Biomedical subjects

Hermann Lage

Publications and source records attributed to Hermann Lage.

50 records · Page 3Linked to original sources

PubMed2003

ABC-transporters: implications on drug resistance from microorganisms to human cancers.

Resistance to chemotherapy is a common clinical problem in patients with infectious diseases as well as in patients with cancer. During treatment of infections or malignant tumors, the drug targets of prokaryotic or eukaryotic microorganisms and neoplastic cells are often found to be refractory to a variety of drugs that have different structures and functions. This phenomenon has been termed multidrug resistance (MDR). The mechanisms leading to MDR are frequently caused by trans-membrane xenobiotic transport molecules belonging to the superfamily of ATP-binding cassette (ABC) transporters. There is an urgent need to understand the structure-function relationships of these efflux pumps that underlie their transport mechanism and drug selectivity. This knowledge may allow the rational design of new drugs that can inhibit or circumvent the activity of these MDR transport molecules. Furthermore, the development of such chemosensitizing agents would help us learn more about the physiological functions and substrates of these pump proteins. This review will discuss the current state of knowledge of the functional and structural similarities among ABC-transporters in prokaryotic and eukaryotic cells and their impact on MDR.

ATP-Binding Cassette Transporters↗

PubMed2003

Overexpression of cMOAT (MRP2/ABCC2) is associated with decreased formation of platinum-DNA adducts and decreased G2-arrest in melanoma cells resistant to cisplatin.

Resistance to various anti-neoplastic agents is a common observation in clinical management of melanoma. The biologic mechanisms conferring these different drug-resistant phenotypes, including resistance against the commonly used anti-cancer drug cisplatin, are unclear. In order to elucidate the role of the membrane adenosine triphosphate binding cassette-transporter cMOAT (canalicular multispecific anion transporter) (MRP2/ABCC2) in cisplatin resistance of melanoma, the expression of this protein was analyzed in the platinum drug-resistant cell line MeWo CIS 1. Cisplatin-resistant melanoma cells showed a distinct overexpression of cMOAT on mRNA and protein level. This observation was accompanied by a reduced formation of platinum-induced intrastrand cross-links in the nuclear DNA measured by an immunocytologic assay. This decrease in DNA platination was accompanied by an accelerated re-entry into the cell cycle after the typical cisplatin-induced G2 arrest, and a resistance to undergo apoptosis. Kinetics of formation and elimination of platinum-DNA adducts suggest that the DNA repair capacity for Pt-d(GpG) adducts was not elevated in platinum drug-resistant melanoma cells. The decrease in platinum-DNA adduct formation in cisplatin-resistant melanoma cells was rather a reflection of the protecting activity of the transporter cMOAT. In conclusion, the functional inhibition of cMOAT might be a promising strategy in the reversal of resistance to platinum-based anti-cancer drugs in human melanoma.

Antineoplastic Agents↗

PubMed2003

Molecular analysis of therapy resistance in gastric cancer.

Therapy resistance is the main cause of therapeutic failure and death in patients suffering from gastric carcinoma. Clinical resistance against systemic chemotherapy of gastric cancer is likely to be multifactorial and heterogenous. So far, no significant resistance factor that predicts the clinical outcome of systemic treatment of gastric carcinoma has been identified. In order to gain further understanding of therapy resistance in gastric carcinoma, various in vitro model systems were established. One of these models consists of the parental, drug-sensitive and thermosensitive human gastric carcinoma cell line EPG85-257P, its classical multidrug-resistant variant EPG85-257RDB, its atypical multidrug-resistant subline EPG85-257RNOV and their thermoresistant counterparts EPG85-257P-TR, EPG85-257RDB-TR, and EPG85-257RNOV-TR. This panel of cells was analyzed using morphological, biochemical, cellular and molecular biological methods to identify potential new factors involved in therapy resistance of gastric carcinoma. Cellular alterations that could be identified in these models were evaluated by functional investigations. This review will discuss the current state of knowledge of these new therapy resistance-associated factors, e.g. glypican-3 (GPC3), as well as the impact of well-known drug resistance-associated factors, such as MDR1/P-glycoprotein, on therapy resistance of gastric carcinoma.

Antineoplastic Agents↗

PubMed2002

Transport of phosphatidylserine via MDR1 (multidrug resistance 1)P-glycoprotein in a human gastric carcinoma cell line.

The ATP-binding cassette transporter multidrug resistance 1 P-glycoprotein (MDR1 Pgp) has been implicated with the transport of lipids from the inner to the outer leaflet of the plasma membrane. While this has been unambigously shown for the fluorescent lipid analogues [N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoyl (C6-NBD)-phosphatidylcholine, -phosphatidylethanolamine, -sphingomyelin and -glucosylceramide, by using a novel approach we have now found significantly increased outward transport also for C6-NBD-phosphatidylserine (C6-NBD-PS) in EPG85-257 human gastric carcinoma cells overexpressing MDR1 (coding for MDR1 Pgp). The increased transport of C6-NBD-PS is mediated by MDR1 Pgp, shown by transport reduction nearly to the level of controls in the presence of MDR1 Pgp inhibitors [PSC 833, cyclosporin A and dexniguldipine hydrochloride (Dex)]. Addition of MK 571, a specific inhibitor of the MDR protein MRP1, does not decrease transport in either of the two cell lines. The plasma-membrane association of FITC-annexin V, a fluorescent protein conjugate binding PS, is significantly increased in MDR1-overexpressing cells as compared with controls, and can be reduced by an MDR1 Pgp inhibitor. This suggests that MDR1 Pgp transports endogenous PS, the lipid exhibiting the most pronounced transverse asymmetry in the plasma membrane.

4-Chloro-7-nitrobenzofurazan↗

PubMed2002

Multiple mechanisms confer different drug-resistant phenotypes in pancreatic carcinoma cells.

PURPOSE: Drug-resistant phenotypes of cancer cells may be caused by complex multimodal mechanisms of resistance. In order to gain further insighte into these mechanisms, a P-glycoprotein-mediated multidrug-resistant phenotype induced by daunorubicin-selection and an alternative drug resistance due to treatment with mitoxantrone were investigated in pancreatic carcinoma-derived cells. METHODS: For assessing cross-resistance against various drugs, cell proliferation assays were performed. Drug accumulation was measured by flow cytometry. Messenger RNA expression was analyzed by Northern blot and RT-PCR, whereas protein expression was determined by Western blot. Catalytic activity of DNA-topoisomerases (Topo) II was determined by the decatenation assay. RESULTS: In mitoxantrone-selected EPP85-181RNOV cells a decreased accumulation of mitoxantrone and daunorubicin was observed in the absence of P-glycoprotein, multidrug resistance protein or breast cancer resistance protein over-expression. An approximately twofold decrease of DNA topoisomerase II catalytic activity could be observed in both drug-resistance-exhibiting cell lines. The reduction of Topo II catalytic activity was reflected by decreased expression of Topo IIalpha and IIbeta mRNAs and proteins. CONCLUSIONS: The decreased drug accumulation in EPP85-181RNOV cells indicates that alternative transport events are occurring. The decreased catalytic activity and expression of Topo II indicate that modulation of Topo II catalytic activity contributes to both drug-resistant phenotypes in pancreatic carcinoma cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2002

Impact of BCRP/MXR, MRP1 and MDR1/P-Glycoprotein on thermoresistant variants of atypical and classical multidrug resistant cancer cells.

The impact of the ABC transporters breast cancer resistance protein/mitoxantrone resistance associated transporter (BCRP/MXR), multidrug resistance-associated protein 1 (MRP1) and multidrug resistance gene-1/P-glycoprotein (MDR1/PGP) on the multidrug resistance (MDR) phenotype in chemoresistance and thermoresistance was investigated in the parental human gastric carcinoma cell line EPG85-257P, the atypical MDR subline EPG85-257RNOV, the classical MDR subline EPG85-257RDB and their thermoresistant counterparts EPG85-257P-TR, EPG85-257RNOV-TR and EPG85-257RDB-TR. Within the atypical MDR subline EPG85-257RNOV expression of BCRP/MXR and of MRP1 were clearly enhanced (vs. parental and classical MDR lines). MDR1/PGP expression was distinctly elevated in the classical MDR subline EPG85-257RDB (vs. parental and atypical MDR sublines). In all thermoresistant counterparts basal expression of BCRP/MXR, MRP1 and MDR1/PGP was increased relative to thermosensitive sublines. Although it could be shown that the overexpressed ABC transporters were functionally active, however, no decreased drug accumulations of doxorubicin, mitoxantrone and rhodamine 123 were observed. Thus, expression of BCRP/MXR, MRP1 and MDR1/PGP was found to be dependent on the appropriate type of chemoresistance; correlating with a classical or atypical MDR phenotype. Within the thermoresistant variants, however, the increase in ABC transporter expression did obviously not influence the MDR phenotype.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2002

YB-1 relocates to the nucleus in adenovirus-infected cells and facilitates viral replication by inducing E2 gene expression through the E2 late promoter.

The adenovirus early proteins E1A and E1B-55kDa are key regulators of viral DNA replication, and it was thought that targeting of p53 by E1B-55kDa is essential for this process. Here we have identified a previously unrecognized function of E1B for adenovirus replication. We found that E1B-55kDa is involved in targeting the transcription factor YB-1 to the nuclei of adenovirus type 5-infected cells where it is associated with viral inclusion bodies believed to be sites of viral transcription and replication. We show that YB-1 facilitates E2 gene expression through the E2 late promoter thus controlling E2 gene activity at later stages of infection. The role of YB-1 for adenovirus replication was demonstrated with an E1-minus adenovirus vector containing a YB-1 transgene. In infected cells, AdYB-1 efficiently replicated and produced infectious progeny particles. Thus, adenovirus E1B-55kDa protein and the host cell factor YB-1 act jointly to facilitate adenovirus replication in the late phase of infection.

Active Transport, Cell Nucleus↗

PubMed2002

Modulation of the atypical multidrug-resistant phenotype by a hammerhead ribozyme directed against the ABC transporter BCRP/MXR/ABCG2.

The phenomenon of multidrug resistance (MDR) in human cancers is one of the major causes of failure of chemotherapy. A recently identified new member of the superfamily of ATP-binding cassette transporters, breast cancer resistance protein (BCRP), was demonstrated to confer an atypical multidrug-resistant phenotype to tumor cells. To overcome the BCRP-mediated drug resistance, a specific anti-BCRP hammerhead ribozyme was introduced into the human gastric carcinoma cell line, EPG85-257RNOV, exhibiting an atypical MDR phenotype. By this approach, the expression levels of the targeted BCRP-encoding mRNA and the BCRP transport protein were decreased to the low constitutive expression level that was observed in highly drug-sensitive parental gastric carcinoma cells. In addition, in the anti-BCRP ribozyme-treated cells, the cellular drug accumulation was dramatically increased to the level measured in drug-sensitive cells. These effects were accompanied by an extensive reversal of the drug-resistant phenotype of more than 80%. Because additional mechanisms contribute to the multimodal-mediated MDR phenotype exhibited by this gastric carcinoma cell line, the data suggest that the BCRP-mediated contingent to the drug resistance was overcome nearly completely. Moreover, the data indicate that ribozyme-based gene therapy may be clinically applicable in preventing and reversing BCRP-mediated atypical MDR.

ATP Binding Cassette Transporter, Subfamily G, Mem↗

PubMed2002

Study of therapy resistance in cancer cells with functional proteome analysis.

Different types of cancer are naturally resistant to many anticancer drugs. Additionally, these tumours develop acquired drug resistance, which includes the classical multidrug resistance (MDR) accompanied by the synthesis of P-glycoprotein, a member of the superfamily of ATP-binding cassette (ABC) transporters. Furthermore, atypical MDR is mediated by several different, some unknown, mechanisms. To overcome chemoresistance problems, antineoplastic drugs are often combined with other modes of therapy, e.g. hyperthermia, where good response has been reported in several experimental tumour models and in advanced cancer patients. The success of this combined anticancer treatment may be limited by an increase in chemoresistance and thermoresistance. A model system to study resistance phenomena is the use of chemoresistant and thermoresistant cancer cell lines. We have established chemoresistant cancer cell lines (gastric and pancreatic carcinoma, fibrosarcoma, melanoma) and now thermoresistant cell lines derived from gastric and pancreatic carcinoma cells and their counterparts that were resistant towards daunorubicin (classical MDR) and mitoxantrone (atypical MDR). Using proteomics, in this paper we evaluate the drug resistance of chemoresistant melanoma cells (parental cell line MeWo and sublines exhibiting drug resistance towards etoposide, cisplatin, fotemustine and vindesine) as a paradigm for analysis of drug resistance phenomena. Additionally, we investigate heat resistance and the interaction of chemoresistance and thermoresistance to identify common pathways using the parental and drug resistant stomach cancer cell lines EPG85-257, EPG85-257RNOV, EPG85-257RDB and their thermoresistant counterparts. Possible implications of differential protein expression will be discussed.

Databases as Topic↗

PubMedDate not supplied

Identification of differentially expressed genes in classical and atypical multidrug-resistant gastric carcinoma cells.

The phenomenon of multidrug resistance (MDR) in human cancers is the major cause of failure of chemotherapy. To better understand the molecular events associated with the development of different types of MDR, a classical MDR P-glycoprotein expressing gastric carcinoma cell line and an atypical MDR P-glycoprotein-negative variant were analyzed by cDNA array hybridization. Of 588 cDNAs spotted on the array, a total of 9 genes showed differences in mRNA expressions. An enhanced expression level of 3 genes could be detected in both different MDR models (HLH, IR21, CCT5, Tx P-1), 4 genes were overexpressed solely in classical MDR cells (Hsp27, Rcl, aldehyde dehydrogenase 1, vimentin), whereas the expression of one gene was increased in atypical MDR cells (ProTa). Furthermore, the mRNA expressions of 3 genes were down-regulated in atypical MDR cells (Hsp27, vimentin, JNK2), whereas a decrease in mRNA expression in classical MDR cells could be determined for none of them. These differences were confirmed by a semiquantitative RT-PCR approach. Further characterization of these factors may provide more insight into the biology and development of different types of MDR in human gastric carcinomas. Moreover, these genes may be potential candidate factors for the diagnostics and/or prognosis of clinical drug resistance.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMedDate not supplied

Effective knock down of very high ABCG2 expression by a hammerhead ribozyme.

BACKGROUND: Ribozymes are an effective tool to reduce the mRNA levels of specific target genes. Overexpression of the drug transport protein, ABCG2, has been associated with multidrug resistance in cancer cells. MATERIALS AND METHODS: An expression plasmid encoding a hammerhead ribozyme against the ABCG2 gene was stably transfected into multidrug-resistant MCF7/MX cells that express very high levels of the ABCG2 protein. The effect of the ribozyme was determined by quantitative real-time RT-PCR, Western blot and cytotoxicity assays. RESULTS: The ribozyme reduced ABCG2 mRNA levels to less than 10% of control values, which resulted in the concomitant reduction of ABCG2 protein levels and sensitization of the cells to mitoxantrone and methotrexate. CONCLUSION: The ribozyme used was highly effective in reducing the expression of its target gene, ABCG2, and was able to modulate the associated multidrug-resistant phenotype.

ATP Binding Cassette Transporter, Subfamily G, Mem↗

PubMedDate not supplied

Transcriptome analysis of different multidrug-resistant gastric carcinoma cells.

Multidrug resistance (MDR) of human cancers is the major cause of failure of chemotherapy. To better understand the molecular events associated with the development of different types of MDR, two different multidrug-resistant gastric carcinoma cell lines, the MDR1/P-glycoprotein-expressing cell line EPG85-257RDB and the MDR1/P-glycoprotein-negative cell variant EPG85-257RNOV, as well as the corresponding drug-sensitive parental cell line EPG85-257P, were used for analyses of the mRNA expression profiles by cDNA array hybridization. Of more than 12,000 genes spotted on the arrays, 156 genes were detected as being significantly regulated in the cell line EPG85-257RDB in comparison to the non-resistant cell variant, and 61 genes were found to be differentially expressed in the cell line EPG85-257RNOV Seventeen genes showed a differential expression level in both multidrug-resistant gastric carcinoma variants. The impact of these alterations in gene expression levels in different multidrug-resistant gastric carcinoma cell variants is discussed.

Cell Line, Tumor↗

PubMedDate not supplied

CD24 expression is specific for tamoxifen-resistant ductal breast cancer cases.

BACKGROUND: In breast cancer, the expression of CD24 represents a poorly recognised unfavourable prognostic factor. CD24 has been described to be potentially down-regulated by estrogen receptor alpha (ER). The present study was aimed at examining the predictive value of CD24 expression in tamoxifen-treated breast cancer cases. MATERIALS AND METHODS: Sixty patients with primary invasive ductal breast cancers with post-operative tamoxifen treatment were enrolled in the study. Immmunohistochemical reactions were performed using monoclonal antibodies directed against CD24 and ER. RESULTS: Cases demonstrating cytoplasmic-membranous expression of CD24 (CD24c-m) proved to be characterised by a significantly lower expression of ER as compared to CD24c-m-negative cases. A multivariate progression analysis based on the Cox proportional hazard model demonstrated that CD24c-m expression is an independent prognostic factor for poor overall survival. CONCLUSION: The data from the present study suggested that CD24c-m expression is specific for tamoxifen-resistant breast cancer cases. CD24 should be subjected to comprehensive studies as a marker of resistance to tamoxifen treatment.

Antineoplastic Agents, Hormonal↗

PubMedDate not supplied

CD46 expression is indicative of shorter revival-free survival for ovarian cancer patients.

BACKGROUND: The membrane cofactor protein CD46 represents a complement inhibitor, which protects autologous cells from complement - mediated cytotoxicity. CD46 may exhibit the potential to protect tumor cells from the immune responses of the host. The present study aimed to evaluate the prognostic significance of CD46 expression in ovarian cancers. MATERIALS AND METHODS: The analyses were performed on 73 ovarian cancer samples. Immunohistochemical reactions were performed on paraffin sections of tumors using monoclonal antibodies directed against CD46. The immunohistochemical reactions and the clinical observations results were subjected to statistical analysis. RESULTS: Expression of CD46 was demonstrated in 60% of primary laparotomy cases and in 70% secondary cytoreduction cases. Kaplan-Meier analysis showed that a significantly shorter revival-free time was linked to cases with CD46 expression at PL (p= 0.01). CONCLUSION: Ovarian cancers manifest CD46 expression that is linked to a less favourable prognosis.

Female↗