Protein expression profile of primary human squamous cell lung carcinomas indicative of the incidence of metastases

The purpose of this investigation was to evaluate firstly whether different protein expression patterns exist in primary squamous cell lung carcinomas of patients with and without lymph node involvement and secondly, whether or not different patterns exist in tumours with positive lymph nodes. For this reason, formalin-fixed, paraffin-embedded specimens from 130 patients with squamous cell lung carcinomas were analyzed by immunohistochemistry. In a first step, proteins were selected which showed a relationship to lymph node involvement. The expression of JUN, ERBB2, MYC, cyclin D, PCNA, bFGF, VEGF and Hsp70 proteins revealed a positive correlation to lymph node involvement. In contrast, caspase-3, Fas ligand, Fas/CD95, and PAI showed an inverse correlation to lymph node involvement. In a second step, these parameters were further analyzed by hierarchical cluster analyses. The resulting clusters were correlated to patients with or without lymph node involvement. The data show that different protein expression patterns exist between primary squamous cell lung carcinomas with and without lymph node involvement and within carcinomas with lymph node involvement. The data suggest that various metastasis profiles exist. This revised version was published online in July 2006 with corrections to the Cover Date.     

Stability of morphine, morphine-3-glucuronide, and morphine-6-glucuronide in fresh blood and plasma and postmortem blood samples

The present study was designed to determine the stability of morphine and its glucuronides in spiked fresh blood and plasma from live individuals as well as in four authentic postmortem blood specimens for a time interval of up to six months. The samples were stored in glass vials at -20 degrees C, 4 degrees C, and 20 degrees C. Additionally, spiked samples were exposed to light through window glass and subjected to a forced-degradation study at 40 degrees C. Data were established using solid-phase extraction and high-performance liquid chromatography coupled to atmospheric pressure ionization mass spectrometry for isolation and quantitation, providing a sensitive and specific detection method for the parent drug in the presence of its glucuronide metabolites. Morphine and its glucuronide metabolites were found to be stable in both blood and plasma at 4 degrees C for the whole observation period. In postmortem blood the analytes were stable only when stored at -20 degrees C. The thermal decomposition of morphine and morphine-6-glucuronide in spiked blood and plasma could be interpreted using pseudo first-order kinetics. Photodegradation of morphine-3-glucuronide in plasma was consistent with a second-order reaction. In postmortem samples the degradation pattern differed completely from that observed in fresh blood and plasma. The elevated morphine levels observed were primarily due to postmortem hydrolysis of morphine glucuronides.     

Relationship of urokinase and urokinase receptor in non-small cell lung cancer to proliferation, angiogenesis, metastasis and patient survival

The purpose of the analysis was to ascertain whether the expression of components of the plasminogen activation system possesses an association with several biological parameters. The expression of urokinase (uPA) and urokinase receptor (uPAR) was analyzed in 137 non-small cell lung carcinomas by immunohistochemistry. No relationship could be observed between the proliferative activity of the carcinomas measured by flow cytometry and the expressions of uPA and uPAR. In addition, there was no association of the expressions of uPA or uPAR and vessel density (angiogenesis), neither any significant correlation between the expressions of uPA or uPAR and metastasis. The median survival was shorter for patients with uPA positive carcinomas than for those with uPA negative tumors (60 vs. 111 weeks; p=0.018). The relative risk estimate for patients with uPA positive tumors was increased by a factor of 1.8. In contrast, the expression of uPAR showed no significant correlation with overall survival.     

Detection of induced tumour-resistance to cyclophosphamide by the in vitro short term test

Treatment of the solid Walker carcinosarcoma of the rat for a period of one year with cyclophosphamide (40 mg/kg at each passage; tumours were transplanted weekly) resulted in the formation of a cyclophosphamide-resistant tumour cell line. Without further treatment, animals injected with non-treated or cyclophosphamide pre-treated cells survived for 10 days on an average. After therapy with cyclophosphamide (2 x 38 mg/kg), rats with non-treated tumour cells survived 22 days whereas those with pre-treated tumours survived only for 17 days. Tumour cells which were shown to be sensitive in vivo also exhibited a larger reduction in 3H-thymidine and 3H-uridine incorporation in the in vitro short term test after incubation with urine from cyclophosphamide-treated rats or with 4-hydroperoxy-cyclophosphamide. The resistance to cyclophosphamide which was detected in animal experiments with Walker carcinosarcoma can therefore also be observed using the in vitro short term test.     

Evaluation of the Sheffield system for identifying children at risk from unexpected death in infancy. Results from Birmingham and Newcastle upon Tyne

The "at birth" system which is used in Sheffield to identify children likely to die unexpectedly in infancy, was tested retrospectively in Birmingham (83 cases) and in Newcastle upon Tyne (56 cases). The discrimination between cases and age-matched controls was poor in both cities. Analysis of the 8 factors used in the system showed that only 2 maintained significant case/control differences in Birmingham and Newcastle. Further investigation showed that other factors from maternity records showed significant case/control differences in these cities. Although the system used in Sheffield would not be of use in a prospective prevention programme in either Newcastle or Birmingham, the possibility of evolving an "at risk" system which might apply more widely is discussed.     

Overexpression of P-glycoprotein and glutathione S-transferase-pi in resistant non-small cell lung carcinomas of smokers.

Ninety-four human non-small cell lung carcinomas (NSCLC) of previously untreated patients were analysed for the presence of P-glycoprotein (P-170) and glutathione S-transferase-pi (GST-pi) by means of immunohistochemistry. The expression of P-170 and GST-pi was compared with the results of doxorubicin resistance of the tumours in vitro and the smoking habits of the patients. A significant relationship between smoking habits of the patients and resistance of NSCLC was found (P = 0.007). Of the 72 tumours of smokers 57 (= 79%) were resistant, whereas of the 22 tumours of non-smokers only 11 (= 50%) showed resistance. Identical results were obtained when the analysis was restricted to patients with epidermoid lung carcinomas (P = 0.004). In contrast to these data, there exists no relationship between resistance and smoking for adenocarcinomas of the lung. Forty-two (= 58%) out of the 72 NSCLC of smokers expressed P-170, whereas out of 22 tumours of non-smokers only two tumours (= 9%) showed P-170 expression (P less than 0.0001). Similar results were obtained with epidermoid carcinomas (P = 0.004) and adenocarcinomas (P = 0.027). Fifty (= 69%) of 72 NSCLC of smokers revealed expression of GST-pi, whereas only nine (= 41%) of 22 tumours of non-smokers showed GST-pi expression (P = 0.015). Significant correlations also exist between resistance in vitro and expression of P-170 (P less than 0.0001) or expression of GST-pi (P less than 0.0001). Furthermore, a significant relationship between both proteins could be demonstrated (P less than 0.0001).     

Cellular pharmacology of mu-[1,2-bis(diphenylphosphino)ethane]bis[(1-thio-beta-D-gluco pyranosato-S)gold(I)]: a novel antitumor agent

SK&F 102912 (mu-[1,2-bis(diphenylphosphino)ethane]bis[(1-thio-beta-D- glucopyranosato-S)gold(I)], [(Autg)2(dppe)]) has shown reproducible and significant activity in transplantable murine tumor models and represents a structurally unique class of antineoplastic agents. A number of in vitro studies were performed to elucidate the cellular pharmacology of this gold-containing complex. [(Autg)2(dppe)] is a potent cytotoxic agent in vitro as demonstrated by its ability to inhibit the clonogenic capacity of a variety of tumor cell lines following a brief exposure to the drug. Cell-cycle analysis using HL-60 cells showed that low concentrations (2 microM) of [(Autg)2(dppe)] induced an S-phase block and higher concentrations induced a secondary block at the G1/S boundary. [(Autg)2(dppe)] had several effects on DNA metabolism and structure including preferential inhibition in cells of DNA synthesis (relative to RNA and protein synthesis) and the production of DNA single- and double-strand breaks as measured by alkaline elution. The cytoxic mechanism of this gold complex appears to be distinct from that of the monophosphine-gold complex auranofin.     

In vivo antitumor activity and in vitro cytotoxic properties of bis[1,2-bis(diphenylphosphino)ethane]gold(I) chloride

We have previously reported the cytotoxicity and antitumor activity of bis(diphenylphosphino)ethane (DPPE) and a variety of its transition metal complexes. During studies of the chemistry of a gold complex of this group [(AuCl)2(DPPE)], it was observed that this complex readily underwent ring closure on reaction with DPPE to form the tetrahedral complex [Au(DPPE)2]+. Various counterion forms (e.g., Cl-) of this cation were isolated and were found to exhibit a remarkably high stability in solution. Evaluation of [Au(DPPE)2]Cl in mice bearing i.p. P388 leukemia demonstrated that the compound produced an average of 87% increase in life span at its maximally tolerated dose (2-3 mumol/kg/day for 5 days). Activity was also seen in i.p. M5076 reticulum cell sarcoma (60% increase in life span) and s.c. mammary adenocarcinoma 16/c. Modest activity was evident in i.p. B16 melanoma and L1210 leukemia. A subline of P388 leukemia resistant to cisplatin was not cross-resistant to [Au(DPPE)2]Cl. In addition, combination therapy of [Au(DPPE)2]Cl and cisplatin against i.p. P388 demonstrated an advantage over single-agent therapy. In vitro studies of [Au(DPPE)2]Cl showed that the compound: is cytotoxic to tumor cell lines; is only minimally inhibited in its cytotoxic activity by the presence of serum; produces DNA protein cross-links and DNA strand breaks in cells; and inhibits macromolecular synthesis with a preferential inhibitory effect on protein synthesis relative to DNA and RNA synthesis. 31P nuclear magnetic resonance spectroscopy indicated that the compound is stable in the presence of serum proteins, thiols, or disulfides and that it reacts with Cu(II) resulting in the formation of a Cu(I)DPPE complex. The results of these in vivo and in vitro experiments suggest that the contrasting pharmacological profile of [Au(DPPE)2]Cl with respect to other gold(I) phosphine complexes may be related to both the kinetic stability of the complex and its stability in the presence of thiols.