of IL-6/STAT3 signaling pathway and downregulation of Axl and Tyro3 RTKs expression might be a therapeutic strategy to overcome taxol resistance in ovarian cancer cells.

Combination of Taxol® and dichloroacetate results in synergistically inhibitory effects on Taxol-resistant oral cancer cells under hypoxia

Cancer cells preferentially catalyze glucose through the glycolytic pathway in the presence of adequate oxygen. This phenomenon is known as the Warburg effect. As is the case with numerous cancer therapeutic agents, resistance remains a significant problem when using Taxol® to treat malignancies. The present study reported that expression of pyruvate dehydrogenase kinase 1 (PDK1) was induced by Taxol treatment at low toxic concentrations in oral cancer cells. In addition, Taxol‑resistant cells exhibited upregulated PDK1 protein and mRNA expression. Elevated PDK1 levels contribute to Taxol resistance under hypoxic conditions. Inhibition of PDK1 expression was observed when oral cancer cells were treated with the PDK1 inhibitor dichloroacetate (DCA). The combination of Taxol with DCA showed synergistic inhibitory effects on Taxol‑resistant cells under hypoxic conditions; these effects were not observed in Taxol‑sensitive oral cancer cells under normoxic conditions. The present study provides a novel mechanism for overcoming Taxol resistance in oral cancer cells, and will contribute towards the development of clinical therapeutics for cancer patients.

RASSF1A and the Taxol Response in Ovarian Cancer

The RASSF1A tumor suppressor gene is frequently inactivated by promoter methylation in human tumors. The RASSF1A protein forms an endogenous complex with tubulin and promotes the stabilization of microtubules. Loss of RASSF1A expression sensitizes cells to microtubule destabilizing stimuli. We have observed a strong correlation between the loss of RASSF1A expression and the development of Taxol resistance in primary ovarian cancer samples. Thus, we sought to determine if RASSF1A levels could dictate the response to Taxol and whether an epigenetic therapy approach might be able to reverse the Taxol resistant phenotype of RASSF1A negative ovarian tumor cells. We found that knocking down RASSF1A expression in an ovarian cancer cell line inhibited Taxol-mediated apoptosis and promoted cell survival during Taxol treatment. Moreover, using a combination of small molecule inhibitors of DNA Methyl Transferase enzymes, we were able restore RASSF1A expression and Taxol sensitivity. This identifies a role for RASSF1A in modulating the tumor response to Taxol and provides proof of principal for the use of epigenetic therapy to overcome Taxol resistance.

MicroRNA-125b Confers the Resistance of Breast Cancer Cells to Paclitaxel through Suppression of Pro-apoptotic Bcl-2 Antagonist Killer 1 (Bak1) Expression

Paclitaxel (Taxol) is an effective chemotherapeutic agent for treatment of cancer patients. Despite impressive initial clinical responses, the majority of patients eventually develop some degree of resistance to Taxol-based therapy. The mechanisms underlying cancer cells resistance to Taxol are not fully understood. MicroRNA (miRNA) has emerged to play important roles in tumorigenesis and drug resistance. However, the interaction between the development of Taxol resistance and miRNA has not been previously explored. In this study we utilized a miRNA array to compare the differentially expressed miRNAs in Taxol-resistant and their Taxol-sensitive parental cells. We verified that miR-125b, miR-221, miR-222, and miR-923 were up-regulated in Taxol-resistant cancer cells by real-time PCR. We further investigated the role and mechanisms of miR-125b in Taxol resistance. We found that miR-125b was up-regulated in Taxol-resistant cells, causing a marked inhibition of Taxol-induced cytotoxicity and apoptosis and a subsequent increase in the resistance to Taxol in cancer cells. Moreover, we demonstrated that the pro-apoptotic Bcl-2 antagonist killer 1 (Bak1) is a direct target of miR-125b. Down-regulation of Bak1 suppressed Taxol-induced apoptosis and led to an increased resistance to Taxol. Restoring Bak1 expression by either miR-125b inhibitor or re-expression of Bak1 in miR-125b-overexpressing cells recovered Taxol sensitivity, overcoming miR-125-mediated Taxol resistance. Taken together, our data strongly support a central role for miR-125b in conferring Taxol resistance through the suppression of Bak1 expression. This finding has important implications in the development of targeted therapeutics for overcoming Taxol resistance in a number of different tumor histologies.

Abstract 2109: Mir-125b confers the resistance of cancer cells to Taxol through suppression of Bak1

Abstract Paclitaxel (Taxol) is one of the most effective chemotherapeutic agents for treatment of cancer patients. Despite impressive initial clinical responses, the majority of patients will eventually develop some degree of resistance to Taxol-based therapy. The mechanisms underlying cancer cells resistance to Taxol are not fully understood. microRNA (miRNA) has emerged to play important roles in tumorigenesis and drug-resistance. However, the interaction between the development of Taxol resistance and miRNA has not been previously explored. In this study, we utilized a miRNA array to screen differentially expressed miRNAs, comparing Taxol-resistant and their Taxol-sensitive parental cells, and investigated the role and mechanisms of miR-125b in Taxol resistance of breast cancer cells. We found that miR-125b was up-regulated in Taxol-resistant cells, with its overexpression causing a marked inhibition of Taxol-induced cytotoxicity and apoptosis, and a subsequent increase in the resistance of cancer cells to Taxol. Moreover, we demonstrated that Bak1 is a direct target of miR-125b in breast cancer cells, and down- regulation of Bak1 suppressed Taxol- induced apoptosis and led to an increased resistance to Taxol. Furthermore, restoring the expression of Bak1 by either miR-125b inhibitor or specific siRNA recovered Taxol sensitivity, overcoming miR-125-mediated Taxol resistance. Taken together, our data strongly supports a central role for miR-125b in conferring Taxol resistance through the suppression of Bak1 expression. This finding has important implications in the development of targeted therapeutics for overcoming Taxol resistance in a number of different tumor histologies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2109.

Abstract 23: Sanguinarine suppresses survivin expression and inhibits the growth of prostate cancer cells

Abstract Background: Prostate cancer is a frequently occurring disease and is the second leading cause of cancer related deaths of men in the United States. Current treatments have proved inadequate in curing or controlling prostate cancer and a search for agents for the management of this disease is urgently needed. Survivin plays an important role in both progression of castration-resistant prostate cancer and resistance to chemotherapy. Altered expression of survivin in prostate cancer cells is associated with cancer progression, drug/radiation resistance, poor prognosis and short patient survival. Methods: We performed a cell based rapid screen of the Prestwick Chemical Library consisting of 1120 FDA-approved compounds with known safety and bioavailability in humans to identify potential inhibitors of survivin and anti-cancer agents for prostate cancer. Results: Sanguinarine, a benzophenanthridine alkaloid derived primarily from the bloodroot plant, was identified as a novel inhibitor of survivin that selectively kills prostate cancer cells over “normal” prostate epithelial cells. We found that sanguinarine inhibits survivin protein expression through protein degradation via the ubiquitin-proteasome system. Sanguinarine induces apoptosis and inhibits tumor formation and growth of human prostate cancer cells. Administration of sanguinarine, beginning 3 days post ectopic implantation of DU145 human prostate cancer cells, reduces both tumor weight and volume by more than 70% compared to the control treated mice with no apparent toxicity. In addition, sanguinarine sensitized prostate cancer cells to paclitaxel-mediated growth inhibition and apoptosis, offering a potential therapeutic strategy for overcoming taxol resistance. Conclusions: These results suggest that sanguinarine may be developed as an agent either alone or in combination with taxol for treatment of castration-resistant prostate cancer overexpressing survivin. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 23.