Genetic diseases seriously threaten human health and have always been one of the refractory conditions facing humanity.Currently,gene therapy drugs such as siRNA,shRNA,antisense oligonucleotide,CRISPR/Cas9 system,plas...Genetic diseases seriously threaten human health and have always been one of the refractory conditions facing humanity.Currently,gene therapy drugs such as siRNA,shRNA,antisense oligonucleotide,CRISPR/Cas9 system,plasmid DNA and miRNA have shown great potential in biomedical applications.To avoid the degradation of gene therapy drugs in the body and effectively deliver them to target tissues,cells and organelles,the development of excellent drug delivery vehicles is of utmost importance.Viral vectors are the most widely used delivery vehicles for gene therapy in vivo and in vitro due to their high transfection efficiency and stable transgene expression.With the development of nanotechnology,novel nanocarriers are gradually replacing viral vectors,emerging superior performance.This review mainly illuminates the current widely used gene therapy drugs,summarizes the viral vectors and non-viral vectors that deliver gene therapy drugs,and sums up the application of gene therapy to treat genetic diseases.Additionally,the challenges and opportunities of the field are discussed from the perspective of developing an effective nano-delivery system.展开更多
This study was designed to measure the multi-drug resistance gene (MDR1) mRNA content and analyze clinical relationship between MDR1 expression and drug resistance in primary ovarian cancer. Reverse transcription PCR ...This study was designed to measure the multi-drug resistance gene (MDR1) mRNA content and analyze clinical relationship between MDR1 expression and drug resistance in primary ovarian cancer. Reverse transcription PCR (RT-PCR) was used to measure MDR1 mRNA content in biopsy sample of 31 primary ovarian cancers (experimental group) and 30 gynecological tumors (control group). The level of 95.2% (20/21) MDR1 expression was relatively low, and the detected rate of MDR1 expression was 67.7%(21/31) in experimental group,which was higher than that in control group (40.0%, P<0.05). The differences of MDR1 expression between the effective group and no effect group after combined chemotherapy was significant (P<0.05). No significant relationship was found between MDR1 expression and clinical stage or histological classification or grade of differentiation in experimental group. We are led to concluded that primary ovarian cancers have drug-resistance clones which might express MDR1 spontaneously and expression of MDR1 may be used as a prognostic and predictive indicator for clinical response of ovarian cancers to combined chemotherapy.展开更多
Objective:The aim of the study was to identify specific chemosensitivity drugs for various molecular subtypes of breast tumors in Chinese women,by detecting the expression of drug resistance genes and by using the dru...Objective:The aim of the study was to identify specific chemosensitivity drugs for various molecular subtypes of breast tumors in Chinese women,by detecting the expression of drug resistance genes and by using the drug sensitivity test on different molecular subtypes of breast cancers.Methods:The expression of drug resistance genes including Topo Ⅱ,GST-π,P-gp,LRP,and CD133 were detected with immunohistochemistry in a tissue microarray.Drug sensitivity tests included those for paclitaxel,epirubicin,carboplatin,vinorelbine,and fluorouracil and were conducted on primary cancer tissue cells and cell lines,including the T47 D,BT-474,and MDA-MB-231 cells and human breast cancer xenografts in nude mice.Results:The different drug resistant genes Topo Ⅱ,GST-π,P-gp,and LRP were differentially expressed among different molecular subtypes of breast cancers(P<0.05).Positive expression of CD133 was highest in basal-like breast cancer(P<0.05).Kaplan-Meier survival analysis showed that positive expressions of Topo Ⅱ and CD133 both correlated with shorter disease-free survival(DFS)(P<0.05)and overall survival(P<0.05),and positive expression of LRP correlated only with shorter DFS(P<0.05).BT-474 showed chemosensitivity to paclitaxel and epirubicin,while MDA-MB-231 showed chemosensitivities to paclitaxel,epirubicin,carboplatin,and fluorouracil(T/C≤50%).The basal-like and HER2+breast cancer primary cells showed chemosensitivities to paclitaxel and epirubicin with significant differences compared with luminal breast cancer primary cells(P<0.05).Conclusions:The differential expression of drug resistance genes and the differential chemosensitivities of drugs in different molecular subtype of breast cancers suggested that individual treatment should be given for each type of breast cancer.展开更多
To establish a cytologic expressing system of rat glutathione S-transferase pi (GST-pi) cDNA for detecting the resistance of HeLa cells to anticancer drugs. Methods The assessment was made with various anticancer drug...To establish a cytologic expressing system of rat glutathione S-transferase pi (GST-pi) cDNA for detecting the resistance of HeLa cells to anticancer drugs. Methods The assessment was made with various anticancer drugs (adriamycin, mitomycin, cisplatinum and vincristine) that showed different cytotoxicities in transfectant HeLa cells with pSV-GT containing rat GST-pi cDNA (HeLa/pSV-GT) or control pSV-neo (HeLa/pSV-neo). Expression levels of GST-pi mRNA in HeLa/pSV-GT and HeLa/pSV-neo were measured by in situ hybridization using Digoxin-labelled cDNA probe. Results HeLa/pSV-GT expressed significantly high degree of GST-pi mRNA, whereas both HeLa/pSV-neo and HeLa cells had very low expression. Cytotoxicities of HeLa/pSV-GT and HeLa/pSV-neo with 4 anticancer drugs were measured by MTT assay. Drug concentrations for yielding 50% inhibition (IC50) in HeLa/pSV-GT by adriamycin, mitomycin and cisplatinum were 70.13 靏/mL, 10.95 靏/mL and 16.52 靏/mL, respectively. In contrast, IC50 in HeLa/pSV-neo was 10.34 靏/mL, 7.48 靏/mL and 13.70 靏/mL, respectively. The cytotoxicities of vincristine on both HeLa/pSV-GT and HeLa/pSV-neo were not significantly different. Conclusions Our findings suggest that HeLa/pSV-GT containing rat GST-pi cDNA is resistant to some anticancer drugs due to overexpression of GST-pi. Also, HeLa/pSV-GT cell line could serve as a useful cytogenetic model for further research.展开更多
Objective: To study on the effect of MRP gene overexpression on prognosis of patients with non-small lung cancer (NSCLC). Methods: Paraffin-embedded tissues from 47 cases of NSCLC who had undergone radical tumor resec...Objective: To study on the effect of MRP gene overexpression on prognosis of patients with non-small lung cancer (NSCLC). Methods: Paraffin-embedded tissues from 47 cases of NSCLC who had undergone radical tumor resection were examined for expression of MRP gene mRNA by in situ hybridization using labelled digoxigenin probes combined with immunohistochemistry. All the patients were retrospectively followed-up. Results: All of the 47 lung cancer specimens were found to have overexpression of MRP gene mRNA. It was significantly correlated with patients’ survival time, response to chemotherapy, recurrence or metastases after surgery, but was not correlated with histology, tumor size, node status, TNM stage, degree of differentiation, age and sex. Conclusion: Overexpression of MRP gene is a marker of prognostic significance in patients with NSCLC.展开更多
Objective:To investigate the influence of GST-π gene transfer on drug-resistance of human cord blood CD34 + cells.Methods:CD34 + cells were purified from cord blood from normal full-term pregnancy.Gene transduction i...Objective:To investigate the influence of GST-π gene transfer on drug-resistance of human cord blood CD34 + cells.Methods:CD34 + cells were purified from cord blood from normal full-term pregnancy.Gene transduction into human cord blood CD34 + cells was carried out using GST-π gene containing retrovirus vector.The GST-π gene expression in transduced CD34 + cell was confirmed by RT-PCR.After confirmation of GST-π gene transfer,the transfected CD34 + cells were cultured by colony assay in the presence of carboplatin.Results:GST-π mRNA was detected in 30% of CFU-GM derived from GST-π gene transduced CD34 + cells.In vitro drug resistance test showed that the number of CFU-GM formed was significantly higher (2~3 fold) in GST-π gene transduced CD34 + cells than untransduced CD34 +cells.Conclusion:GST-π gene transfer can confer resistance to hematopoietic progenitors against carboplatin in vitro.展开更多
Objective: To investigate the co-expression of drug resistance- and apoptosis-related genes of cisplatin (CDDP)-selected lung adenocarcinoma cell line A549DDP for compared to the parental cell line A549, and reverse...Objective: To investigate the co-expression of drug resistance- and apoptosis-related genes of cisplatin (CDDP)-selected lung adenocarcinoma cell line A549DDP for compared to the parental cell line A549, and reverse of drug resistance by antisense s-oligodeoxynucleotides (S-ODNs) of differentially expressed genes. Methods: Sense and antisense S-ODN were transferred into A549DDP cells by lipofectin. The expression of drug resistance and apoptosis related genes was examined by RT-PCR, immunocytochemistry and flow cytometry, respectively. Apoptostic cells were identified by DNA electrophoresis and terminal deoxynucleotidyl transferase (TdT)-mediated biotin dUTP nick end- labeling(TUNEL). Drug resistance of tumor cells was detected by a cell viability (MTT) assay. Results: The expression of bc1-2 was positive and that of multidrug resistance-associated protein (MRP) at mRNA and protein level was increased in A549DDP compared to A549 cells. MDRI, c-myc and topoisomeras Ⅱ (TOPO Ⅱ) were similarly co-expressed in two cell lines. Both cell lines were negative for c-erbB-2 expression. In A549DDP cells, the expression of bc1-2 and MRP was sigllificantly inhibited by their respective antisense S-ODNs. Antisensc S-ODNs could also decrease significantly drug resistance of A549DDP cells to CDDP by promoting cell apoptosis. Conclusion: Both intrinsic and acquired drug resistance were involved in co-expression of multiple MDR-related genes in lung adenocarcinoma. Co- operation of hcf-2 and MRP genes appeared to play an important action to confer the resistance of A549DDP cells to CDDP. Their antisense S-ODNs are responsible for the decrease of drug resistance of this cell line by promoting apoptosis.展开更多
Summary: The expression and functional activity of multiple drug resistance (MDR1) gene in human normal bone marrow CD34+ cells was observed. Human normal bone marrow CD34+ cells were enriched with magnetic cell sorti...Summary: The expression and functional activity of multiple drug resistance (MDR1) gene in human normal bone marrow CD34+ cells was observed. Human normal bone marrow CD34+ cells were enriched with magnetic cell sorting (MACS) system, and then liposome-mediated MDR1 gene was transferred into bone marrow CD34+ cells. Fluorescence-activated cell sorter was used to evaluate the expression and functional activity of P-glycoprotein (P-gp) encoded by MDR1 gene. It was found that the purity of bone marrow CD34+ cells was approximately (91±4.56) % and recovery rate was (72.3±2.36) % by MACS. The expression of P-gp in the transfected CD34+cells was obviously higher than that in non-transfected CD34+ cells. The amount of P-gp in non-transfected CD34+ cells was (11.2±2.2) %, but increased to (23.6±2.34) % 48 h after gene transfection (P<0.0l). The amount of P-gp was gradually decreased to the basic level one week later. The accumulation and extrusion assays showed that the overexpression of P-gp could efflux Rh-123 out of cells and there was low fluorescence within the transfected cells. The functional activity of P-gp could be inhibited by 10 μg/ml verapamil. It was suggested that the transient and highly effective expression and functional activity of P-gp could be obtained by liposome-mediated MRD1 transferring into human normal bone marrow CD34+ cells.展开更多
AIM: To study the action of the genes associated with drug-induced liver diseases at the gene transcriptional level during liver regeneration (LR) in rats. METHODS: The genes associated with drug-induced liver disease...AIM: To study the action of the genes associated with drug-induced liver diseases at the gene transcriptional level during liver regeneration (LR) in rats. METHODS: The genes associated with drug-induced liver diseases were obtained by collecting the data from databases and literature, and the gene expression changes in the regenerating liver were checked by the Rat Genome 230 2.0 array. RESULTS: The initial and total expression numbers of genes occurring in phases of 0.5-4 h after partial hepatectomy (PH), 4-6 h after PH (G0/G1 transition), 6-66 h after PH (cell proliferation), 66-168 h after PH (cell differentiation and structure-function reconstruction) were 21, 3, 9, 2 and 21, 9, 19, 18, respectively. It is illustrated that the associated genes were mainly triggered at the initial stage of LR and worked at different phases. According to their expression similarity, these genes were classified into 5 types: only up- regulated (12 genes), predominantly up-regulated (4 genes), only down-regulated (11 genes), predominantly down-regulated (3 genes), and approximately up-/ down-regulated (2 genes). The total times of their up- and down-expression were 130 and 79, respectively, demonstrating that expression of most of the genes was increased during LR, while a few decreased. The cell physiological and biochemical activities during LR were staggered according to the time relevance and were diverse and complicated in gene expression patterns. CONCLUSION: Drug metabolic capacity in regenerating liver was enhanced. Thirty-two genes play important roles during liver regeneration in rats.展开更多
基金supported by the National Natural Science Foundation of China(No.51472115)Double Firstclass Innovation Team of China Pharmaceutical University(CPU2018GY40).
文摘Genetic diseases seriously threaten human health and have always been one of the refractory conditions facing humanity.Currently,gene therapy drugs such as siRNA,shRNA,antisense oligonucleotide,CRISPR/Cas9 system,plasmid DNA and miRNA have shown great potential in biomedical applications.To avoid the degradation of gene therapy drugs in the body and effectively deliver them to target tissues,cells and organelles,the development of excellent drug delivery vehicles is of utmost importance.Viral vectors are the most widely used delivery vehicles for gene therapy in vivo and in vitro due to their high transfection efficiency and stable transgene expression.With the development of nanotechnology,novel nanocarriers are gradually replacing viral vectors,emerging superior performance.This review mainly illuminates the current widely used gene therapy drugs,summarizes the viral vectors and non-viral vectors that deliver gene therapy drugs,and sums up the application of gene therapy to treat genetic diseases.Additionally,the challenges and opportunities of the field are discussed from the perspective of developing an effective nano-delivery system.
文摘This study was designed to measure the multi-drug resistance gene (MDR1) mRNA content and analyze clinical relationship between MDR1 expression and drug resistance in primary ovarian cancer. Reverse transcription PCR (RT-PCR) was used to measure MDR1 mRNA content in biopsy sample of 31 primary ovarian cancers (experimental group) and 30 gynecological tumors (control group). The level of 95.2% (20/21) MDR1 expression was relatively low, and the detected rate of MDR1 expression was 67.7%(21/31) in experimental group,which was higher than that in control group (40.0%, P<0.05). The differences of MDR1 expression between the effective group and no effect group after combined chemotherapy was significant (P<0.05). No significant relationship was found between MDR1 expression and clinical stage or histological classification or grade of differentiation in experimental group. We are led to concluded that primary ovarian cancers have drug-resistance clones which might express MDR1 spontaneously and expression of MDR1 may be used as a prognostic and predictive indicator for clinical response of ovarian cancers to combined chemotherapy.
基金supported by the National Natural Science Foundation of China(Grant No.81502309)。
文摘Objective:The aim of the study was to identify specific chemosensitivity drugs for various molecular subtypes of breast tumors in Chinese women,by detecting the expression of drug resistance genes and by using the drug sensitivity test on different molecular subtypes of breast cancers.Methods:The expression of drug resistance genes including Topo Ⅱ,GST-π,P-gp,LRP,and CD133 were detected with immunohistochemistry in a tissue microarray.Drug sensitivity tests included those for paclitaxel,epirubicin,carboplatin,vinorelbine,and fluorouracil and were conducted on primary cancer tissue cells and cell lines,including the T47 D,BT-474,and MDA-MB-231 cells and human breast cancer xenografts in nude mice.Results:The different drug resistant genes Topo Ⅱ,GST-π,P-gp,and LRP were differentially expressed among different molecular subtypes of breast cancers(P<0.05).Positive expression of CD133 was highest in basal-like breast cancer(P<0.05).Kaplan-Meier survival analysis showed that positive expressions of Topo Ⅱ and CD133 both correlated with shorter disease-free survival(DFS)(P<0.05)and overall survival(P<0.05),and positive expression of LRP correlated only with shorter DFS(P<0.05).BT-474 showed chemosensitivity to paclitaxel and epirubicin,while MDA-MB-231 showed chemosensitivities to paclitaxel,epirubicin,carboplatin,and fluorouracil(T/C≤50%).The basal-like and HER2+breast cancer primary cells showed chemosensitivities to paclitaxel and epirubicin with significant differences compared with luminal breast cancer primary cells(P<0.05).Conclusions:The differential expression of drug resistance genes and the differential chemosensitivities of drugs in different molecular subtype of breast cancers suggested that individual treatment should be given for each type of breast cancer.
基金the National Natural Science Foundation of China.
文摘To establish a cytologic expressing system of rat glutathione S-transferase pi (GST-pi) cDNA for detecting the resistance of HeLa cells to anticancer drugs. Methods The assessment was made with various anticancer drugs (adriamycin, mitomycin, cisplatinum and vincristine) that showed different cytotoxicities in transfectant HeLa cells with pSV-GT containing rat GST-pi cDNA (HeLa/pSV-GT) or control pSV-neo (HeLa/pSV-neo). Expression levels of GST-pi mRNA in HeLa/pSV-GT and HeLa/pSV-neo were measured by in situ hybridization using Digoxin-labelled cDNA probe. Results HeLa/pSV-GT expressed significantly high degree of GST-pi mRNA, whereas both HeLa/pSV-neo and HeLa cells had very low expression. Cytotoxicities of HeLa/pSV-GT and HeLa/pSV-neo with 4 anticancer drugs were measured by MTT assay. Drug concentrations for yielding 50% inhibition (IC50) in HeLa/pSV-GT by adriamycin, mitomycin and cisplatinum were 70.13 靏/mL, 10.95 靏/mL and 16.52 靏/mL, respectively. In contrast, IC50 in HeLa/pSV-neo was 10.34 靏/mL, 7.48 靏/mL and 13.70 靏/mL, respectively. The cytotoxicities of vincristine on both HeLa/pSV-GT and HeLa/pSV-neo were not significantly different. Conclusions Our findings suggest that HeLa/pSV-GT containing rat GST-pi cDNA is resistant to some anticancer drugs due to overexpression of GST-pi. Also, HeLa/pSV-GT cell line could serve as a useful cytogenetic model for further research.
基金the Natural Science Foundation of Shanghai, China! (96ZB14043).
文摘Objective: To study on the effect of MRP gene overexpression on prognosis of patients with non-small lung cancer (NSCLC). Methods: Paraffin-embedded tissues from 47 cases of NSCLC who had undergone radical tumor resection were examined for expression of MRP gene mRNA by in situ hybridization using labelled digoxigenin probes combined with immunohistochemistry. All the patients were retrospectively followed-up. Results: All of the 47 lung cancer specimens were found to have overexpression of MRP gene mRNA. It was significantly correlated with patients’ survival time, response to chemotherapy, recurrence or metastases after surgery, but was not correlated with histology, tumor size, node status, TNM stage, degree of differentiation, age and sex. Conclusion: Overexpression of MRP gene is a marker of prognostic significance in patients with NSCLC.
文摘Objective:To investigate the influence of GST-π gene transfer on drug-resistance of human cord blood CD34 + cells.Methods:CD34 + cells were purified from cord blood from normal full-term pregnancy.Gene transduction into human cord blood CD34 + cells was carried out using GST-π gene containing retrovirus vector.The GST-π gene expression in transduced CD34 + cell was confirmed by RT-PCR.After confirmation of GST-π gene transfer,the transfected CD34 + cells were cultured by colony assay in the presence of carboplatin.Results:GST-π mRNA was detected in 30% of CFU-GM derived from GST-π gene transduced CD34 + cells.In vitro drug resistance test showed that the number of CFU-GM formed was significantly higher (2~3 fold) in GST-π gene transduced CD34 + cells than untransduced CD34 +cells.Conclusion:GST-π gene transfer can confer resistance to hematopoietic progenitors against carboplatin in vitro.
基金the Grant from Beijing Natural Science Foundation (No.7992005)and a Grantfrom Postdoctoral Foundation of National Committee of E
文摘Objective: To investigate the co-expression of drug resistance- and apoptosis-related genes of cisplatin (CDDP)-selected lung adenocarcinoma cell line A549DDP for compared to the parental cell line A549, and reverse of drug resistance by antisense s-oligodeoxynucleotides (S-ODNs) of differentially expressed genes. Methods: Sense and antisense S-ODN were transferred into A549DDP cells by lipofectin. The expression of drug resistance and apoptosis related genes was examined by RT-PCR, immunocytochemistry and flow cytometry, respectively. Apoptostic cells were identified by DNA electrophoresis and terminal deoxynucleotidyl transferase (TdT)-mediated biotin dUTP nick end- labeling(TUNEL). Drug resistance of tumor cells was detected by a cell viability (MTT) assay. Results: The expression of bc1-2 was positive and that of multidrug resistance-associated protein (MRP) at mRNA and protein level was increased in A549DDP compared to A549 cells. MDRI, c-myc and topoisomeras Ⅱ (TOPO Ⅱ) were similarly co-expressed in two cell lines. Both cell lines were negative for c-erbB-2 expression. In A549DDP cells, the expression of bc1-2 and MRP was sigllificantly inhibited by their respective antisense S-ODNs. Antisensc S-ODNs could also decrease significantly drug resistance of A549DDP cells to CDDP by promoting cell apoptosis. Conclusion: Both intrinsic and acquired drug resistance were involved in co-expression of multiple MDR-related genes in lung adenocarcinoma. Co- operation of hcf-2 and MRP genes appeared to play an important action to confer the resistance of A549DDP cells to CDDP. Their antisense S-ODNs are responsible for the decrease of drug resistance of this cell line by promoting apoptosis.
文摘Summary: The expression and functional activity of multiple drug resistance (MDR1) gene in human normal bone marrow CD34+ cells was observed. Human normal bone marrow CD34+ cells were enriched with magnetic cell sorting (MACS) system, and then liposome-mediated MDR1 gene was transferred into bone marrow CD34+ cells. Fluorescence-activated cell sorter was used to evaluate the expression and functional activity of P-glycoprotein (P-gp) encoded by MDR1 gene. It was found that the purity of bone marrow CD34+ cells was approximately (91±4.56) % and recovery rate was (72.3±2.36) % by MACS. The expression of P-gp in the transfected CD34+cells was obviously higher than that in non-transfected CD34+ cells. The amount of P-gp in non-transfected CD34+ cells was (11.2±2.2) %, but increased to (23.6±2.34) % 48 h after gene transfection (P<0.0l). The amount of P-gp was gradually decreased to the basic level one week later. The accumulation and extrusion assays showed that the overexpression of P-gp could efflux Rh-123 out of cells and there was low fluorescence within the transfected cells. The functional activity of P-gp could be inhibited by 10 μg/ml verapamil. It was suggested that the transient and highly effective expression and functional activity of P-gp could be obtained by liposome-mediated MRD1 transferring into human normal bone marrow CD34+ cells.
基金Supported by the National Natural Science Foundation of China, No. 30270673
文摘AIM: To study the action of the genes associated with drug-induced liver diseases at the gene transcriptional level during liver regeneration (LR) in rats. METHODS: The genes associated with drug-induced liver diseases were obtained by collecting the data from databases and literature, and the gene expression changes in the regenerating liver were checked by the Rat Genome 230 2.0 array. RESULTS: The initial and total expression numbers of genes occurring in phases of 0.5-4 h after partial hepatectomy (PH), 4-6 h after PH (G0/G1 transition), 6-66 h after PH (cell proliferation), 66-168 h after PH (cell differentiation and structure-function reconstruction) were 21, 3, 9, 2 and 21, 9, 19, 18, respectively. It is illustrated that the associated genes were mainly triggered at the initial stage of LR and worked at different phases. According to their expression similarity, these genes were classified into 5 types: only up- regulated (12 genes), predominantly up-regulated (4 genes), only down-regulated (11 genes), predominantly down-regulated (3 genes), and approximately up-/ down-regulated (2 genes). The total times of their up- and down-expression were 130 and 79, respectively, demonstrating that expression of most of the genes was increased during LR, while a few decreased. The cell physiological and biochemical activities during LR were staggered according to the time relevance and were diverse and complicated in gene expression patterns. CONCLUSION: Drug metabolic capacity in regenerating liver was enhanced. Thirty-two genes play important roles during liver regeneration in rats.