[Management of chronic pain using extended release tilidine : Quality of life and implication of comedication on tilidine metabolism]. / Management von chronischem Schmerz mit retardiertem Tilidin : Lebensqualität und Bedeutung der Komedikation für den Tilidinmetabolismus.

BACKGROUND AND OBJECTIVES: The synthetic opioid tilidine is often used in chronic pain treatment. However, the activation via metabolism in patients with concomitant medication and reduced liver or kidney function is not thoroughly investigated. We therefore studied pain treatment efficacy, health-related quality of live and the metabolism of tilidine in patients with chronic pain. METHODS AND MATERIALS: In all, 48 patients, who were on a stable dose of oral prolonged release tilidine for at least 7 days, were included in this observational multicenter study. Liver and kidney function were assessed in routine blood samples, concentrations of tilidine, nortilidine and bisnortilidine were determined using a validated LC/MS/MS method. Comedication was registered and patients experience with regard to quality of life, pain, gastrointestinal symptoms and adverse events was assessed in standardised questionnaires. RESULTS: On average a daily dose of 180 mg tilidine was taken. Dose normalized plasma concentrations of the active metabolite nortilidine ranged between 1.6 ng/ml and 76.5 ng/ml (mean 29.2 ± 25.1 ng/ml). Ratios between tilidine and nortilidine were on average 0.28 (median = 0.13, standard deviation = 0.67). Patients were on 1 to 14 different concomitant medications. About 66% of the patients had sufficient pain treatment. Almost no opioid-induced constipation was observed. Only few patients had decreased kidney or liver function which did not result in elevated nortilidine concentrations. CONCLUSION: Pain treatment using tilidine resulted in variable nortilidine concentrations which are obviously not strongly influenced by comedication or reduced liver or kidney function. Only a few side effects were observed with almost no opioid-induced constipation.

Expression of the retinoblastoma tumor suppressor gene (RB-1) in acute leukemia.

In this report we review current studies concerning the RB-1 gene expression in acute leukemias. The RB-1 gene was analyzed in several studies by protein-, RNA and DNA-techniques in acute lymphoblastic leukemia (ALL) as well as in acute myelogenous leukemia (AML). The frequency of RB-1 inactivation in ALL-patients ranged between 30% and 64% in several studies. Structural abnormalities of the RB-1 gene were reported in 18% of ALL-patients and in 27% of Philadelphia chromosome-positive ALL, respectively. The proportion of AML-patients with absent RB-1 protein expression ranged between 19% and 55%. Structural RB-1-abnormalities in AML were predominantly reported in leukemias with monocytic differentiation. Furthermore, the prognostic value of an abnormal RB-1 gene expression was also estimated in some studies. In childhood ALL RB-1 inactivation was reported to have prognostic significance while in contrast, in another study on adults no prognostic value of RB-1 was found. In 4 out of 5 documented studies AML-patients with RB-1 inactivation generally had a poorer prognosis. In conclusion, RB-1 inactivation is frequently observed in acute leukemia. The prognostic value of low RB-1 expression is controversial but the majority of published studies found low RB-1 expression to be a negative prognostic predictor, in acute leukemia.

Spontaneous apoptosis in ovarian cancer: an unfavorable prognostic factor.

Apoptosis and cell proliferation play important roles in the cellular response to chemotherapy, and may have prognostic value. The percentage of apoptotic cells [apoptotic index (AI)] was evaluated in ovarian carcinomas of 25 patients by the terminal desoxynucleotidyl transferase-mediated dUTP-nick end labeling (TUNEL) technique. All patients were surgically treated and received postoperatively chemotherapy consisting of cytoxan and cisplatin or carboplatin. As a marker of proliferation the mRNA expression of histone H3 in tumor tissue was determined. In addition, tumor vascularity was assessed by immunohistochemistry and factor VIII. Patients with high AI had significantly shorter survival times (p=0. 0001) or recurrence-free intervals (p=0.004) than patients with low AI. No significant relationship was found between AI and histone H3 mRNA expression, however, an inverse correlation of AI with microvessel density was detected. Tumors with high AI had significantly lower microvessel count than tumors with low AI (p=0. 002). The obtained data suggest that AI can be predictive of treatment outcome in ovarian cancer.

Prognostic implications of cyclins (D1, E, A), cyclin-dependent kinases (CDK2, CDK4) and tumor-suppressor genes (pRB, p16INK4A) in childhood acute lymphoblastic leukemia.

Immunohistochemistry was used to analyze samples of 40 newly diagnosed childhood acute lymphoblastic leukemias (ALL) for their expression of cyclins (D1, E, A), cyclin-dependent kinases (cdk2, cdk4) and tumor-suppressor genes (pRb, p16INK4A), in order to discover whether or not the expression of these various proteins may be of prognostic relevance for the survival of children with ALL. Patients with ALL who were strongly positive for cyclin D1 had a lower probability of remaining in first continuous remission than ALL patients who were negative or weakly positive for this trait. There was also a significant correlation between expression of cyclin D1 and frequency of recurrence. For cyclin E and cyclin A, in contrast, there was no difference in the duration of relapse-free-intervals or the frequency of recurrence in patients. Children with cdk4-positive ALL had a lower probability of remaining in first continuous remission than children with cdk4-negative ALL. No prognostic relevance was found for cdk2. Patients with ALL who expressed pRb had a higher probability and patients who expressed p16 a lower probability of remaining in first continuous remission, but the results were not statistically significant. This investigation demonstrated that cyclin D1 and cdk4 were the most important prognostic factors for children with ALL, and that the combination of them showed the strongest prognostic relevance.

Expression of lung resistance-related protein (LRP) in initial and relapsed childhood acute lymphoblastic leukemia.

Thirty-eight children with initial and 25 children with relapsed lymphoblastic leukemia (ALL) were analyzed for the expression of LRP (lung resistance protein or p110 major vault protein) using immunocytochemistry. LRP expression was found in 18 of 38 (47%) children with initial ALL and in 17 of 25 (68%) children with relapsed ALL. Children with initial ALL and without LRP expression had significantly longer relapse-free intervals than patients with LRP expression.

Deoxycytidine kinase mRNA expression in childhood acute lymphoblastic leukemia.

Initial and relapsed childhood acute lymphoblastic leukemias (ALL) were investigated for mutations and expression of deoxycytidine kinase (dCK). dCK mediating toxicity of 1-beta-D-arabinofuranosylcytosine was analyzed semiquantitatively by RT-PCR and by single-strand conformation polymorphism. We found that patients with initial ALL experienced more frequently a relapse if dCK expression was low or absent. No mutations were found in initial and relapsed ALL.

Prognostic value of basic fibroblast growth factor and its receptor (FGFR-1) in patients with non-small cell lung carcinomas.

Tumour specimens of 206 patients with untreated non-small cell lung carcinomas (NSCLC) were analysed immunohistochemically for the expression of the basic fibroblast growth factor (bFGF) and for its receptor (FGFR-1, Flg). Seventy of the tumours showed weak expression, 109 moderate and 27 high expression of bFGF. Thirty-eight tumours had low expression of FGFR-1, 116 had moderate and 52 cases high expression. Patients with high FGFR-1 expression had significantly shorter survival times than patients with weak or moderate expressions (P < 0.05), but there was no significant correlation between bFGF expression and patient survival. The results of the multivariate analysis demonstrated that FGFR-1 in the presence of stage is not an independent prognostic factor.

Green tea catechins (EGCG and EGC) have modulating effects on the activity of doxorubicin in drug-resistant cell lines.

The chemopreventive effect of polyphenols from green tea [e.g. (-)-epigallocatechin gallate (EGCG) and (-)-epigallocatechin (EGC)] against cancer has been demonstrated in several studies. The aim of this investigation was to prove whether these compounds modulate the activity of antineoplastic drugs. Therefore, the influence of EGCG and EGC was tested on doxorubicin-resistant murine sarcoma (S180-dox) and human colon carcinoma (SW620-dox) cell lines. Both substances showed a sensitizing effect on the cell lines if they had been treated with doxorubicin. These results suggest that protein kinase C may be inhibited by EGCG and EGC, and this may lead to a reduced expression of some drug resistance related proteins.

Messenger RNA expression of resistance proteins and related factors in human ovarian carcinoma cell lines resistant to doxorubicin, taxol and cisplatin.

Doxorubicin- (OAW-dox, SK-OV-dox), taxol- (OAW-tax, SK-OV-tax) and cisplatin- (SK-OV-cis) resistant cells derived from the parental OAW-42 and SK-OV-3 cell lines were established. OAW-42 sublines showed high resistance, the SK-OV-3 sublines only low resistance. OAW-42 sublines showed a cross-resistance profile typical of multidrug resistance (MDR). The sublines of SK-OV-3 showed a cross-resistance profile different from the OAW-42 sublines. The mRNA expression of several resistance proteins and related factors was analyzed. An overexpression of P-glycoprotein 170 (P-170), glutathione-S-transferase-pi (GST-pi), thymidylate synthase (TS), glutathione peroxidase (GP) and c-jun was found in OAW-dox and OAW-tax cells. Additionally, OAW-tax cells expressed a higher mRNA level of protein kinase Cbeta2. DNA analysis revealed a 2-fold gene amplification of P-170, whereas the genes for GST-pi, TS and GP were not amplified. SK-OV-dox and SK-OV-tax cells showed a decreased level of histone 3 (H3) and TS mRNA. This shows that the sublines of OAW-42 developed resistance by co-expression of several resistance-related proteins and proto-oncogenes whereas the sublines of SK-OV-3 expressed resistance by decreased expression of the proliferation-dependent proteins H3 and TS.

Cyclin A is associated with an unfavourable outcome in patients with non-small-cell lung carcinomas.

Specimens of formalin-fixed, paraffin-embedded non-small-cell lung carcinomas (NSCLCs; n = 187) were analysed immunohistochemically for expression of cyclin A. The analysis was intended to determine whether cyclin A has additional prognostic value for predicting patients' survival and drug response. Of the 187 NSCLCs, 141 cases (75%) showed expression of cyclin A. Patients with cyclin A-positive carcinomas had significantly shorter median survival times than patients with cyclin A-negative carcinomas (79 vs 129 weeks, P = 0.045). Similar results were obtained with more homogeneous groups of patients: patients with only T3 tumours, patients with epidermoid carcinomas and patients with lymph node involvement. The clinical parameters (age, stage, histology, extent of tumour size, lymph node involvement) had no influence on expression of cyclin A. A direct correlation between cyclin A and the proportion of S-phase cells (P = 0.08) and an inverse relationship between cyclin A and the proportion of G0/G1-phase cells (P = 0.04) were found. Furthermore, a significant correlation between the expression of cyclin A and the response of NSCLC to doxorubicin in vitro was detected (P = 0.026).

Association of vascular endothelial growth factor expression with tumor cell proliferation in ovarian carcinoma.

Vascular endothelial growth factor (VEGF) is a potent angiogenic factor that may also function as an autocrine growth regulator. Thirty-one ovarian carcinomas were investigated for mRNA expression of VEGF and of a proliferation-dependent gene (histone H3) using slot-blot analysis. Tumor vascularity was assessed by immunohistochemistry and factor VIII. All tumors were demonstrated to express VEGF and histone H3, though to various degrees. There was a good correlation between VEGF mRNA values and histone H3 mRNA values (r = 0.71, p < 0.05). No correlation was found between tumor cell proliferation and tumor vascularity. There was no significant difference in relapse-free interval or overall survival between tumors with low and high VEGF expression. The close correlation of VEGF expression with tumor cell proliferation in this study raises the possibility of autocrine stimulation of ovarian carcinoma.

Angiogenic growth factors and their receptors in non-small cell lung carcinomas and their relationships to drug response in vitro.

Tumor specimens of non-small cell lung carcinomas (NSCLC) from previously untreated patients (n = 153) were analysed immunohistochemically for expression of vascular endothelial growth factor (VEGF), VEGF-receptor (Flt-1), basic fibroblast growth factor (bFGF) and FGF-receptor (FGFR-1, (Flg). Expression of the proteins was compared with the in vitro response of the tumors against doxorubicin. The data clearly demonstrate that a significant relationship exists between VEGF (p < 0.001) and Flt-1 expression (p < 0.01) and drug response. The expression of VEGF and Flt-1 was lower in resistant than in sensitive tumors. In contrast, no significant interrelationship was found between expression of bFGF and response to doxorubicin. However, there exists a significant correlation between the expression of FGF-receptor (FGFR-1) and the response of the carcinomas to doxorubicin (p < 0.05). Expression of FGFR-1 was more frequently negative or weak in resistant and more frequently moderate or high in sensitive NSCLC. The data from this investigation clearly demonstrate that a significant interrelationship exists between the expression of VEGF, VEGF-receptor Flt-1 and FGF-receptor, FGFR-1 and the response of NSCLC to doxorubicin in vitro.

Apoptotic index, Fas and bcl-2 in initial and relapsed childhood acute lymphoblastic leukaemia.

Seventy-two samples with initial and 23 samples with relapsed childhood acute lymphoblastic leukaemia (ALL) were investigated for apoptotic index (AI) and for Fas expression. AI was determined by DNA-fragmentation using deoxynucleotidyl terminal transferase and Fas expression by immunocytochemistry. bcl-2 mRNA expression was measured in 50 initial and 20 relapsed ALL. The patients were discriminated in groups with low and high AI, Fas-protein expression and bcl-2mRNA expression by the mean value. AI was higher in relapsed than in initial ALL. The mean survival was significantly higher in patients with low AI (p= 0.034). This was also true for the relapse-free interval, however, this result was borderline significant (p= 0.064). AI was directly correlated with expression of Fas and inversely correlated with bcl-2 mRNA expression. These results suggest that Fas and - with limitations - bcl-2 may influence the apoptotic process in childhood ALL and that enhanced apoptotic activity predicts poor prognosis.

Expression of resistance-related proteins in tumoral and peritumoral tissues of patients with lung cancer.

Twenty tumoral and peritumoral tissues from patients with lung cancer were analyzed immunohistochemically for the drug resistance-related proteins P-glycoprotein (P-170), topoisomerase II (Topo-II), glutathione S-transferase-pi (GST-pi), metallothionein (MT), heat shock protein-70 (HSP-70) and the putative regulators of resistance (ErbB1, Fos and Jun). Protein expression of Topo-II, GST-pi, MT, HSP-70, ErbB1, Fos and Jun was elevated in tumor tissue in comparison to normal tissue. The different expression of the proteins between tumoral and normal tissues was statistically significant for Topo-II (P = 0.05), MT (P = 0.03), and HSP-70 (P = 0.01), whereas ErbB1 showed a borderline significance. The expression of the proteins was frequently increased in smokers in comparison to non-smokers. In general, the increase of the proteins of smokers corresponded in tumoral and non-tumoral tissue. Different expression was only found with MT and HSP-70 which were higher in tissues of smokers.

Expression and prognostic value of the retinoblastoma tumour suppressor gene (RB-1) in childhood acute lymphoblastic leukaemia.

In a retrospective analysis, acute lymphoblastic leukaemia (ALL) blast cells of 102 children were investigated for the expression of the retinoblastoma susceptibility (RB)-1 gene at mRNA level by dot blot hybridization and semiquantitative RT-PCR. 56 patients were analysed by dot blot hybridization and 35 representative patients out of this group by semiquantitative RT-PCR. Two additional groups of patients (23 patients with initial and 23 patients with relapsed ALL) were also investigated by semiquantitative RT-PCR. RB-1 gene expression was detectable in all investigated ALL at different levels. According to the relative mRNA expression the patients were discriminated by the median value in groups with low or high RB-1 expression. The Kaplan-Meier estimates showed that patients with low RB-1 expression had a lower probability of remaining in first remission (P = 0.03) and a significantly higher risk to succumb to their disease (P = 0.03). Furthermore, the comparison of the results between initial and relapsed ALL showed that the relapses had significantly lower RB-1 mRNA expression (P = 0.02). The overall survival of the patients was shorter in both groups when RB-1 gene expression was low. A multivariate analysis, including age, sex, immunological subtype, initial white blood cell count and RB-1 expression, identified RB-1 as an independent prognostic predictor (P = 0.017) in addition to the initial white blood cell count (P = 0.00001). In conclusion, low RB-1 expression is an unfavourable prognostic predictor in initial and relapsed childhood ALL. The RB-1 gene expression in relapsed ALL is significantly lower than in initial ALL.

Microvessel density, expression of proto-oncogenes, resistance-related proteins and incidence of metastases in primary ovarian carcinomas.

Relationships between the incidence of metastatic spread and microvessel density, expression of proto-oncogene products, or expression of resistance-related proteins were investigated in human ovarian carcinomas by immunohistochemistry. Ovarian carcinomas with a high microvessel density showed a significantly increased formation of metastases (P = 0.005). Tumors with positive immunoreactivity of c-jun and c-myc products had a higher metastatic spread; however, these results were not statistically significant. A marginally significant correlation existed between the expression of erbB1 (EGFR) and metastatic spread (P = 0.05). No significant relationship was found between the expression of the resistance-related proteins P-glycoprotein or glutathione S-transferase-pi and the incidence of metastases. Furthermore, no correlation was detected between expression of the heat shock protein 70 and the occurrence of metastases.

Retinoblastoma (Rb) protein expression and resistance in squamous cell lung carcinomas.

The purpose of this study was to prove the value of Rb protein expression as a prognostic factor for patients with squamous cell lung carcinoma. The expression of Rb in 75 carcinomas was analyzed immunohistochemically and the resistance to doxorubicin was investigated in vitro. Rb-negative carcinomas were more frequently resistant than Rb-positive carcinomas (p<0.05). A trend for a correlation between the expression of Rb and the expression of glutathione S-transferase-pi was detectable (p=0.08). Patients with Rb-positive carcinomas had a trend for better prognosis, but this result was not statistically significant.

Expression of heat shock proteins, glutathione peroxidase and catalase in childhood acute lymphoblastic leukemia and nephroblastoma.

In this study we analyzed the mRNA expression of the heat shock proteins 27 and 70, and the expression of the radical scavenging enzymes catalase and glutathione peroxidase (GPX) in childhood acute lymphoblastic leukemia (ALL, n = 54) and in nephroblastoma (n = 34). We found a significant positive correlation between both heat shock proteins and also between glutathione peroxidase and both heat shock proteins in ALL and nephroblastoma. There was also a significant correlation between catalase and glutathione peroxidase detectable. Furthermore, we investigated whether the expression of the heat shock proteins and the antioxidant enzymes glutathione peroxidase and catalase have implications in the clinical outcome in ALL. However, we found no significant correlation between the expression of these proteins and relapse rate, the relapse free intervals or the overall survival.

Messenger RNA expression of resistance factors in human tumor cell lines after single exposure to radiation.

Resistance of tumor cells to chemotherapeutic drugs can not only be caused by treatment with antineoplastic agents but also by radiotherapy. The aim of this study was to analyze whether ionizing radiation can influence the mRNA expression of proteins which have been found to be involved in drug resistance of tumor cells. Human tumor cell lines (MCF-7, LXF and Sk-Mel) were treated with single doses of irradiation (5, 10 and 20 Gy). The expression of the resistance related proteins glutathione S-transferase-pi (GST-pi), topoisomerase II alpha (Topo II), thymidylate synthase (TS), O6-methylguanine-DNA-methyltransferase (MGMT), P-glycoprotein (Pgp), glutathione peroxidase (GPX) multidrug resistance-associated protein (MRP) and also of the heat-shock protein 70 (HSP 70) were determined at the mRNA level during the time interval from 1.5 to 72 h post-irradiation and compared with their corresponding controls. We also examined whether a relationship exists between these proteins and the proliferative activity (histone 3, Ki-67, statin) of the cells. We found that exposure of MCF-7, LXF and Sk-Mel cells to ionizing radiation increases the expression of the mRNA of GST-pi. Topo II, TS, HSP 70 and proliferation markers were also altered by exposure to ionizing radiation, but there was no common response of the three cell lines. No significant changes were observed in the expression of MGMT, Pgp, GPX and MRP after radiation treatment. Drug resistance tests revealed that irradiated MCF 7 cells were less sensitive to doxorubicin than non-irradiated control cells. Our results indicate that ionizing irradiation modifies the expression of some proteins involved in drug resistance and the response of MCF 7 cells to doxorubicin and may, therefore, play a role in clinical drug response.