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ГоловнаРізне → Особливості функціонального стану нирок у підлітків із первинною артеріальною гіпертензією (автореферат) - Реферат

Особливості функціонального стану нирок у підлітків із первинною артеріальною гіпертензією (автореферат) - Реферат

Ключові слова: рак яєчників, мікроваскуляризація, цитокіни, ФНП-, ТФР-β, BRCA1, p57, p53, антиангіогенна терапія, виживаність.

SUMMARY

Lomnytska M.I. Angiogenesis in ovarian cancer: cytokine regulation and prognostic value.- Manuscript.

Thesis for the degree of Candidate of Medical Sciences by specialty 14.01.07 - Oncology. - R.E.Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology of the National Academy of Sciences of Ukraine, Kyiv, 2005.

The aim: to study the role of growth-inhibiting cytokines and tumor-suppressor proteins in the regulation of angiogenesis in ovarian cancer (OC), to estimate the prognostic value of the microvessels' density in OC for the individualization of the patients' treatment.

The 125 OC patients, treated in 1995-2005 were studied. The average age of patients was 54,70,9 (113 of patients of the age of 45 - 65). 48(38%) patients were diagnosed with І-ІІ stage (FIGO classification) of OC, and 77(62%) – with III-IV stage. The treatment of patients was combined: radical surgical (extirpation of uterus with adnexes, omentectomy) with adjuvant chemotherapy (ChT) or neoadjuvant ChT with the surgery and adjuvant ChT (6 cycles: cys-platinum 75.0 mg/m2, cyclophosphamide 750.0 mg/m2). Antiangiogenesis therapy with Laferon (Interferon-2b, "Biofarma", Ukraine) (1 mln IU/day over 6 months) was performed to 12 patients with chemoresistant OC. We have used immunohistochemistry (with monoclonal antibodies to f. VIII, proteins BRCA1, p53, p57), cell culture methods (cell lines L929, Mv1Lu (CCL64) and microvascular endothelial cells of human derma; primary cell culture purification and biologic assay for the measurement of the concentration of TNF- and TGF-, [3Н]-incorporation assay, methyltertazolium assay), immunocytochemistry, proteomix studies (two-dimensional protein electrophoresis, mass-spectrometry, immunobloting), statistical (STATISTICA 6.0.).

We have estimated the prognostic value of the microvessels' density (MD) of OC for the overall and 3- and 5-year survival of patients, and suggested to use this prognostic factor in clinical practice. The significant increase of the MD of OC was shown at advanced (III – IV) stages of disease. The MD of the serous ovarian cancer was lower than MD in serous-papillary and mucinous OC. Nondifferentiated OC was characterized by significant higher tumor MD than compared histological types. The tumor MD was the highest at OC with G3 than with G2 and G1. The microenvironment of OC was characterized by the composition of supernatants of primary OC cell lines. The negative correlation (r= -0.4713, p=0.007) between the MD of OC and concentration of TNF- in supernatants was shown. Same dose-dependent tendency was found in in vitro study with human microvascular endothelial cells. This suggests the "dose-dependent" regulatory role for TNF- in OC angiogenesis. Only tendency to negative correlation (r=-0.356, p=0.095) between MD of OC and production of TGF-β by primary OC cell lines was shown. However, in in vitro system TGF-β in a dose 0.05 ng/ml significantly inhibited the proliferation of human microvascular endothelial cells. For the detailed estimation of role of TGF-β in the regulation of angiogenesis, the analysis of proteome of human microvascular endothelial cells was performed. We have identified 54 proteins, which determine the final effect of TGF-β in studied cells. Identified proteins were divided into 11 functional groups: signal transducers (13 proteins), cell cycle regulators (р57Kip2, E2F6 etc), cytoskeleton components (7 proteins), regulators of methabolism etc. Additionally, 25 Smad3-interacting proteins were identified.

The negative correlation (r= - 0.231, p=0.061) between the MD of OC and the level of expression of BRCA1 in OC cells was shown. It means the association of the positive expression of BRCA1 with the low MD of OC, and of low or negative expression of BRCA1 with the high MD in OC. The level of expression of BRCA1 was shown to decrease with the disease progression (r= -0.370, p=0.0096). The 1-, 3- and 5-year survival of OC patients was shown to be associated with the level of expression of BRCA1, and immunohistochemical detection of BRCA1 expression was suggested for use in clinics for prognosis of OC patients' survival. The positive correlation between the MD of OC and the expression of p53 in OC cells (r=0,39, p=0,011) was shown: hyperexpression of p53 in OC cells was associated with high MD of OC. The decrease in expression of p57 in OC cells was shown to be associated with the activated angiogenesis in OC.

Interferon-2b (IFN-2b) inhibited proliferative potential of human microvascular endothelial cells in a dose-dependent manner. The inhibitory activity of IFN-2b in a dose of 2.5-5.0 IU/ml was 1.5-fold and in higher doses (50-75 IU/ml) was 1.5-2.0-fold compared to control. With the increase of the dose of IFN-2b over 200.0 IU/ml, its inhibitory effect did not increase. On contrary, the antiproliferative activity of IFN-2b to cancer cells of lines MCF-7 and OVCAR-3 was minimal or absent at low doses, and appeared at the dose of 5000 IU/ml. Cytostatics cys-platinum, paclitaxel and cyclophosphamide in low doses inhibited the proliferative potential of human microvascular endothelial cells. Obtained results of studies in vitro and of investigations about the antiangiogenesis activity of IFN-2b in vivo (Slaton J. et al., 2001; Tedjarati S. et al., 2002; Streck J. et al., 2004) were the substantiation for the suggestion of use of this cytokine in a „metronomic antiangiogenesis regimen". The possibility of therapeutic use of IFN-α2b (Laferon) in low doses (1 mln IU/day) for a long period (over 6 months) for the inhibition of angiogenesis was suggested to OC patients with chemoresistant nonresectable tumor lesions.

Key words: ovarian cancer, microvessels' density, cytokines TNF-α, TGF-β, BRCA1, p57, p53, antiangiogenesis therapy, survival.

АННОТАЦИЯ

Ломницкая М.И. Ангиогенез в раке яичника: цитокинная регуляция и прогностическое значение. - Рукопись.

Диссертация на соискание ученой степени кандидата медицинских наук по специальности 14.01.07. – онкология. – Институт экспериментальной патологии, онкологии и радиобиологии им. Р.Е. Кавецкого НАН Украины, Киев, 2005.

Определено прогностическую ценность степени МВ РЯ для виживаемсти больных. Предложено использование этого показателя в клинической практике. Определено, что между степенью МВ РЯ и концентрацией цитокина ФНП- в супернатантах первичных клеточных культур РЯ существует оборотная связь, а между МВ РЯ и концентрацией ТФР-β – тенденция к оборотной связи. В системе in vitro ТФР-β ингибирует пролиферацию эндотелиоцитов микрососудов. Проанализировано протеом этих клеток и идентифицировано 54 белки, активностью которых определяется конечный эффект ТФР-β. Идентифицировано белки, которые взаимодействуют с медиатором передачи сигнала ТФР-β - Смад3. Показано, что снижение экспрессии белков BRCA1, р57 и гиперэкспрессия белка р53 в клетках РЯ сопровождается активацией ангиогенеза в опухоли. Предложено определять степень экспрессии белка BRCA1 в клетках РЯ для прогноза виживаемости больных. Обосновано возможность терапевтического использования интерферона-α2b (Лаферона) с антиангигенной целью в низких дозах на протяжении длительного времени в программе комбинированного лечения больных на химиорезистентный РЯ.

Ключевые слова: рак яичника, микроваскуляризация, цитокины ФНП-, ТФР-β, BRCA1, p57, p53, антиангиогенная терапия, выживаемость.

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