Inhibition of ADAM-17 more effectively down-regulates the Notch pathway than that of γ-secretase in renal carcinoma

Background

Our study is to research the effect of inhibited ADAM-17 expression through the Notch pathway in renal carcinoma.

Methods

Immunohistochemistry and western blot were used to examine the expression of ADAM-17 protein in renal cancer tissues. Proliferation and cell invasion of 786-o cells, as well as OS-RC-2 cells, after treatment with two different inhibitors of the Notch pathway, were examined by CCK-8 assay and Transwell assay, respectively. 786-o cell apoptosis was measured using the FCM test.

Results

ADAM-17 was highly expressed in RCC tissues. Compared with blocking γ-secretase, a known mechanism of impairing Notch, blockade of ADAM-17 more effectively down-regulated the expressions of Notch1 and HES-1 proteins. Similarly, we found that the ADAM-17 inhibitor, Marimastat, could more efficiently reduce renal cell proliferation and invasive capacity in comparison with the γ-secretase inhibitor DAPT when used at the same dose. Similar results were obtained when apoptosis of 786-o was measured.

Conclusion

Compared with γ-secretase, inhibition of ADAM-17 expression more effectively inhibits Notch pathway-mediated renal cancer cell proliferation and invasion. ADAM-17 may be a new target for future treatment of renal carcinoma.     

Up-regulation of γ-glutamylcysteine synthetase activity in melphalan-resistant human multiple myeloma cells expressing increased glutathione levels

Levels of intracellular glutathione (GSH) and the GSH-related enzymes -glutamylcysteine synthetase (-GCS) and -glutamyltranspeptidase (-GT) were measured in the melphalan-resistant human multiple myeloma cell line 8226/LR-5 and were compared to those measured in the drug-sensitive 8226/S and doxorubicin-resistant 8226/Dox40 cell lines. Both GSH and -GCS activity, the rate-limiting step in the de novo synthesis of GSH, were elevated by a factor of approximately 2 in the melphalanresistant 8226/LR-5 cells relative to the other two lines. -GT activity was not elevated significantly in the /LR-5 cells. Northern analysis with a probe specific for the large subunit of human liver -GCS identified two bands (3.2 and 4.0 kb), both of which were increased by a factor of 2–3 in the 8226/LR-5 line. Levels of -GCS mRNA expression were comparable in the /S and /Dox40 cell lines. Levels of -GT mRNA were similar in the /S and /LR-5 lines but were reduced in the /Dox40 cells. These data suggest that the increased GSH levels associated with resistance to melphalan in the 8226/LR-5 myeloma cells is attributable to up-regulation of -GCS. This observation is consistent with recent demonstrations of up-regulation of -GCS in melphalan-resistant prostate carcinoma cells and cisplatinum-resistant ovarian carcinoma cells, suggesting that increased expression of -GCS may be an important mediator of GSH-associated resistance mechanisms.     

Chemoradiation in cervical carcinoma: a must?

Cervical carcinoma is one of the most frequent gynecological malignancies. Literature shows that while the rate is exceedingly low in systematically screened populations, the incidence remains high because of large populations of at-risk women--particularly in underserved nations and in medically indigent subpopulations of Western nations--who are not screened. Recently, a series of randomized trials has demonstrated the possibility to dramatically improve the prognosis of these patients by using concurrent chemoradiation. In particular, concurrent chemoradiation represents a major treatment option in patients with bulky IB-IIA disease, IIB-IVA disease and resected IB-IIA disease with poor prognostic factors. However, further studies are necessary to optimize treatment schedules and particularly to define the best drug combination to be used during radiation therapy, improve patients selection by the analysis of anatomical (TNM stage) and non-anatomical (tumor oxygenation, genetic markers, tumor angiogenesis) prognostic factors, explore novel treatment strategies, such as use of neoadjuvant chemoradiation in locally advanced tumors, integration of antiangiogenetic therapies in chemoradiation schedules, use of supportive treatments aimed to overcome tumor hypoxia, to evaluate the possibility of 'cure' of locally recurrent tumors by chemoradiation and finally to define the best 'second-line' treatment for patients failing after chemoradiation with or without surgery.     

Expressions of the γ_2 chain of laminin-5 and secreted protein acidic and rich in cysteine in esophageal squamous cell carcinoma and their relation to prognosis

Previous studies have shown that the expressions of the γ2 chain of laminin-5 and secreted protein acidic and rich in cysteine (SPARC) play important roles in oncogenesis and the development of carcinoma. To assess the expressions of laminin-5 γ2 chain and SPARC in esophageal squamous cell carcinoma (SCC), and to clarify the prognostic significance of the expressions of laminin-5 γ2 chain and SPARC in esophageal SCC, we detected the expressions of laminin-5 γ2 chain and SPARC in cancer tissue and correspond...     

Role of zinc and ��2macroglobulin on thymic endocrine activity and on peripheral immune efficiency (natural killer activity and interleukin 2) in cervical carcinoma

Decreased natural killer (NK) activity as well as interleukin 2 (IL-2) are risk factors for the progression of cervical carcinoma. NK activity and IL-2 may be thymus controlled. Plasma levels of active thymulin, a zinc-dependent thymic hormone (ZnFTS), are reduced in cancer because of the low peripheral zinc bioavailability. Zinc and thymulin are relevant for normal immune functions. α2-Macroglobulin is an inhibitor of matrix metalloproteases (MMPs) against invasive tumour proliferation. Because α₂-macroglobulin has a binding affinity (K_d) for zinc that is higher than does thymulin, it may play a key role in immune efficiency in cancer. Plasma samples of 22 patients (age range 35–60 years) with locally advanced squamous cervical carcinoma and with FIGO stage Ib2–IIb were examined. They showed reduced active thymulin, decreased NK activity and IL-2 production, increased soluble IL-2 receptor (sIL-2R) and augmented α₂-macroglobulin in the circulation, whereas plasma zinc levels were within the normal range for age. Significant positive correlations were found between zinc or active thymulin and α₂-macroglobulin (r = 0.75, P< 0.01, r = 0.78, P< 0.01, respectively) in cancer patients. In vitro zinc increases IL-2 production from peripheral blood mononuclear cells (PBMCs) of cancer patients. These data suggest that an increase in α₂-macroglobulin, which competes with thymulin for zinc binding, may be involved in causing a thymulin deficit with a consequent decrease of IL-2 and NK cytotoxicity. Thus, physiological zinc treatment in cervical carcinoma maybe restores impaired central and peripheral immune efficiency. © 1999 Cancer Research Campaign     

Specific changes in the parameters of blood antioxidant status in patients with cervical carcinoma in the course of cancer therapyĭ

Parameters of the blood and plasma antioxidant systems were investigated at different stages of specific therapy for T3NxM0 cervical tumors in 23 patients to identify mechanisms of "relative" drug resistance. Treatment included preoperative telegammatherapy (10 Gy), aregional and regional lymphinfusion with low and high doses of cytostatics, respectively, complemented with endolymphatic injection of tocopherol acetate 100 ng. As a result, surgery after Wertheim became feasible in 14 out of 23 patients following external exposure to 30-40 Gy. Morphological examination detected metastases into the regional lymph nodes in 3 patients. The advantages offered by the treatment modality include adequate response to therapy as well as access to control of free-radical processes taking place in membranes of immunocompetent cells which in turn assures good clinical results.     

Is the International Federation of Gynecology and Obstetrics staging system for cervical carcinoma able to predict survival in patients with cervical carcinoma?: an assessment of clinimetric properties

BACKGROUND: The objective of this article was to assess the clinimetric properties of the International Federation of Gynecology and Obstetrics (FIGO) staging system of cervical carcinoma to determine whether it is an adequate prognostic tool for the survival of patients with cervical carcinoma. METHODS: The FIGO staging system for cervical carcinoma was evaluated with regard to item generation, item reduction, sensibility, reliability, and validity. RESULTS: Many statistically significant and clinically important variables have been omitted from the current staging system for cervical carcinoma. The item-reduction step for the formulation of the prognostic tool has not been described by the authors of the FIGO staging system, but a consensus process is assumed. There are no studies currently available to assess the reliability of interobserver and intraobserver variability in applying the staging system to patients with cervical carcinoma. A trial to assess the reliability of this tool is proposed by the authors. Although there are no prospective trials to assess the criterion validity of the FIGO staging system, there is enough literature to suggest that the staging system is not capable of discriminating with regard to patient survival within and between stages. CONCLUSIONS: The current FIGO staging system for cervical carcinoma does not fully meet the majority of methodologic criteria for a strong predictive tool. Developing an improved prognostic index containing a complete array of independently prognostic variables is suggested.     

Indomethacin overcomes doxorubicin resistance by decreasing intracellular content of glutathione and its conjugates with decreasing expression of γ-glutamylcysteine synthetase via promoter activity in doxorubicin-resistant leukemia cells

Drug resistance continues to be a serious problem in cancer therapy. We investigated whether indomethacin, which inhibits cyclooxygenases, is able to overcome doxorubicin resistance in K562/ADR leukemia cells. Indomethacin at 10 μM increased the cytotoxicity of doxorubicin and vincristine in K562/ADR cells. Intracellular glutathione content was elevated in K562/ADR cells. Indomethacin treatment decreased glutathione content and glutathione-conjugates in K562/ADR cells. Increased expression of γ-glutamylcysteine synthetase (γ-GCS) was observed in K562/ADR cells, but this expression was decreased by indomethacin treatment. The activity of the γ-GCS promoter from K562/ADR cells decreased after indomethacin treatment in MDA231 cells. These data strongly suggest that the cyclooxygenase inhibitor indomethacin increases the cytotoxicity of doxorubicin by decreasing the intracellular contents of glutathione and its conjugates with decreasing expression of γ-GCS by inhibiting γ-GCS promoter activity.     

Does histology influence prognosis in patients with early-stage cervical carcinoma?

BACKGROUND: The objective of this study was to examine the influence of histology on the outcome of patients with surgically treated, Stage IA-IB carcinoma of the uterine cervix. METHODS: All patient information was collected prospectively and was extracted subsequently from the University of Toronto cervical carcinoma surgery data base. Selection criteria for surgery were based on tumor size and were independent of histology. Patients with adenocarcinoma were separated into two groups: those with mucinous/endometrioid adenocarcinoma (M/E AC) and those with adenosquamous/clear cell adenocarcinoma (AS/CC AC). Statistical analysis used Wilcoxon rank tests, Mantel-Hanzel tests, chi-square tests, and Cox regression analyses. RESULTS: Between July 1984 and January 2000, 880 patients with Stage IA-IB cervical carcinoma underwent radical surgery, including pelvic lymphadenectomy, as the primary treatment. Two hundred fifty-five patients had M/E AC (29%), 81 patients had AS/CC AC (9%), and 544 patients had squamous cell carcinoma (SCC; 62%). Compared with patients who had SCC, patients with M/E AC had significantly more favorable prognostic characteristics: age (median, 39 years vs. 41 years; P < 0.03), depth of invasion (3.7 mm vs. 5.5 mm; P < 0.001), vascular space involvement (24% vs. 57%; P < 0.0001), Grade 2-3 tumor (40% vs. 78%; P < 0.0001), and pelvic lymph node metastases (4% vs. 8%; P < 0.04), respectively. Characteristics among patients with AS/CC AC tended have values similar to the median values for patients with SCC (or intermediate between the values for patients with M/E AC and the values for patients with SCC): age (38 years), depth of invasion (6 mm), vascular space involvement (40%), Grades 2-3 (70%), and pelvic lymph node metastases (6%). The 2-year and 5-year recurrence free survival rate was similar between patients with M/E AC and patients with SCC (95% vs. 94% and 90% vs. 90%, respectively); however, both were significantly superior to the rates for patients with AS/CC AC (2-year recurrence free survival rate: 86%, P < 0.03; 5-year recurrence free survival rate: 81%, P % 0.03). There were no differences in the pattern of first recurrence by histology. CONCLUSIONS: Patients with surgically treated Stage IA-IB cervical carcinoma with M/E AC and SCC histology have a similar prognosis. For patients with disease with AS/CC AC histology, the current results and the literature indicate that patients with uncommon histologies have an inferior recurrence free survival rate. Although the optimal therapy for these patients remains undefined, there is no obvious rationale for altering the treatment strategies from those currently employed for patients with M/E AC and SCC.     

Inhibition of γ-synuclein (SNCG) expression in breast cancer MDA-MB231 cell line

Objective  

The aim of the study was to evaluate the inhibition of different chemotherapy drugs on Γ-synuclein (SNCG) positive expression of breast cancer cell line MDA-MB231, and the effects on cell cycle and apoptosis, and to explore the related mechanism as well.     

Oral therapy with proteolytic enzymes decreases excessive TGF-? levels in human blood

Therapy with oral proteolytic enzymes (OET) with combination drug products containing papain, bromelain, trypsin, and chymotrypsin has been shown to be beneficial in clinical settings such as radiotherapy-induced fibrosis, bleomycin pneumotoxicity and immunosuppression in cancer, all of which are nowadays known to be accompanied by excessive transforming growth factor-? (TGF-?) production. It has been demonstrated that proteolytic enzymes reduce TGF-? levels in serum by converting the protease inhibitor a2 macroglobulin (a2M) from the "slow" form into the "fast" form, whereby the "fast" form binds and inactivates TGF-? irreversibly. In this study we have investigated the effect of OET on the concentration of TGF-?1 in serum of patients with rheumatoid arthritis (RA) (n=38), osteomyelofibrosis (OMF) (n=7) and herpes zoster (HZ) (n=7). Seventy-eight healthy volunteers served as controls. TGF-?1 levels in serum were assessed by enzyme-linked immunosorbent assay (ELISA). We have demonstrated that in healthy volunteers and in patients there exists a correlation between active and latent TGF-?1 in serum (r=0.8021; P<0.0001). Treatment with OET had no significant effect on TGF-?1 concentration in healthy volunteers or patients with a normal level of TGF-?1. In patients with elevated TGF-?1 concentration (>50 ng/ml serum), OET reduced TGF-?1 in RA (P<0.005), in OMF (P<0.05) and in HZ (P<0.05). Conclusion: These results support the concept that OET is beneficial in diseases characterized in part by TGF-?1 overproduction.     

No synergistic activity of epirubicin and interferon-α2b in the treatment of hepatocellular carcinoma

Single-agent activity for anthracyclines reflected by response rates of 10%–30% has been reported in patients with advanced hepatocellular carcinoma (HCC). Preclinical data indicate that -interferon could enhance the cytotoxic activity of the anthracycline Adriamycin or its analog epirubicin. In a phase I/II study, 31 patients with biopsy-proven inoperable HCC were treated with interferon-2b given s. c. at a dose of 3×10⁶ units/m² per day for 5 days per week plus weekly epirubicin given at 25 mg/m² as an i. v. bolus. The protocol called for 4 consecutive weeks of treatment followed by 1 week off treatment. In all, 15 patients had been previously treated; 6 patients had failed hormonal therapy (tamoxifen), 5 patients had failed prior anthracycline treatment, and 4 patients had received chemoembolization of the tumor and had subsequently progressed. A total of 30 patients were evaluable for response. In all, 1 patient (3%) achieved a partial response for 8+ months and 11 patients (35%) achieved stabilization of disease. Six patients had a fall in alpha-fetoprotein (AFP) values of >50% during therapy. The median survival for all patients was 9.5 months (range, 3–34+ months). The main side effects were hematological toxicity and fever, both of which were considered tolerable. As an indicator of the immunostimulatory effects of interferon, an elevation in serum markers of inflammation [C-reactive protein (CRP), ₂-microglobulin] was found in 15%–20% of patients. All patients had measurable Mx protein production during therapy, but these effects were not correlated to the clinical response. The clinical response rate achieved in this trial indicates that the combination of interferon and epirubicin, at least when used on the schedule reported herein, is not superior to treatment with either agent alone for patients with advanced HCC. However, single patients achieved a prolonged progression-free interval (8–10+ months) on this therapy, and it may therefore be an option for patients who have failed prior hormonal or single-agent anthracycline therapy.     

Oestrogen receptors and peripheral serum levels of oestradiol-17β in patients with mammary carcinoma

Oestradiol receptors were determined by isoelectric focusing in cytosol from breast cancer tissue from 12 pre- and 70 postmenopausal patients. Simultaneous determination of the peripheral serum levels of oestradiol-17β was carried out in the 12 premenopausal and 57 of the postmenopausal patients. At oestradiol-17β levels below 150 pM the receptor values ranged from below 0.01 to 7.19 fmole bound [³H] oestradiol-17β/μg of DNA. At oestradiol-17β levels above 150 pM only low receptor values (<0.25 fmole bound [³H] oestradiol-17β/μg of DNA) were recorded. The results indicate an influence of the endogenous oestrogen on the receptor assay probably due to blockade of the hormone binding site. It is proposed that an endocrine evaluation of the patient should be performed together with the oestrogen receptor assay.     

UFT and its metabolite γ-butyrolactone (GBL) inhibit angiogenesis induced by vascular endothelial growth factor in advanced cervical carcinoma

OBJECTIVE: The aim of this study was to evaluate the potential role of UFT and its metabolite gamma-butyrolactone (GBL) for inhibition of angiogenesis induced by vascular endothelial growth factor (VEGF) in advanced cervical carcinoma by the determination of serum GBL and VEGF, and by immunohistochemical staining to assess VEGF protein expression, before and after UFT therapy. METHODS: The subjects were 35 patients with an advanced cervical carcinoma and five healthy volunteers between 2002 and 2003 at Hiroshima University Hospital, under informed consent. The patients received two courses of oral fluoropyrimidine (UFT) therapy at a dose of 600 mg/day for 5 and 2 days off treatment. Serum GBL and VEGF was measured before and after UFT therapy by the gas chromatography mass spectrometry and ELISA-kit in 22 patients and five healthy volunteers, respectively. Immunohistochemical detection of VEGF protein was done in 35 cervical cancers. Results The mean serum GBL level before and after UFT therapy was 21.9 +/- 2.3 and 79.3 +/- 6.2 ng/ml, respectively, and it was significantly increased after UFT administration (P < 0.0001). The mean serum VEGF level before and after UFT therapy was 95.3 +/- 28.1 and 67.5 +/- 11.2 pg/ml, respectively, and it was decreased by UFT administration. In 20 out of 33 (66.6%) patients who were detected with VEGF protein, VEGF protein expression was decreased by UFT therapy. The Delta GBL value (GBL after UFT--GBL before UFT therapy) showed a significant inverse correlation with Delta VEGF value (VEGF after therapy--VEGF before therapy) (r2 = 0.940). CONCLUSIONS: Our findings suggest that UFT and its metabolite GBL inhibit angiogenesis induced by VEGF to have an antitumor effect on cervical cancer.     

Inhibition of invasion-associated thromboxane synthase sensitizes experimental gliomas to γ-radiation

The invasion- and apoptosis-associated thromboxane synthase gene encoding an enzyme of the arachidonic acid pathway has been implicated in glioma progression. Furegrelate, a specific inhibitor of thromboxane synthase, blocks cell motility, induces apoptosis and increases sensitivity to drug induced apoptosis in human glioma cells in vitro. The impact of furegrelate on the sensitivity of human glioma cells to gamma-irradiation was analyzed using colony formation assay in vitro and an orthotopic mouse model in vivo. Pre-treatment of glioma cells with furegrelate increases radiation sensitivity of cultured glioma cells. Treatment of experimental gliomas with suboptimal doses of radiation and furegrelate results in a significant decrease in tumor volumes compared to untreated controls. Thus, the specific thromboxane synthase inhibitor furegrelate increases death response induced by gamma-radiation in glioma cells in vitro and sensitizes experimental gliomas to radiation treatment in vivo.     

Therapeutic Effect of γ-Secretase Inhibition in Kras(G12V)-Driven Non-Small Cell Lung Carcinoma by Derepression of DUSP1 and Inhibition of ERK

Here, we have investigated the role of the Notch pathway in the generation and maintenance of Kras(G12V)-driven non-small cell lung carcinomas (NSCLCs). We demonstrate by genetic means that γ-secretase and RBPJ are essential for the formation of NSCLCs. Of importance, pharmacologic treatment of mice carrying autochthonous NSCLCs with a γ-secretase inhibitor (GSI) blocks cancer growth. Treated carcinomas present reduced HES1 levels and reduced phosphorylated ERK without changes in phosphorylated MEK. Mechanistically, we show that HES1 directly binds to and represses the promoter of DUSP1, encoding a dual phosphatase that is active against phospho-ERK. Accordingly, GSI treatment upregulates DUSP1 and decreases phospho-ERK. These data provide proof of the in?vivo therapeutic potential of GSIs in primary NSCLCs.