Sensitivity Of Drug-resistant Cells Research Articles

The regulation of hsacirc_004413 promotes proliferation and drug resistance of gastric cancer cells by acting as a competing endogenous RNA for miR-145-5p.

BackgroundWhether circRAN, which acts as a microRNA sponge, plays a role in 5-fluorouracil (5-Fu) resistant gastric cancer has not been reported. In this study, a 5-Fu resistant cell line with an IC50 of 16.59 µM was constructed.MethodsUsing comparative analysis of circRNA in the transcriptomics of resistant and sensitive strains, 31 differentially expressed circRNAs were detected, and the microRNA interacting with them was predicted.ResultsHsacirc_004413 was selected for verification in drug resistant and sensitive cells. By interfering with hsacirc_004413 using antisense RNA, the sensitivity of drug resistant cells to 5-Fu was significantly promoted, and the apoptosis and necrosis of the cells were significantly increased. In sensitive cells, inhibition by inhibitors enhanced the resistance of cells to 5-Fu. We hypothesize that hsacirc_004413 makes gastric cancer cells resistant to 5-Fu mainly through adsorption of miR-145-5p.

Exosomes derived from chronic myeloid leukemia cells: roles in disease progression, survival, and treatment

Exosomes, biologically active extracellular vesicles, are derived from normal and neoplastic cells. Emerging evidence revealed that exosomes modulate cell-cell communication and involve in hemostatic and pathologic processes. Recent studies have shown that exosomes released from cancer cells such as chronic myeloid leukemia cells could act as a key mediator in tumor induction and progression. Myeloid cells-derived exosomes affect different processes including angiogenesis, neoplastic proliferation, tumor cell survival, and imatinib resistance. These exosomes induce angiogenesis and tumor progression by IL-8 overexpression in both leukemic and bone marrow stromal cells. Exosomes cargo could alter the expression of different adhesion molecules, anti-and pro-apoptotic molecules, cytokines, and chemokines such as VCAM-1, ICAM-1, BCL, BAD, BAX, TGF-β, TNF-α, CXCL12 which affect tumor migration, homing, survival, and growth. CML-derived exosomes can also regulate signal transduction pathways, such as ERK/MAPK, ERK/Akt, EGFR/Ras, and Wnt. Furthermore, they can be applied as a vehicle for drug delivery or sensitization of drug-resistant cells. Here, we reviewed the role of chronic myeloid cell-derived exosomes in tumor growth, survival, and resistance to treatment. Keywords: Chronic myeloid leukemia, Exosome, Angiogenesis, Tyrosine kinase inhibitor, Drug resistance

Aspirin attenuates YAP and β-catenin expression by promoting β-TrCP to overcome docetaxel and vinorelbine resistance in triple-negative breast cancer.

e13099 Background: YAP and β-catenin play critical roles in the carcinogenesis of several cancers, but their functions in chemoresistance remain unclear in triple-negative breast cancer (TNBC). Herein, we examined the expression and function of the two proteins in the chemoresistance of TNBC. Methods: We examined the expression levels of YAP and β-catenin in TNBC samples by immunohistochemistry (IHC) and their roles in prognosis. Dynamic changes in the two proteins in TNBC tissues were determined by RT-PCR. CCK-8 assays, colony formation assays, flow cytometry, migration assays and nude mouse models were used to investigate the functions of YAP and β-catenin in drug-resistant TNBC cells. The involved signalling pathways were detected by western blot assays. Results: TNBC patients with high YAP and/or β-catenin expression had a higher risk of relapse or mortality than patients with low YAP and/or β-catenin expression, and changes in the expression of the two proteins might be a promising predictive biomarker for neoadjuvant chemotherapy sensitivity. Inhibition of YAP or β-catenin enhanced the cytotoxicity of the anti-microtubule agents docetaxel and vinorelbine against TNBC cells in vitro and in vivo. Moreover, YAP and β-catenin were upregulated in docetaxel- or vinorelbine-resistant TNBC cells, while inhibition of YAP or β-catenin enhanced the sensitivity of drug-resistant cells. Interestingly, aspirin, a kind of acetylsalicylic acid, reversed the drug resistance of cells and in combination with aspirin and docetaxel or vinorelbine significantly inhibited the growth of drug-resistant TNBC cells. The involved mechanism is that aspirin impaired YAP or β-catenin expression by upregulating the E3 ubiquitin ligase β-TrCP. Conclusions: Both YAP and β-catenin act as key biomarkers of prognosis and anti-microtubule drug resistance in TNBC, and the combined use of aspirin and anti-microtubule drugs inhibits YAP and β-catenin, presenting several promising therapeutic approaches for TNBC treatment.

Low-Dose Crizotinib, a Tyrosine Kinase Inhibitor, Highly and Specifically Sensitizes P-Glycoprotein-Overexpressing Chemoresistant Cancer Cells Through Induction of Late Apoptosis in vivo and in vitro.

We investigated possible conditions or drugs that could target P-glycoprotein (P-gp)-overexpressing drug-resistant KBV20C cancer cells. Specifically, we focused on identifying a single treatment with a relatively low half maximal inhibitory concentration (IC50). Our approach utilized repurposing drugs, which are already used in clinical practice. We evaluated 13 TKIs (gefitinib, imatinib, erlotinib, nilotinib, pazopanib, masatinib, sunitinib, sorafenib, regorafenib, lapatinib, vandetanib, cediranib, and crizotinib) for their sensitizing effects on P-gp-overexpressing drug-resistant KBV20C cells. We found that crizotinib had a much greater sensitization effect than the other tested drugs at relatively low doses. In a detailed quantitative analysis using both lower doses and time-duration treatments, we demonstrated that crizotinib, which increased the levels of apoptosis and G2 arrest, was the best TKI to induce sensitization in P-gp-overexpressing KBV20C cells. Upon comparing resistant KBV20C cells and sensitive KB parent cells, crizotinib was found to markedly sensitize drug-resistant KBV20C cancer cells compared with other TKIs. This suggests that crizotinib is a resistant cancer cell-sensitizing drug that induces apoptosis. In mice bearing xenografted P-gp-overexpressing KBV20C cells, we confirmed that crizotinib significantly reduced tumor growth and weight, without apparent side effects. In addition, although lapatinib and crizotinib have a high P-gp inhibitory activity, we found that co-treatment with crizotinib and vincristine (VIC) did not have much of a sensitization effect on KBV20C cells, whereas lapatinib had a high sensitization effect on VIC-treated KBV20C cells. This suggests that crizotinib is a single-treatment specific drug for resistant cancer cells. These findings provide valuable information regarding the sensitization of drug-resistant cells and indicate that low-dose crizotinib monotherapy may be used in patients with specific P-gp-overexpressing chemoresistant cancer.

Upregulation of miR-34c after silencing E2F transcription factor 1 inhibits paclitaxel combined with cisplatin resistance in gastric cancer cells

BACKGROUNDMicroRNA 34c (miR-34c) has been reported to be associated with malignant types of cancer, however, it remains unknown whether miR-34c is involved in chemoresistance in gastric cancer (GC).AIMTo investigate the effect of miR-34c and its upstream transcription factor E2F1 on paclitaxel combined with cisplatin resistance in GC cells.METHODSPaired GC tissues and adjacent normal tissues were randomly sampled from 74 GC patients. miR-34c and E2F1 were detected by real-time quantitative PCR (qPCR) and Western blot. In addition, the drug resistance of GC cells to paclitaxel and cisplatin was induced by concentration gradient increasing methods, and changes in miR-34c and E2F1 during this process were measured. Furthermore, E2F1 and miR-34c overexpression or underexpression vectors were constructed and transfected into drug-resistant GC cells. MTT was employed to test the sensitivity of cells to paclitaxel combined with cisplatin, qPCR was adopted to detect the expression of miR-34c, Western blot was applied to detect the expression levels of E2F1, drug resistance-related proteins and apoptosis-related proteins, and flow cytometry was used for the determination of cell apoptosis and cell cycle status.RESULTSE2F1 was overexpressed while miR-34c was underexpressed in GC. After inducing GC cells to be resistant to paclitaxel and cisplatin, E2F1 expression increased while miR-34c expression decreased. Both silencing E2F1 and over-expressing miR-34c could increase the sensitivity of drug-resistant GC cells to paclitaxel combined with cisplatin, promote cell apoptosis and inhibit cell proliferation. Among which, silencing E2F1 could reduce the expression of drug resistance-related proteins and apoptosis-related proteins, while over-expression of miR-34c could upregulate the expression of apoptosis-related proteins without affecting the expression of MDR-1, MRP and other drug resistance-related proteins. Rescue experiments demonstrated that inhibiting miR-34c could significantly weaken the sensitization of drug resistant cells, and Si E2F1 to paclitaxel combined with cisplatin.CONCLUSIONE2F1 inhibits miR-34c to promote the proliferation of GC cells and enhance the resistance to paclitaxel combined with cisplatin, and silencing E2F1 is conducive to improving the efficacy of paclitaxel combined with cisplatin in GC cells.

Histamine Receptor Antagonists, Loratadine and Azelastine, Sensitize P-gp-overexpressing Antimitotic Drug-resistant KBV20C Cells Through Different Molecular Mechanisms.

Previously, we showed that KBV20C cancer cells highly resistant to antimitotic drugs were sensitized by co-treatment with a repositioned drug fluphenazine. Considering that fluphenazine plays a role as a histamine receptor antagonist, we investigated low doses of 21 other histamine receptor antagonists (lidocaine, cimetidine, chlorpromazine, diphenhydramine, promethazine, ranitidine, famotidine, clemastine, chlorpheniramine, desloratadine, loratadine, cyproheptadine, azelastine, brompheniramine, carbinoxamine, fexofenadine, hydroxyzine, levocetirizine, meclizine, nizatidine, and pemirolast) to identify repositioned drugs for their sensitizing effects on antimitotic drug-resistant KBV20C cells at relatively low doses. Co-treatment with loratadine, and with azelastine highly sensitized KBV20C cells to vincristine treatment. Loratadine and azelastine reduced cell viability, increased G2 arrest, and up-regulated apoptosis when co-administered with vincristine. In detailed quantitative analysis, combination of vincristine with loratadine had a higher sensitization effect than that with azelastine. Azelastine had a higher P-glycoprotein (P-gp)-inhibitory activity, similar to that of verapamil, indicating that sensitization by vincristine-azelastine involved the P-gp-inhibitory effects of azelastine. However, loratadine had a very low P-gp-inhibitory activity, suggesting that loratadine sensitization to vincristine is independent of the P-gp-inhibition. Co-treatment with eribulin and loratadine increased the sensitization of KBV20C cells, suggesting that loratadine can be combined with other antimitotic drugs to sensitize resistant cancer cells. These findings provide important information regarding the sensitization of drug-resistant cells and indicate that loratadine may be used in patients with potentially resistant cancer without any toxic effects from P-gp inhibition.

Long non-coding RNA FAM84B-AS promotes resistance of gastric cancer to platinum drugs through inhibition of FAM84B expression

PurposeTo investigate the expression and significance of the long non-coding RNA (lncRNA) FAM84B-AS in gastric cancer tissues and its effect on platinum drug resistance in gastric cancer. Methods(1) The expression of FAM84B-AS was detected in samples from 228 cases of fresh gastric cancer to analyze its association with clinical data and gastric cancer prognosis. (2) An lncRNA interference cell line model was established and used to study the effects of FAM84B-AS on the malignant biological behaviors of gastric cancer cells and platinum drug resistance at the cellular level. (3) The mechanisms underlying the effect of FAM84B-AS on gastric cancer were investigated using Western blotting. Results(1) FAM84B-AS was closely associated with the differentiation level, T stage, and N stage of gastric cancer and could be used as an independent risk factor of the prognosis of gastric cancer. (2) FAM84B-AS interference significantly inhibited the proliferation and invasion abilities of gastric cancer cells and significantly increased the percentage of apoptosis. FAM84B-AS could partially restore the sensitivity of drug resistant cells to platinum drugs. The above results were also confirmed in in vivo studies. (3) Western blot results demonstrated that FAM84B-AS interference could activate apoptosis signaling pathways in gastric cancer cells to promote apoptosis in gastric cells. ConclusionFAM84B-AS could be used as a molecular marker of the malignancy of gastric cancer and offers a certain predictive function for gastric cancer prognosis. FAM84B-AS promotes gastric cancer proliferation through inhibition of apoptosis signaling pathways and promotes drug resistance of gastric cancer cells to platinum drugs through its apoptosis-inhibiting functions.

Sequential therapy with redox-responsive glucolipid nanocarrier separately delivering siRNA and doxorubicin to overcome multidrug resistance

P-glycoprotein (P-gp) is a major efflux transporter overexpressed on multidrug resistant tumor cells and responsible for pumping drugs out. If anti-tumor drugs are given when P-gp level is low, satisfactory treatment efficiency may be achieved. Thus, a P-gp down-regulating siRNA (siMDR1) and doxorubicin (DOX) were applied to eliminate multidrug resistant breast cancer cells (MCF-7/ADR). A redox-responsive glucolipid conjugate (CSO-ss-SA) was used to condense siRNA (CSO-ss-SA/siRNA) and encapsulate DOX (CSO-ss-SA/DOX) separately. They responded to the high reducing environment of tumor cells and fast released the payload. CSO-ss-SA/siMDR1 silenced MDR1 gene and resulted in a transient decrease of P-gp. Sequentially, DOX formulation (CSO-ss-SA/DOX or DOX·HCl) was delivered when P-gp was reduced to the lowest level. After pretreatment by CSO-ss-SA/siMDR1, cytotoxicity of CSO-ss-SA/DOX and DOX·HCl against MCF-7/ADR cells were 6.4 or 3.4-fold respectively of that treated by DOX·HCl alone, which exhibited increased cytotoxin sensitivity of drug resistant cells and maximized therapeutic outcomes. These results revealed that the sequential treatment strategy of CSO-ss-SA/siRNA and CSO-ss-SA/DOX hold potential in achieving an optimal overcoming multidrug resistance efficiency.

Effect of Y6, an epigallocatechin gallate derivative, on reversing doxorubicin drug resistance in human hepatocellular carcinoma cells.

Cancer cells can acquire resistance to a wide variety of diverse and unrelated drugs, this phenomenon is termed multidrug resistance (MDR). Multidrug resistance has been an obstacle to the success of cancer chemotherapy. The present study investigated the reversal effect of Y6, a new compound obtained by chemically modifying the structure of epigallocatechin-3-gallate (EGCG) extracted from green tea. Y6 was proven to be effective in inhibiting cell proliferation and reversing drug resistance in doxorubicin (DOX) resistant human hepatocellular carcinoma cells (BEL-7404/DOX). BEL-7404/DOX cells were treated with either doxorubicin combination regimen (doxorubicin plus Y6 or epigallocatechin-3-gallate or verapamil separately) or doxorubicin alone. The results showed that cell proliferation was inhibited and late cell apoptosis increased in the combination treatment group, especially in the group treated with doxorubicin plus Y6. Further analysis revealed that the expressions of hypoxia-inducible factor-1α and multidrug resistance 1/P-glycoprotein decreased at both messenger RNA and protein levels by treatments with combined drugs compared to doxorubicin alone. Our results indicated that Y6, as a drug resistance reversal agent, increased the sensitivity of drug resistant cells to doxorubicin. The mechanisms of actions of Y6 in reversal effect were associated with the decreased expression of hypoxia-inducible factor-1α and multidrug resistance 1/P-glycoprotein.

Co-treatment with Celecoxib or NS398 Strongly Sensitizes Resistant Cancer Cells to Antimitotic Drugs Independent of P-gp Inhibition.

Inhibition of cyclooxygenase-2 (COX-2) has been investigated in clinical trials. Currently, NS398 and celecoxib are the most commonly used COX-2 inhibitors. The purpose of this study was to identify conditions that would increase the sensitivity of resistant cancer cells to antimitotic drugs. We tested whether COX-2 inhibitors can sensitize drug-resistant KBV20C cancer cells. We also compared the efficacy of NS398 with that of celecoxib. Both NS398 and celecoxib could sensitize KB and KBV20C cells to a similar extent, suggesting that COX-2 inhibitors could be used for sensitive, as well as resistant, cancer cells. We demonstrated that the NS398 and celecoxib sensitization mechanism is independent of the inhibition of p-glycoprotein (P-gp), suggesting that resistant KBV20C cells are sensitized through targeting of signaling pathways by both drugs. Furthermore, through using microscopic observation, assessment of cleaved poly ADP ribose polymerase (C-PARP) and annexin V staining we determined that both COX-2 inhibitors strongly sensitized resistant KBV20C cells to vinblastine (VIB) or paclitaxel (PAC) treatment. These results suggest that antimitotic drug-resistant cancer cells can be strongly sensitized by co-treatment with COX-2 inhibitors, without P-gp inhibitory activity. These findings provide important information regarding the sensitization of drug-resistant cells and indicate that COX-2 inhibitors may be used for potentially resistant cancer patients, without the toxic effects of P-gp inhibition.

Abstract 2896: Nanoparticle co-delivery of RNAi and chemotherapy for the treatment of drug-resistant cancers

Abstract The emergence of drug resistance has been a critical barrier for effective cancer chemotherapy. Recently, RNA interference (RNAi) has shown promise in suppressing the over-expression of drug resistance-associated proteins such as P-glycoprotein and Prohibitin1, and in treating multidrug resistant cancers through combination with chemotherapy. Nevertheless, the safe and effective delivery of RNAi therapeutics (e.g., siRNA and miRNA) to target cells remains a major hurdle for their clinical applications, and the co-delivery of RNAi and chemotherapeutic agents in a controlled fashion requires specific delivery vehicles. Herein, we present a novel nanoparticle (NP) platform to tackle the challenges associated with the delivery of siRNA and chemotherapeutic drugs, using biodegradable and biocompatible polymers and lipids. The hybrid lipid-polymer NP has a differentially charged hollow core/shell nanostructure which provides the delivery system with three distinct functional features: (i) a positively charged inner hollow core for dense loading of siRNA; (ii) a middle hydrophobic polymer layer for the encapsulation of anti-cancer drugs (e.g., taxanes) and for the controlled release of both siRNA and drug; and (iii) a relatively neutral lipid-polyethylene glycol (PEG) surface to keep the NP stable and prolong its systematic circulation. Moreover, such hybrid NPs can be formulated in a simple and robust way that could facilitate future scale-up. By screening various factors, such as inner cationic lipids, outer PEG chain length, and formulation parameters, the hybrid lipid-polymer NPs show excellent in vitro knockdown efficacy at low doses of siRNA, with greater than 80% luciferase silencing at an siRNA dose of 0.4 pmol (4.0 nM) and ∼ 95% silencing at a dose of 2.0 pmol (20 nM). These NPs can also efficiently reduce the expression of drug resistance-associated proteins (e.g., MDR1 and Prohibitin1), as evidenced by qPCR. Cellular cytotoxicity experiments further confirm that this NP co-delivery strategy can drastically improve the sensitivity of drug-resistant cells to chemotherapy. In addition, these hybrid NPs demonstrate promising in vivo results for co-delivering siRNA and chemotherapeutic drugs to treat drug-resistant cancers. 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 2896. doi:1538-7445.AM2012-2896

Paclitaxel sensitization of multidrug-resistant cells to chemotherapy is independent of the cell cycle

Recent studies have shown that paclitaxel (Taxol) is an active chemotherapeutic in the treatment of small cell lung cancer. Paclitaxel binds to tubulin and prevents depolymerization. This causes cells to arrest in the G(2)/M phase of the cell cycle, resulting in sensitization of cells to drug or radiation treatment. A drug-resistant H69 small cell lung cancer subline was established. Cytotoxicity of cisplatin and chlorambucil was determined using the MTT cell viability assay and distribution of DNA in the cell cycle. DNA distribution was analyzed by flow cytometry after treatment with paclitaxel or the other tubulin-binding drugs, vinblastine and navelbine. The H69-EPR drug-resistant subline was resistant to epirubicin (sixfold) and was cross-resistant to cisplatin (7.5-fold) and chlorambucil (7.5-fold). Pretreatment with paclitaxel or vinblastine, but not navelbine, sensitized the subline to cisplatin and chlorambucil (P < 0.05), with no effect on parental H69 cells. Sensitization was dose dependent and occurred at doses below those that caused a G(2)/M block in the cell cycle. Sensitization of drug-resistant cells by paclitaxel was not associated with its ability to cause a G(2)/M block in the cell cycle. Sensitization by paclitaxel and vinblastine, but not navelbine, which preferentially targets mitotic tubulin, suggests that sensitization may involve changes in the tubulin-dependent intracellular transport processes rather than changes in mitotic tubulin and the G(2)/M block.

Patterns of anthracycline retention modulation in human tumor cells.

Laser excitation of cellular doxorubicin and daunomycin content (with or without incubation in the presence of efflux blockers such as phenothiazines or verapamil) was studied in cells from leukemic peripheral blood, bone marrow aspirates, and ascites and pleural fluid of solid tumor patients. Selected examples are presented to show that heterogeneity in cellular anthracycline retention as well as sensitivity to efflux modulators is seen in human tumor cells. Several tumor subpopulations differing in their cellular retention of anthracyclines or sensitivity to modulators were seen. In serial tumor samples from patients (pre- and post-treatment with anthracycline-containing protocols), initial drug retention and sensitivity to efflux modulators was followed by lack of drug retention and insensitivity to modulators. The present study shows that in view of the variables encountered in drug-retention characteristics and sensitivity to efflux modulators, one needs to screen tumor samples before recommending use of any particular transport modulator to enhance drug retention and sensitivity of drug-resistant cells.

Verapamil and adriamycin in the treatment of drug-resistant ovarian cancer patients.

Eight patients with refractory ovarian cancer were treated on a pilot protocol of verapamil plus Adriamycin (Adria Laboratories, Columbus, OH). This trial was based on our previous laboratory studies which demonstrated that Adriamycin resistance in human ovarian cancer cell lines could be partially reversed by exposure of the cells to high concentrations of verapamil (3,000 ng/mL). Patients were treated in an intensive care unit with continuous cardiovascular monitoring. The dose of verapamil was escalated in each patient until hypotension or heart block developed, and this dose was maintained for 72 hours. Adriamycin (50 mg/m2) was infused over 24 hours during the second day of the verapamil infusion and verapamil alone was administered on the third day in an effort to block efflux from drug-resistant cells. This intensive approach led to a median plasma verapamil level of 1,273 ng/mL (range, 720 to 2,767). However, the high infusion rates of verapamil (9 micrograms/kg/min) required to achieve these plasma levels produced an unacceptable degree of cardiac toxicity. Two patients developed transient atropine-responsive complete heart block and four patients developed transient congestive heart failure with increases in pulmonary capillary wedge pressure. There was no evidence that the noncardiac toxicities of Adriamycin were enhanced by verapamil. There were no objective responses to therapy. Future studies should use less cardiotoxic calcium channel blockers that can be safely administered to produce the plasma levels required for in vitro sensitization of drug resistant cells.