Lung Resistance-related Protein Expression Research Articles

Grape seed proanthocyanidin extract reverses multidrug resistance in HL-60/ADR cells via inhibition of the PI3K/Akt signaling pathway

Background and purposeMultidrug resistance (MDR) is a great challenge and obstacle in cancer treatment. It is a common problem in the treatment of acute myeloid leukemia (AML). Whether grape seed proanthocyanidin extract (GSPE) could reverse MDR in patients with AML is still unknown. The aim of this study was to investigate the MDR reverse ability of GSPE and its possible mechanism in vitro. Materials and methodsHuman leukemia cell line HL-60 cells and HL-ADR cells were used. MTT assay were employed to identify the cytotoxic effects of different chemotherapeutic drugs and reverse ability of GSPE. Flow cytometry assays were used to verify the cell apoptosis induced by GSPE. MDR-related genes expression was tested by real-time polymerase chain reaction (Q-PCR). MDR-related protein expression was assessed by Western blotting assays. The genes and their related protein expression of multidrug resistance-associated protein 1 (MRP1), multidrug resistance protein 1 (MDR1) and lung resistance-related protein (LRP) were tested in this study. Key resultsWe found that HL-60/ADR cells were resistant to a variety of chemotherapeutic drugs, including cytarabine (Ara-C), adriamycin (ADR), vincristine (VCR), daunorubicin (DNR), mitoxantrone (MTZ), pirarubicin (THP), homoharringtonine (HHT) and etoposide (VP16). Co-treatment with GSPE could significant lower the IC50 of Ara-C and ADR in HL-60/ADR cells (P < 0.01). MDR related mRNA and their protein expression of MRP1 and MDR1 were significant highly expressed in HL-60/ADR cells than HL-60 cells (P < 0.01). But only protein expression of LRP was higher in HL-60/ADR cells than HL-60 cells (P < 0.05). GSPE could induce a higher intracellular level of ADR in HL-60/ADR cells. It could also inhibit Akt phosphorylation resulted in the down regulation of MRP1, MDR1 and LRP and induce cell apoptosis. 25.0 μg/mL GSPE significant inhibited the Akt phosphorylation (P < 0.05). Conclusion and implicationsGSPE-reversed MDR of HL-60/ADR cells might be associated with the inhibition of the PI3K/Akt signaling pathway, which resulted in the down-regulation the expression of MRP1, MDR1 and LRP. These results provide that GSPE may serve as a combination therapy in AML chemotherapy for future study.

Lung resistance-related protein (LRP) predicts favorable therapeutic outcome in Acute Myeloid Leukemia

There is conflicting evidence that MDR1, MRP2 and LRP expression is responsible for chemotherapy resistance. We conducted this study to explore their role in AML therapy outcomes. Bone marrow and peripheral blood samples of 90 AML patients, receiving chemotherapy, were analyzed by real time PCR. Gene expression was calculated by the 2−ΔΔCt method. The patients who had a persistent remission were labelled ‘Good Responder’ (GRes) whereas, those with relapse or drug resistance were labelled ‘Poor Responders’ (PRes). Higher LRP expression in bone marrow, but not in peripheral blood, was positively associated with persistent remission (p = 0.001), GRes (p = 0.002), 1-year overall as well as disease-free survival (p = 0.02 and p = 0.007, respectively). Marrow and blood MDR1 and MRP2 expression did not differ significantly between the above groups. Logistic regression analysis showed that only a diagnosis of acute promyelocytic leukemia (APL; M3) or high marrow LRP expression significantly predicted a favorable therapeutic outcome. This is the first report showing that high bone marrow LRP expression predicts significant favorable therapeutic outcome. Peripheral blood LRP expression as well as marrow and blood MDR1 and MRP2 expression have no predictive value in AML patients treated with standard dose cytarabine and daunorubicin 3+7 regimen.

Expression of LRP Gene in Breast Cancer Patients Correlated with MRP1 as Two Independent Predictive Biomarkers in Breast Cancer

Background:Breast cancer is the most common malignancy in women. Multidrug resistance (MDR) is still a great obstacle of breast cancer chemotherapy. We have previously shown that multidrug resistance-associated protein 1 (MRP1) is associated with response to neoadjuvant chemotherapy. The lung resistance-related protein (LRP) is identified as a prognostic marker and response to treatment factor which has been studied mainly in hematological malignancy and leukemia. In this study, we aimed to analyze LRP expression and possible correlation between the expression level of this gene with MRP1 as a candidate marker for chemotherapy resistance.Materials and Methods:We collected 54 breast tumors and adjacent normal tissues from Iranian breast cancer patients and Real time RT-PCR was employed to measure the gene expression level in our samples.Results:MRP1 and LRP expression level were significantly lower in tumor tissues of the patients responding to chemotherapy compared to non-responding patients. No relation between the expression level of either of these genes and clinicopathology markers was found.Conclusion:Our results suggest that LRP gene expression is correlated to MRP1 in human breast cancer cells and may affect the clinical response to treatment.

Characterization of Drug Resistance Protein Expression Levels in Human Melanoma Cells and Subsequent Treatment with Antifungal Agents

The multidrug resistance (MDR) phenotype allows cancer cells to display decreased sensitivity to chemotherapy. A well‐characterized drug resistance gene in cancer is MDR1, which encodes for the p‐glycoprotein/ATP‐binding cassette (ABC) transporter. Lesser known are the roles of the multidrug resistance‐associated protein 1 (MRP1) and lung resistance‐related protein (LRP) in melanoma. Because of the previously demonstrated presence of MRP1 and LRP in human melanoma cells and the continued difficulty of overcoming MDR in melanoma treatment, here we characterized the expression levels of drug resistance genes in melanoma cell lines derived from human patients and determined their susceptibility to novel antifungal treatment. Expression levels were determined by real‐time PCR (qPCR) and immunoblot analyses. Susceptibility of cell lines to antifungal agents was analyzed by flow cytometry, cell proliferation assays, and immunoblotting after drug treatments.Several lines of human melanoma and normal skin cells were analyzed. MDR1 expression was observed in multiple melanoma lines, while increased levels of MRP1 were observed in normal skin cells and one melanoma line. Interestingly, LRP expression was highest in normal skin cells and several melanoma lines. Novel treatment with clotrimazole, an agent previously shown to eradicate melanoma cells, caused significant decreases in proliferation and increases in cell death in all MDR melanoma lines. No extensive cell death was induced by clotrimazole treatment in noncancerous human skin cells. These results indicated that clotrimazole selectively induces cytotoxicity in drug‐resistant melanoma lines. However, decreased efficacy was observed in melanoma lines that overexpressed LRP, as compared to all other melanoma lines that overexpressed MDR1 and/or MRP1. These results indicated that LRP, mostly known for facilitating drug resistance in lung and breast cancer, may also have a significant role in the ability of melanoma cells to resist the effects of conventional melanoma treatments.In conclusion, this study demonstrated the existence and differential expression levels of MDR1, MRP1, and LRP in melanoma and normal skin cell lines. Although all melanoma lines were susceptible to treatment with clotrimazole, decreased efficacy was observed in melanoma lines that overexpressed LRP. Taken together, we conclude that clotrimazole has in vitro efficacy toward the treatment of drug‐resistant human melanoma cells. However, these preliminary studies also indicated that LRP is potentially a significant facilitator of drug resistance in human melanoma.Support or Funding InformationThis research was funded by the Ohio Northern University Summer Research Stipend.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Kinesin KIF4A is associated with chemotherapeutic drug resistance by regulating intracellular trafficking of lung resistance-related protein.

Multidrug resistance (MDR) is the major impediment to cancer chemotherapy. The expression of lung resistance-related protein (LRP), a non-ATP-binding cassette (ABC) transporter, is high in tumor cells, resulting in their resistance to a variety of cytotoxic drugs. However, the function of LRP in tumor drug resistance is not yet explicit. Our previous studies had shown that Kinesin KIF4A was overexpressed in cisplatin (DDP)-resistant human lung adenocarcinoma cells (A549/DDP cells) compared with A549 cells. The expression of KIF4A in A549 or A549/DDP cells significantly affects cisplatin resistance but the detailed mechanisms remain unclear. Here, we performed co-immunoprecipitation experiments to show that the tail domain of KIF4A interacted with the N-terminal of LRP. Immunofluorescence images showed that both the ability of binding to LRP and the motility of KIF4A were essential for the dispersed cytoplasm distribution of LRP. Altogether, our results shed light on a potential mechanism in that motor protein KIF4A promotes drug resistance of lung adenocarcinoma cells through transporting LRP-based vaults along microtubules towards the cell membrane. Thus KIF4A might be a cisplatin resistance-associated protein and serves as a potential target for chemotherapeutic drug resistance in lung cancer.

Synergistic Effect of Baicalin and Adriamycin in Resistant HL-60/ADM Leukaemia Cells

Background/Aims: The present study was designed to investigate the expression of multidrug resistance (MDR)-related genes, verify the synergistic effects of baicalin and Adriamycin (ADM) and investigate the related mechanisms in ADM-resistant leukaemic HL-60/ADM cells. Methods: We used a HL-60/ADM cell line. Cytotoxicity and flow cytometry assays were employed to verify the cytotoxic effects of baicalin. Real-time polymerase chain reaction and Western blotting assays were used to assess the expression of MDR-related genes and the changes in gene expression (both MDR-related and PI3K/Akt pathway-related) induced by administration of baicalin. Results: We found that only multidrug resistance protein 1 (MRP1), lung resistance-related protein (LRP) and Bcl-2 genes were expressed in both HL-60 and HL-60/ADM cells. HL-60/ADM cells exhibited significantly higher expression (p < 0.05). We also observed that low-dose baicalin (5 and 10 µmol/L) can induce growth inhibition and apoptotic effects on HL-60/ADM cells by increasing the intracellular accumulation of ADM. The synergistic effect of baicalin and ADM was verified. Concerning the potential mechanisms involved in this process, we showed that baicalin down-regulated the expression of several MDR-related and PI3K/Akt pathway-related genes. Conclusions: We confirmed the increased expression of MRP1, LRP and Bcl-2 genes in HL-60/ADM cells compared to regular HL-60 cells, which are recommended for future investigation on MDR. The present study provided evidence of the synergistic effect of baicalin and ADM in HL-60/ADM cells. Therefore, baicalin may be considered as a potential therapeutic agent against resistant leukaemia. Suppression of the PI3K/Akt signalling pathway, followed by inhibition of the expression of MDR-related genes may be a common mechanism in combination treatments with ADM for the reduction of resistance to ADM.

ABCB1, ABCC1, and LRP gene expressions are altered by LDL, HDL, and serum deprivation in a human doxorubicin-resistant uterine sarcoma cell line

Multidrug resistance (MDR) is the major cause of cancer treatment failure. The ATP-binding cassette-B1 (ABCB1) transporter, also known as MDR1 or P-glycoprotein, is thought to promote the efflux of drugs from cells. MDR is also associated with the multidrug resistance-associated protein 1 (ABCC1) and the lung resistance-related protein (LRP), a human major vault protein. Moreover, MDR has a complex relationship with lipids. The ABCB1 has been reported to modulate cellular cholesterol homeostasis. Conversely, cholesterol has been reported to modulate multidrug transporters. However, results reported to date are contradictory and confusing. The aim of this study was to investigate whether LDL, HDL, and serum deprivation could influence ABCB1, ABCC1, and LRP expression in a human doxorubicin-resistant uterine sarcoma cell line.ABCB1 and ABCC1 expression increased after 24 h of serum deprivation, and expression returned to basal levels after 72 h. LDL, depending on concentration, increased ABCB1, ABCC1, and LRP expression. ABCB1 expression increased at low HDL, and decreased at high HDL concentrations.We demonstrated that serum deprivation and lipoproteins, particularly LDL, modulated ABCB1 expression and, to a lesser extent, ABCC1 expression. This finding may link the phenomena of drug transport, cholesterol metabolism and cancer.

Expression of ERCC1, RRM1 and LRP in non-small cell lung cancers and their influence on chemotherapeutic efficacy of gemcitabine concomitant with nedaplatin.

To explore the clinical efficacy of gemcitabine concomitant with nedaplatin and drug resistance in the treatment of non-small cell lung cancer (NSCLC) and associated molecular predicators. A total of 68 patients diagnosed with NSCLC by histology served as the study objects and were randomly divided into an observation group treated with gemcitabine concomitant with nedaplatin and a control group with cisplatin concomitant with gemcitabine, 34 cases for each group. Short-term and long-term efficacies, adverse responses as well as the expression of nucleotide excision repair cross complementing 1 (ERCC1), ribonucleotide reductase subunit M1 (RRM1) and lung resistance-related protein (LRP) in NSCLC tissues in both groups were assessed. The short-term objective response rate (ORR) and disease control rate (DCR) were 35.3% (12/34) and 76.5% (26/34) in the observation group and 38.2% (13/34) and 85.3% (29/34) in the control group, respectively, the differences not being statistically significant. The time to progression (TTP) in both groups were 1~12 months, while the median TTP was 135 d and 144 d, respectively. Though the survival was slightly higher in the control group, there were no significant differences in TTP and survival time. The rates of decreased hemoglobin, vomiting and nausea as well as renal toxicity were evidently lower in the observation group, while other adverse responses demonstrated no significant difference. The positive expression rates of ERCC1, RRM1 and LRP were 47.1% (16/34), 61.8% (21/34) and 64.7% (22/34) in the observation group, respectively. Compared with negative ERCC1 expression, ORR had decreasing trend and the overall survival time (OS) decreased significantly in patients with positive ERCC1 expression, which were markedly decreased by the positive expressions of RRM1 and LRP. Gemcitabine concomitant with nedaplatin has significant effects in the treatment of NSCLC, with an adverse response rate obviously lower than for cisplatin concomitant with gemcitabine, suggesting that wider use in the clinic is warranted. Additionally, the positive expressions of ERCC1, RRM1 and LRP may increase patient drug resistance, so they can be applied as the chemotherapeutic predicators to guide individualized therapy of NSCLC patients.

Lung Resistance-Related Protein (LRP) Expression in Malignant Ascitic Cells as a Prognostic Marker for Advanced Ovarian Serous Carcinoma

Ovarian serous carcinoma is an aggressive cancer that often presents with metastatic disease. Although primary tumor and established metastatic foci in the omentum are generally compared to identify proteins involved in drug resistance, we investigated a potential bridge, the malignant cells from ascites, as facilitator of drug resistance and recurrence. We evaluated the expression of drug resistance markers P-glycoprotein (P-gp), canalicular multispecific organic anion transporter (MRP2), and lung resistance-related protein (LRP) in malignant cells from ascites and matched omental metastasis from 25 patients with advanced-stage ovarian serous carcinoma who were chemotherapeutic naïve and undergoing initial cytoreductive surgery. Cell viability in vitro, patient response to chemotherapy, and patient survival were correlated with these biomarkers. Of the 25 patients evaluated for a correlation of LRP to 1-year recurrence, we correctly predicted the 1-year recurrence of 24 patients based solely on the presence of LRP in ascitic tumor cells (p=0.01). P-gp and MRP2 were not expressed in malignant cells of ascites or omental metastases. Malignant cells from ascites had higher expression of LRP and were found to be more resistant to carboplatin treatment than cells from omental metastasis (p=0.00375) by in vitro assay. LRP expression in the malignant cells of ascites correlated with carboplatin resistance (p=0.001) by in vitro assay and recurrence at 1 year (p=0.0125). LRP expression in malignant cells of ascites is a promising marker to predict response to first-line chemotherapy in patients with advanced ovarian serous carcinoma.

Multidrug Resistance in Primary Tumors and Metastases in Patients with Esophageal Squamous Cell Carcinoma

Studies have demonstrated that radical esophagectomy can significantly prolong disease-free survival and improve the survival rate of patients with T3 or T4 esophageal cancer and lymph node metastasis. Multidrug resistant cancer cells have active efflux mechanisms that prevent the accumulation of chemotherapeutic drugs in the cells. The purpose of this study was to compare the expression of five MDR related proteins between primary tumors in patients with thoracic esophageal squamous cell carcinoma (ESCC) and metastatic cancer in lymph nodes to explore the clinical significance of heterogeneity in MDR metastatic cancer cells. Fifty-four patients with ESCC and lymph node metastasis were included. All patients underwent subtotal esophagectomy and D2/D3 lymph node resection. The expression of lung resistance-related protein (LRP), P-glycoprotein, topoisomerase-II, thymidylate synthase, and glutathione S-transferase P1-1 (GST-π) were determined in the primary tumors and lymph nodes via immunohistochemistry. The expression of LRP was significantly different between the primary tumors and lymph nodes (P = 0.026). No significant differences were found for the other four proteins, and protein expression was not associated with either degree of differentiation or disease stage. It was also found that GST-π was expressed in all patients in both the primary tumors and lymph nodes, suggesting that the design and application of chemotherapeutic protocols capable of reducing GST-π expression may be beneficial for patients with ESCC. Additional research regarding the clinical utility of MDR protein expression in ESCC is warranted to design effective chemotherapeutic protocols.

Effect of Src tyrosine kinase inhibition on the drug-resistance as well as MDR1 and LRP expression of the human cis-platinum-resistant lung cancer cell line A549/DDP

背景与目的本研究旨在探讨抑制Src酪氨酸激酶活性对人肺癌A549/DDP细胞耐药性及多药耐药蛋白（muti-drug resistance 1, MDR1）和肺耐药相关蛋白（lung resistance-related protein, LRP）表达的影响。方法以Src酪氨酸激酶抑制剂作用于A549/DDP细胞，应用Western blot法检测肿瘤细胞Src酪氨酸激活性的变化，CellTiter-Glo发光法检测肿瘤细胞药物敏感性的变化，流式细胞仪检测肿瘤细胞Rh-123含量变化，Western blot法和RT-PCR检测肿瘤细胞MDR1和LRP表达变化。结果Src酪氨酸激酶抑制剂可下调A549/DDP细胞中Src酪氨酸激活性，2.5 μM和10 μM Src酪氨酸激酶抑制剂作用肿瘤细胞后，肿瘤细胞药物敏感性提高，逆转倍数（reversal fold, RF）分别为1.59倍和2.10倍，肿瘤细胞中Rh-123含量分别提高了1.21倍和1.59倍，MDR1 mRNA表达分别是对照组的53.8%和27.5%，LRP mRNA表达分别是对照组的59.3%和21.4%，MDR1和LRP蛋白表达水平明显下降。结论抑制A549/DDP细胞中Src酪氨酸激酶活性可逆转肿瘤细胞多药耐药性，提高肿瘤细胞药物敏感性，其机制可能与降低细胞MDR1和LRP表达有关。

Reversal Effect of Melanoma Differentiation Associated Gene‐7/Interleukin‐24 on Multidrug Resistance in Human Hepatocellular Carcinoma Cells

Multidrug resistance is the main cause for failure of chemotherapy. Melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) has been implicated in the inhibition of human tumor cell proliferation. However, the reversing effect of mda-7/IL-24 on multidrug resistance of human hepatocellular carcinoma (HCC) is not fully clear. In this study, we investigated the effects of overexpression of the mda-7/IL-24 gene in human HCC. We established mda-7/IL-24 overexpressing BEL-7402/5-fluorouracil (5-FU) cell lines and their drug sensitivity to 5-FU and doxorubicin (DOX) which were investigated by MTT. Furthermore, we investigated the apoptotic rate and the intracellular accumulation of Rhodamine-123 and DOX by flow cytometry. We also studied the expression of multidrug resistance gene 1 (MDR1), lung resistance-related protein (LRP), and multidrug resistance-related protein 1 (MRP1) by real-time polymerase chain reaction and Western blotting. Transcriptional activation of AP-1 and NF-κB was determined by luciferase reporter assay. The drug sensitivity of 5-FU or DOX, the apoptotic rate, and the intracellular accumulation of Rhodamine-123 and DOX were increased, while the mRNA and protein expression levels of MDR1, LRP, and MRP1 were reduced. The transcriptional activation of AP-1 and NF-κB was suppressed in mda-7/IL-24 overexpressing BEL-7402/5-FU cells. Our results demonstrated that mda-7/IL-24 could restore the drug sensitivity through the downregulation of MDR1, MRP1, and LRP expression, as well as the transcriptional activation of AP-1 and NF-κB and effectively reverse MDR.

P-0075 18F-FDG Uptake in Hepatocellular Carcinoma Cell Lines: Correlation with Multidrug Resistance Proteins Expression

IntroductionMalignant cells require high energy via glycolytic generation of ATP for cell proliferation and survival. Positron emission tomography (PET) using radiolabelled glucose analogue 18F-FDG allows detecting increased glycolytic events in tumor cells. Hepatocellular carcinoma (HCC) is highly resistant to chemotherapy, which is due in part to the overexpression of multidrug resistance proteins (MDR). It is known that 18F-FDG uptake in HCC is associated with tumor differentiation and with the expression of these proteins. This study aims to evaluate the uptake and retention of 18F-FDG into two human HCC cell lines with different expression levels of p53 and to correlate with the expression of three MDR proteins: the P-Glycoprotein (Pgp), the multidrug resistance–associated protein (MRP1) and the lung resistance-related protein (LRP). MethodsThe human HCC cell lines used were HepG2 (wp53) and Huh7 (mp53). Cell suspensions were incubated with 2x106 cells per ml with 18F-FDG (25 µCi/mL). Samples of 200 µl were collected to Eppendorf tubes at different periods of time, which were centrifuged in order to separating the supernatant from the pellet. Radioactivity of pellet and supernatant was measured in a well counter. The retention of 18F-FDG was obtained incubating cell suspension with the radiopharmaceutical for 60 minutes. Then, the cells were centrifuged and the medium replaced, the following procedure was similar to the uptake studies. The protein levels of Pgp, MRP1 and LRP were determined by flow cytometry. ResultsIt was noted that the HuH7 cell line, that express a mutant form of p53, has the higher levels of 18F-FDG uptake and retention. The HepG2 cell line has a lower uptake and retention, and a higher expression of MRP1. The levels of Pgp and LRP expression are identical for two cell lines. ConclusionAlthough these are preliminary results it can be concluded that there is an inverse relationship between the expression of MRP1 and the levels of uptake and retention of 18F-FDG. These data may be useful in clinical practice

Association of Nuclear YB-1 Localization With Lung Resistance-related Protein and Epidermal Growth Factor Receptor Expression in Lung Cancer

BackgroundY-box binding protein 1 (YB-1) is an oncogenic transcription factor that is activated in response to various genotoxic stresses. The purpose of this study was to elucidate whether YB-1 correlates with the expression of lung resistance–related protein (LRP) and epidermal growth factor receptor (EGFR) in primary lung cancer. Patients and MethodsOne hundred and five non–small-cell lung cancer (NSCLC) specimens were analyzed by immunohistochemistry. Knockdown of YB-1 messenger RNA by small interfering RNA(siRNA) was tested for the lung cancer cell lines A549 and Calu-3. ResultsNuclear YB-1 expression significantly correlated with positive LRP and EGFR expression (P < .001). Nuclear YB-1 expression and positive LRP and EGFR expression were independent adverse prognostic factors in patients with NSCLC. Furthermore, patients with tumors positive for nuclear YB-1 and LRP had a significantly worse prognosis than those negative for nuclear YB-1 and LRP (P < .001). In addition, patients with tumors positive for nuclear YB-1 and EGFR had a significantly worse prognosis than those negative for nuclear YB-1 and EGFR (P < .001). In in vitro analyses that use the NSCLC cell lines A549 and Calu-3, the downregulation of YB-1 with siRNAs drastically decreased the expression of EGFR. However, downregulation of YB-1 remarkably decreased the expression of LRP in A549 cells; however, a slight decrease in LRP was induced by the downregulation of YB-1 in Calu-3 cells. ConclusionOur data demonstrate that nuclear YB-1 localization is associated with LRP and EGFR expression in NSCLC, and nuclear YB-1 localization and LRP and EGFR expression are of prognostic significance in NSCLC.

P53 suppresses lung resistance-related protein expression through Y-box binding protein 1 in the MCF-7 breast tumor cell line

Lung resistance-related protein (LRP) has roles in multi-drug resistance of tumor cells. Understanding the mechanisms that regulate LRP expression in tumor cells is an important research area. A putative p53 response element in the LRP promoter has been found. Thus, p53-related regulation of LRP expression was explored in this study. We first demonstrated that p53 overexpression inhibited LRP expression both at the protein and mRNA levels. Then, using a dual-luciferase reporter assay, we located the p53 response element to the Y-box (-263~-407) of the LRP promoter, the YB-1 binding site, but not the putative p53 response element. Furthermore, coimmunoprecipitation and chromatin immunoprecipitation showed p53 could bind to the Y-box of the LRP promoter through interaction of p53 with YB-1. YB-1 coexpression with p53 facilitated p53-induced suppression of endogenous LRP expression in MCF-7 cells. HDAC2, a corepressor of p53, was found to also interact with YB-1, and this interaction was mediated by p53. These results showed that the p53-HDAC2 transcriptional repressor complex can bind to the Y-box of the LRP promoter and repress LRP expression through interaction with YB-1. p53-related suppression of LRP expression was completely reversed by doxorubicin treatment and Adr, whereas CP and VP-16 treatment induced LRP expression increased significantly. Inhibition of LRP expression by siRNA facilitated Adr induced apoptosis of MCF-7 cells. All these findings indicated that loss of p53-related suppression of LRP may be the reason for LRP expression increase, and, therefore, chemotherapy resistance in tumor cells.

Multidrug resistance-associated biomarkers PGP, GST-π, Topo-II and LRP as prognostic factors in primary ovarian carcinoma

This study aims to investigate the expression of P-glycoprotein (PGP), glutathione S-transferase π (GST-π), DNA topoisomerase II (Topo-II) and lung resistance-related protein (LRP) in ovarian carcinoma, thus providing better chemotherapy choice and post-operative prognosis for ovarian carcinoma patients. A total of 80 primary ovarian carcinoma, 16 benign ovarian epithelial neoplasm, and 12 normal ovarian tissue samples were collected. Immunohistochemistry was used to detect the expression of PGP, GST-π, Topo-II and LRP, and the results were analysed by correlation with clinicopathological parameters. Positive expression rates of PGP, GST-π, Topo-II and LRP in patients with ovarian carcinoma (57.5%, 58.8%, 76.3% and 73.8%, respectively) were all higher than those found in normal and benign tissue (P<0.05). In clinical stages I/II vs. III/IV, the expression rates of PGP, GST-π, Topo-II and LRP were 40.7% vs. 66% (P<0.05), 40.7% vs. 67.9% (P<0.05), 66.7% vs. 81.1% (P>0.05) and 55.6% vs. 83.0% (P<0.05), respectively. Carcinoma differentiation ranged from well to poor, and expression levels of each marker were as follows: PGP, 57.9%, 62.1% and 53.1% (P>0.05); GST-π, 36.8%, 55.2% and 75.0% (P<0.05); Topo-II, 52.6%, 79.3% and 87.5% (P<0.05); and LRP, 84.2%, 69.0% and 71.9% (P>0.05). Ovarian carcinoma patients with PGP-, GST-π-, Topo-II- and LRP-positive expression had a shorter median survival time than those who were negative for these markers (PGP: 36 months vs. 48 months [P=0.0017]; GST-π: 36 months vs. 41 months [P=0.0103]; Topo-II: 37 months vs. 39 months [P=0.3811]; LRP: 37 months vs. 55 months [P=0.002]). COX regression analysis demonstrated that the clinical stage of the tumour, and the expression of PGP, GST-π or LRP, may influence patient survival time after surgery. The relative death risk for patients with clinical stage III/IV tumours increased 9.46-fold compared to those with stage I/II tumours. The relative death risk in the PGP-, GST-π- and LRP-positive groups increased by 2.049-, 2.452- or 2.609-fold, respectively, compared with the corresponding negative groups. PGP, GST-π, Topo-II and LRP are all expressed in primary ovarian carcinoma, indicating the presence of multidrug resistance in this disease. Combined evaluation of PGP, GST-π, Topo-II and LRP expression may enable better chemotherapeutic choice and provide an accurate prognosis for ovarian carcinoma patients.

MDR modulation in colorectal adenocarcinoma cell lines: kinetic studies using 99mTc-sestamibi

Multidrug resistance (MDR) represents one of the major setbacks to chemotherapy and can occur due to the overexpression of efflux pumps as P-glycoprotein (Pgp), multiple resistance-related protein 1 (MRP1) and lung resistance-related protein (LRP). Verapamil, a substrate for Pgp, modulates its activity. L-buthionine-sulfoximine (BSO) can be used as blocker for MRP1. In this study we aim to compare transport kinetics for sensitive and resistant human colorectal adenocarcinoma cell lines, in the presence and absence of verapamil and BSO, through 99mTc-Sestamibi. Pgp, MRP1 and LRP expression was evaluated in resistant (LS1034) and sensitive (WiDr) human colorectal adenocarcinoma cell lines using flow-cytometry. Western blot was also used to analyze Pgp expression. Cellular transport kinetics was assessed in the presence and absence of verapamil and BSO. Retention studies were performed after cell incubation with those drugs, for different time intervals (10 and 60 minutes) and concentrations (10, 25, 50 and 100 µM), using 99mTc-Sestamibi. Pgp expression was significantly higher (p < 0.05) in resistant cells, although LRP was also expressed. Western blotting analysis confirmed flow-cytometry results. 99mTc-Sestamibi retention percentage was significantly higher (p < 0.05) in the resistant cell line for all time-points considered. In resistant cells incubated with MDR modulators there were statistically significant differences (p < 0.05) among the modulators used but only for the first minutes after incubation with the radiotracer. The data obtained suggest that these modulators should be used immediately before the cytotoxic drugs are administrated.

Study on the signal pathway of signal transducer and activator Oftranscription 5in Iungadenocayinoma

Objective To elucidate the relationship of signal transducer and activator of transcription 5 (STAT5) in hyperplasia and differentiation,anti-apoptos and drug resistance of lung Adenocarcinoma.Methods CARP-resistant A549 (A549/CARP)was developed by means of different dosages and the sensititvty of anticancer of anticancer drugs were screened in the A549/CARP by MTT method.Immunohi stochemican detetion of signal transducer and activator of transcription 5,lung resistance-related protein (LRP) and multiple resistance protein(MRP) was carried out in A549 and A549/CARP.Results Resistant subline A549/CARP was est lished.Mmunohistemistry; there was no significant difference in the expression of STAT5 between A549/ and A549/CARP(P ＞ 0.05).There was significant difference in the expression of LRP and MRP between A549/CARP P ＜ 0.01.The expression of STAT5 is not correlated with LRP and MRP(P ＞ 0.05).There was no significant in the expression of STA5 in A549/CARP(P ＞ 0.05).The expression of STAT5 is not correlated with LRP and MRP(P ＞ 0.05).Conclusion The overexpression of LRP and MRP in drug resistance cell subline and low expression in adenocarcinoma may play a positive role in drug resistance.The drug resistan mechanism of LR-Pand MRP is not correlated with the pathway of STAT5.It is suggested that STAT5 play a positive role in hyperplasia,differentiation and anti-apoptosis,but not in drug resistance,which will contribute to the study of treatment of lung carcinoma. Key words: Adenocarcinoma; Resistant cells; Signal transducer and activator of transcription 5; Lung resistance-related protein; multiple resistance protein

Immunohistochemical Detection of Proteins Associated with Multidrug Resistance to Anti-Cancer Drugs in Canine and Feline Primary Pulmonary Carcinoma

Fifty-two canine and eighteen feline primary pulmonary carcinomas were evaluated immunohistochemically for the expression of proteins associated with multidrug resistance to anti-cancer drugs. P-glycoprotein (PGP), multidrug resistance-related protein (MRP) and lung resistance-related protein (LRP) expression were frequently observed in neoplastic cells of all carcinoma types, and metallothionein (MT) expression was observed in about half of each carcinoma type. Furthermore, overlapping expression was detected in all positive cases. These results indicate that most canine and feline primary pulmonary carcinomas may have strong multidrug resistance, which is related to the PGP, MRP, LRP or MT expression. It might be difficult to treat canine and feline primary pulmonary carcinomas using anti-cancer drugs because of multidrug resistance.

Expression of MRP1, BCRP, LRP, and ERCC1 in Advanced Non–Small-Cell Lung Cancer: Correlation With Response to Chemotherapy and Survival

Purpose We investigated the prognostic value of the expression of multidrug resistance protein-1 ( MRP1), breast cancer resistance protein ( BCRP), lung resistance-related protein ( LRP), and excision repair cross-complementing group-1 ( ERCC1) in patients with advanced Non–Small-cell lung cancer (NSCLC) treated with cisplatin-based chemotherapy. Patients and Methods Semiquantitative reverse-transcriptase polymerase chain reaction (RT-PCR) was used for detecting the expression of MRP1, BCRP, LRP, and ERCC1 mRNA in 66 transbronchial biopsy (TBB) samples from untreated patients with advanced NSCLC. All of the patients received cisplatin-based chemotherapy. Response to chemotherapy, progression-free survival (PFS), and overall survival (OS) were compared in relation to expression of each gene and clinicopathologic factors. Results Results showed that tumor stage ( P = .028) and the expression of MRP1 ( P = .046) and LRP ( P = .012) correlated with response to chemotherapy. Poor performance status (PS; P = .016), advanced tumor stage ( P = .004), and the high expression of MRP1 ( P = .012) and LRP ( P = .002) predicted poorer PFS. Performance status ( P = .009); tumor stage ( P = .003); and the expression of MRP1 ( P = .017), LRP ( P = .005), and ERCC1 ( P = .002) were predictive for OS. In a Cox proportional hazards multivariable analysis, PS ( P = .042), tumor stage ( P = .007), and the expression of LRP ( P = .011) and ERCC1 ( P = .026) were identified as independent prognostic factors for OS. Conclusion Our data suggested that determination of MRP1, LRP, and ERCC1 mRNA expression using RT-PCR in TBB samples might be helpful in predicting outcome of patients with advanced NSCLC treated with cisplatin-based chemotherapy and in optimizing therapeutic strategy based on the expression of these genes.