O2*- and NO-associated mechanisms of selective action of redox-active cobalt complexes on tumor tissue.

AIM: To study the influence of redox-active cobalt(III) complex with tetradentate Schiff base and nicotinamide as an axial ligand on the rate of superoxide radical-anions generation and levels of NO in tumor and normal tissues of Lewis lung carcinoma bearing mice as well as activity of matrix metalloproteinases 2 and 9 (MMPs) in tumor. METHODS: The superoxide radical-anions formation and NO level in tissues were assessed by EPR method with the use of 1-hydroxy-2,2,6,6-tetramethyl-4-oxopiperidin and diethyldithiocarbomate spin traps, respectively. MMPs activities were determined by zymography in polyacrylamide gel. RESULTS: It was observed that the rate of superoxide radical-anions generation was selectively increased in tumor tissue (by a factor of 6-7) accompanied with the decrease of NO level (by a factor of 2) due to tested complex administration. Activities of MMPs in tumor were significantly decreased. CONCLUSION: It is supposed that the one of mechanisms of detected earlier antimetastatic effect of complex is based on its ability to induce the formation of high level of superoxide radical-anions selectively in the tumor tissue that results in the damage of its regulatory functions, in particular alteration in the regulation of NO-synthase, decrease of NO generation as well as activities of MMPs.

ADAM8 expression is associated with increased invasiveness and reduced patient survival in pancreatic cancer.

ADAM8 belongs to a family of transmembrane proteins implicated in cell-cell interactions, proteolysis of membrane proteins, and various aspects of carcinogenesis. In the present study, we aimed to evaluate the expression and function of ADAM8 in pancreatic cancer. ADAM8 mRNA levels were analysed by quantitative RT-PCR and correlated to patient survival. Immunohistochemistry was performed to localize ADAM8 in pancreatic tis-sues. Silencing of ADAM8 expression was carried out by transfection with specific siRNA oligonucleotides. Cell growth and invasion assays were used to assess the functional consequences of ADAM8 silencing. SELDI-TOF-MS was performed to detect the proteolytic activity of ADAM8 in pancreatic cancer cells. ADAM8 mRNA was significantly overexpressed in pancreatic ductal adenocarcinoma (PDAC) compared with normal pancreatic tissues (5.3-fold increase; P= 0.0008), and high ADAM8 mRNA and protein expression levels correlated with reduced survival time of PDAC patients (P= 0.048 and P= 0.065, respectively). Silencing of ADAM8 expression did not significantly influence pancreatic cancer cell growth but suppressed invasiveness. In addition, decreased proteolytic activity was measured in cell culture supernatants following silencing of ADAM8. In conclusion, ADAM8 is overexpressed in PDAC, influences cancer cell invasiveness and correlates with reduced survival, suggesting that ADAM8 might be a potential target in pancreatic cancer therapy.

Effects of radical oxygen species and NO: formation of intracellular hypoxia and activation of matrix metalloproteinases in tumor tissues.

AIM: To establish the association between the radical oxygen species (ROS) and NO levels in the tumor cells mitochondria, between cell hypoxia development and activation of matrix metalloproteinases-2 and -9. MATERIALS AND METHODS: Electron paramagnetic resonance (EPR) at room temperature and at the temperature of liquid nitrogen (77 degrees K), spin traps technology, enzymography in polyacrylamide gel were applied. RESULTS: Redox-centers in the respiration cascade of mitochondria have been revealed, multiple oxidative damage of which in breast and liver cancer tissues of experimental animals as well as in tumor tissue from patients with gastric cancer promote the development of cell hypoxia. Involvement of ROS and NO in activation of latent forms of matrix metalloproteinases in gastric tumor tissues has been shown. CONCLUSION: We hypothesize that superoxide radical-anions participate in development of cell hypoxia in tumors and surrounding normal tissues inducing activation of latent forms of matrix metalloproteinases.

High formation of superoxide anion and nitric oxide, and matrix metalloproteinases activity in vascular wall of rectal carcinoma vessels.

AIM: To study the relationship between the level of generation of reactive oxygen species (ROS) and nitric oxide (NO) and activity of matrix metalloproteinases (MMPs) MMP-2 and MMP-9 in the vessels isolated from rectal tumors and Arteria rectalis superior. METHODS: EPR at the room temperature and 77 degrees K, Spin Traps technology and zymography in polyacrylamide gels were applied. RESULTS: In the vessels isolated from rectal tumors and Arteria rectalis superior high levels of ROS, NO and formation of complexes of NO with FeS-proteins at the sites of electron-transporting chain of mitochondria have been detected. High activities of MMP-2 and MMP-9 in vascular wall were also observed. The direct positive correlation between the rate of NO generation and formation of complexes of NO with FeS-proteins as well as between ROS and NO formation and MMPs activities have been revealed. CONCLUSION: Altered oxidative equilibrium in mitochondria of cells in vascular wall promotes formation of cell hypoxia and its autocatalytic potentiation accompanied with activation of MMPs.

Hypoxia level and matrix metalloproteinases-2 and -9 activity in Lewis lung carcinoma: correlation with metastasis.

AIM: To investigate the relationship between tumor hypoxia in vivo, activity of matrix metalloproteinases (MMPs), and metastatic potential of tumor. MATERIALS AND METHODS: Lewis lung carcinoma (3LL) was used in this study. Total activity of MMP-2 and -9 in tumor was measured biochemically, tumor hypoxia level was assessed by (31)P NMR spectroscopy in tissue perchloric extracts. RESULTS: It was determined that hypoxia level in primary tumor has been concomitantly increasing along with tumor growth and correlated with metastasis level in lung. The positive correlation between hypoxia level and activities of MMP-2 and MMP-9 in primary tumor was registered. Moreover, the activity of MMP-2 and -9 in 3LL (primary tumor) directly correlates with metastasis level in lung. CONCLUSION: This study demonstrated that the growth of primary tumor is distinctly accompanied by an increase of tumor hypoxia level which positively correlates both with the activity of MMP-2 and -9 in primary tumor and metastatic efficiency.

Modifying effect of organocobalt complexes on the tumour response to anticancer treatments.

A new type of agents are proposed for combined cancer therapy. They are organocobalt (III) chelates containing a sigma-bounded organyl group and a mixed tridentate ligand derived from a Schiff base. These complexes generate free radicals due to the action of protons in physiological ranges of pH and temperature, and hence are conceivably capable of selectively attacking a malignant neoplasm that is slightly acidic and can be made even more so by introducing some means intensifying glycolysis. An in vivo examination was performed using transplanted rat tumours (Guerin and Walker 256 carcinomas, Sarcoma 45). The modifying effect of one of these complexes on the tumour response to cis-DDP, radiation and/or local hyperthermia was tested by means of tumour growth delay assay and local tumour control. The potentiating effect of the complex was maximal when it was administered 60-90 minutes prior to other agents (cisDDP, X-irradiation heat). The enhancement ratio was found to be ca. 2.0-4.0 for cisDDP and 2.0 for radiation. In conclusion, in our tumour models, an increase of the antitumour effect was obtained for conventional antitumour agents when they were supplemented with organocobalt complex. It can be hypothesised that DNA in tumour cells may be considered to be the main target for organocobalt complexes.

Tumour pH drop after cryotreatment and enhancement of hyperthermia antitumour effect.

Our previous study demonstrated a selective drop of pH, PO2 and blood flow in tumours under induced hyperglycaemia that enhanced the antitumour effect of hyperthermia. However, there are contraindications restricting the use of hyperglycaemia. Cryotreatment, as a method of altering tumour microphysiology similar to hyperglycaemia, was investigated. Guerin carcinoma pH was measured after cryotreatment (-15 to -20 degrees C at the tumour and normal tissue boundary; -110 to -120 degrees C at the cryoinstrument tip). It was determined that tumour pH immediately after complete tumour thawing (10 degrees C) was 6.08 (range 5.5-6.3), by 2 h it was 6.34 (range 6.2-6.4), by 4 h it was 6.42 (range 6.1-6.8) and by 24 h it was 6.54 (range 6.3-6.6). Complete tumour regression (%) and survival of rats with both the tumour regrowth and the partial tumour regression (days) after a single treatment were as follows: control -zero, 28 +/- 4; hyperthermia (2450 MHz, 43 degrees C, 60 min) -zero, 30 +/- 3; cryotreatment -14, 35 +/- 5; cryotreatment+hyperthermia -41, 46 +/- 5. It is planned to use combined cryotreatment+hyperthermia in a clinical trial for certain accessible tumours.

Induced hyperglycemia and tumor chemotherapy: experimental and clinical studies.

In order to clarify the influence of induced hyperglycemia upon antitumor effects of chemotherapy we studied some animal and human tumors. We observed that hyperglycemia enhances the antitumor effects of some cytostatic drugs several fold, due in part to the changes in the microphysiology induced by the hyperglycemia. Time doubling of some transplanted rat tumors and survival time of rats, rabbits and dogs bearing the transplanted and spontaneous tumors increased two to ten fold when the chemotherapy was used under hyperglycemia. Remission duration in patients with malignant brain glioma was enhanced by an average of six months; two years survival increased two-fold and the three years survival in patients with ovarian carcinoma in Stage III-IV increased two-fold; five years survival, 1.5-fold. The number of complications due to the combination of hyperglycemia and chemotherapy were not increased significantly. The problems of combination chemotherapy and hyperglycemia for oncologic patients' treatment are under discussion.