Application of C60 Fullerene-Doxorubicin Complex for Tumor Cell Treatment In Vitro and In Vivo.

Development of nanocarriers for effective drug delivery to molecular targets in tumor cells is a real problem in modern pharmaceutical chemistry. In the present work we used pristine C60 fullerene as a platform for delivery of anticancer drug doxorubicin (Dox) to its biological targets. The formation of a complex of C60 fullerene with Dox (C60 + Dox) is described and physico-chemical characteristics of such complex are presented. It was found that Dox conjugation with C60 fullerene leads to 1.5-2-fold increase in Dox toxicity towards various human tumor cell lines, compared with such effect when the drug is used alone. Cytotoxic activity of C60 + Dox complex is accompanied by an increased level of cell produced hydrogen peroxide at early time point (3 h) after its addition to cultured cells. At the same time, cellular production of superoxide radicals does not change in comparison with the effect of Dox alone. Cytomorphological studies have demonstrated that C60 + Dox complexes kill tumor cells by apoptosis induction. The results of in vivo experiments using Lewis lung carcinoma in mice confirmed the enhancement of the Dox toxicity towards tumor cells after drug complexation with C60 fullerene. The effect of such complex towards tumor-bearing mice was even more pronounced than that in the in vitro experiment with targeting human tumor cells. The tumor volume decreased by 2.5 times compared with the control, and an average life span of treated animals increased by 63% compared with control. The obtained results suggest a great perspective of application of C60 + Dox complexes for chemotherapy of malignant tumors.

THE ROLE OF REACTIVE OXYGEN SPECIES IN TUMOR CELLS APOPTOSIS INDUCED BY LANDOMYCIN A.

Landomycin A (LA) is a new antitumor antibiotic of angucycline group, possessing high antitumor activity against cancer cells of different origin, which induces early apoptosis in target cells. It was shown that under LA action the level of reactive oxygen species (ROS) in human T-leukemia cells had increased 5.6 times in comparison to control already at the 1st hour after the addition of studied antibiotic to the culture medium. At the 6th hour after incubation of cells with LA the nucleosomal DNA cleavage, chromatin condensation and nucleus fragmentation were observed, indicating apoptotic cell death. Catalase (scavenger of hydrogen peroxide), mannitol (scavenger of hydroxyl radicals) and superoxide dismutase (scavenger of superoxide radicals) reduced the level of ROS production under LA, suggesting the generation of H2O2, OH* and O2- radicals, respectively. It was revealed that catalase and mannitol effectively inhibited LA-mediated tumor cell death, increasing 2.5 times the percentage of alive cells in comparison to LA. However, superoxide dismutase had no significant inhibitory effect on cytotoxic activity of LA, indicating the minor role of superoxide anions in the implementation of antitumor activity of this antibiotic. Combination of catalase, mannitol and superoxide dismutase with LA increased 4-fold the percentage of alive cells in comparison to the action of LA. Dynamics of ROS formation confirms that the increase of ROS is a very rapid process, but at the same time it is not a direct consequence of apoptosis triggering, mediated by mitochondria.

Study of antineoplastic action of novel isomeric derivatives of 4-thiazolidinone.

Pyrazole- and aryl-substituted derivatives of 4-thiazolidinone belong to a perspective group of compounds with potential antitumor action. Earlier, we have demonstrated high toxicity in vitro of several 4-thiazolidinones derivatives towards tumor cell lines. To further enhance the antitumor activity of novel 4-thiazolidinones, their chemical scaffold was optimized, and new pyrazole-thiazolidinones were synthesized. That allowed us to combine in one molecule the potential pharmacophore centres of previously tested compounds. As a result, "hybrid" 4-thiazolidinones exhibit higher toxicity in vitro toward tumor cells of various origin. The molecular mechanisms of antineoplastic activity of these compounds and intensity of induction of apoptosis strongly depended on the position of the substituent in the thiazolidinone cycle. In particular, Les-3661 compound, containing pyrazoline fragment in the 4th position of thiazolidinone core, exhibits 14 times higher cytotoxic activity towards tumor cells (LC50 = 3 µM) in comparison to its 2-substituted isomer Les-3713 (LC50 = 42 µM). It is demonstrated that in terms of underlying molecular mechanisms for cytotoxic effect the Les-3661 compound induced caspase-8 and caspase-9 dependent mixed-type of apoptosis, while Les-3713 induced apoptosis mediated only by the caspase-8.

[Antitoxic and antioxidant effects of N-stearoylethanolamin in the content of nanocomposite complex with doxorubicin in organs of mice with Lewis carcinoma].

The aim of the study was to evaluate the possibility to reduce the doxorubicin toxic effects by its immobilization with N-stearoylethanolamine (NSE) on nanocarier polyethylene glycol. The studied parameters of the doxorubicin toxicity were: the level of creatinine in the mice blood plasma and activity of alanine aminotransferase and aspartate aminotransferase in the blood plasma of mice. The activity of catalase superoxide dismutase, glutathione peroxidase and intensity of lipid peroxidation was determined in the tissues of the heart, kidneys and liver. Doxorubicin in the content of nanocarrier alone caused an increase of serum creatinine and aspartateaminotrasferase activity in plasma of experimental animals with carcinoma. Nanocomposite which contained doxorubicin and NSE, did not cause an increase of these parameters. It has been shown that the administration of a carrier containing doxorubicin to mice with Lewis lung carcinoma caused the decrease of catalase activity in mice with carcinoma. The combination of NSE and doxorubicin on the carrier led to the normalization of this parameter to the level of intact animals. NSE immobilized on a carrier together with doxorubicin caused a decrease in the activity of superoxide dismutase in the kidney tissue of mice with tumor. The tumor growth caused the increase of the of superoxide dismutase in mice. The administration of a carrier which contained doxorubicin and NSE normalized superoxide dismutase in heart tissue contrary of kidney. The obtained results show the antitoxic and antioxidant effects of N-stearoylethanolamine immobilized in the nanocarrier complex together with doxorubicin.

[Protective effect of N-stearoylethanolamine in suspension and in nanocomposite complex in the organs of mice with the Lewis carcinoma under doxorubicin intoxication administration].

The antioxidant effects of N-stearoylethanolamine (NSE) in the nanocomplex composition and in suspension are shown on the model of intoxication by doxorubicin in conditions of development of the Lewis carcinoma in the heart, kidneys and liver tissue and in the blood plasma of female mice. The NSE suspension reduces the level of urea in the blood plasma of mice with the Lewis carcinoma, which growth was revealed as a result of introduction of doxorubicin. Under introduction of nanocomplex the amount of urea remains at the level of that in the intact mice. In the blood plasma of mice with the Lewis carcinoma the NSE suspension and nanocomplex reduce activity of aspartate aminotransferase, the basic marker of necrosis of the heart tissue, growth of which was caused by the tumour development. Doxorubicinum increases activity of alanine aminotransferase, the marker of the liver lesion; introduction of NSE in the nanocomplex composition prevents the growth of the enzyme activity. N-stearoylethanolamine, both in the nanocomplex and in suspension, modulates activity of enzymes of antioxidantive protection of the heart, kidney and liver tissue of mice with the Lewis carcinoma.

Changes in signaling pathways of cell proliferation and apoptosis during NK/Ly lymphoma aging.

Expression of specific proteins involved in regulation of cell proliferation and apoptosis was studied at the initial (7-8 days after tumor inoculation), median (13-14 days), and terminal (20-21 days) stages of murine NK/Ly lymphoma development. Western-blot analysis using antibodies to MEK-ERK signaling pathway, E2F-1/2 and c-Myc, pSTAT1, pSTAT3, pSTAT5, anti-apoptotic Bcl-XL and pro-apoptotic p53 and Rb proteins, as well as active cleaved forms of caspases-3, -6, -7, was carried out to investigate the growth and survival status of NK/Ly cells. There was a marked increase in the expression of E2F-1/2 transcription factors, MAPK signaling cascade and c-Myc, which suggests intensive proliferation of lymphoma cells at terminal stage of tumor development. However, cytomorphological investigation and electrophoretic study of DNA fragmentation have shown degeneration of NK/Ly lymphoma cells and increase in their death. No expression of p53 protein or cleaved forms of caspases-3, -6, -7 was found, which suggests a caspase-independent type of apoptosis in these cells. Ascitic fluid collected at a terminal stage of NK/Ly lymphoma development was significantly weaker in supporting tumor cell growth than ascitic fluid collected at the initial stage of tumor development. It is suggested that uncontrolled cell proliferation at terminal stage of the NK/Ly lymphoma development causes nutrient deprivation and deficiency of specific growth factors in the ascitic fluid, due to overexpression of MEK-ERK, E2F and c-Myc, thereby leading to the induction of apoptosis.

[Polypeptide growth factors in embryogenesis and tumor growth]. / Polipeptidnye faktory rosta v protsessakh émbrional'nogo razvitiia i opukholevogo rosta.

Polypeptide growth factors, which belong to different families (epidermal growth factors, insulin-like growth factors, fibroblast growth factors, transforming growth factors-beta, and some others), were characterized regarding their specific role in embryogenesis and tumor growth. Differences and parallels of the functioning of growth factors in these processes have been noted. Potential significance of the described characteristics of growth factors for directed modulation of embryogenesis and tumor growth is discussed.

[The similarities and the differences of embryogenesis and carcinogenesis]. / Skhodstvo i razlichie émbriogeneza i kantserogeneza.

The main similarities and embryonic and tumor cells, as well as the mechanisms preventing the malignant transformation of embryonic cells, are presented in this review. Special attention is paid to the role of specific polypeptide growth factors in reciprocally excluding processes: embryogenesis and carcinogenesis. Based on the presented analysis, new potential targets for antitumor drugs are considered.