Analiza ekspresije antiangiogenog FKBPL proteina, vaskularnog endotelnog faktora rasta i estrogenog receptora u endometrioidnom karcinomu uterusa i benignoj hiperplaziji endometrijuma
Analysis of the expression of antiangiogenic FKBPL protein, vascular endothelial growth factor and estrogen receptor in uterine endometrioid carcinoma and benign endometrial hyperplasia
Докторанд
Obradović, DaniloМентор
Oprić, DejanЧланови комисије
McClements, LanaTatić, Svetislav
Glumac, Sofija
Vukomanović, Biserka
Метаподаци
Приказ свих података о дисертацијиСажетак
ANALIZA EKSPRESIJE ANTIANGIOGENOG FKBPL PROTEINA, VASKULARNOG
ENDOTELNOG FAKTORA RASTA I ESTROGENOG RECEPTORA U
ENDOMETRIOIDNOM KARCINOMU UTERUSA I BENIGNOJ HIPERPLAZIJI
ENDOMETRIJUMA
Sažetak
Uvod: FKBPL (FK506-binding protein-like), pripada familiji imunofilina, FK506 vezujućih
proteina, i predstavlja aktuelno široko ispitivani negativni regulator rasta tumora, angiogeneze i
metastaza, sa perspektivom u razvoju ciljane onkološke terapije. Viši nivo ekspresije FKBPL kod
pacijenata sa karcinomom dojke je dokazan kao nezavisni prognostički faktor dužeg preživljavanja,
kao i dužeg intervala bez relapsa u grupi sa visokom ekspresijom estrogenskog receptora, dok je
niži nivo ekspresije FKBPL povezan sa lošijom prognozom preživljavanja. Pokazano je da je
povećana ekspresija FKBPL kod pacijenata sa karcinomom dojke praćena nižom ekspresijom
estrogenog receptora (ER), većom zavisnošću rasta tumora od estrogene stimulacije i većom
senzitivnošću tumora na endokrinu terapiju Tamoksifenom. U okvir...u predkliničkih studija
dokazano je inhibitorno dejstvo FKBPL i njegovih derivata na rast kancerskih stem ćelija
karcinoma dojke. Ekspresija FKBPL takođe je ispitivana na kulturama kancerskih stem ćelija
karcinoma ovarijuma i ciljana terapija bazirana na FKBPL je pokazala antiangiogeno dejstvo kojim
dovodi do usporavanja rasta kolonija stem ćelija karcinoma ovarijuma posredstvom inhibicije
migracije endotelnih ćelija. Dokazana je genska ekspresija FKBPL u stromi karcinoma
endometrijuma, dok su trenutno jedini izvor o imunohistohemijskoj ekspresiji FKBPL u
endometrijumu i karcinomu endometrijuma enciklopedijske baze podataka. Prema dostupnim i
relevantnim izvorima, do sada nije izvedena analiza značaja nivoa ekspresije antiangiogenog
FKBPL proteina u karcinomima endometrijuma, sa detaljnim sagledavanjem ključnih prognostičkih
parametara.
Ciljevi: Ciljevi ovog istraživanja su bili da se ispita nivo ekspresije FKBPL, VEGF-A i ER , kao i
pojedinačna povezanost nivoa ekspresije VEGF-A i ER sa nivoom eks presije FKBPL, u
endometrioidnom karcinomu i benignoj hiperplaziji endometrijuma; da se ispita povezanost nivoa
ekspresije FKBPL sa prosečnom gustinom krvnih sudova u endometrioidnom karcinomu i benignoj
hiperplaziji endometrijuma; i da se ispita povezanost nivoa ekspresije FKBPL u endometrioidnom
karcinomu endometrijuma sa dubinom invazije miometrijuma, prisustvom limfovaskularne invazije,
histološkim gradusom tumora i kliničkim stadijumom bolesti.
Metode: Studijsku populaciju je činilo 90 slučajeva endometrioidnog karcinoma i 40 slučajeva
benigne hiperplazije endometrijuma. Kalupi su izdvojeni iz arhive službe za patologiju a klinički i
histološki podaci su dobijeni uvidom u dokumentaciju pacijenata u Kliničko - bolničkom centru
Zemun. Imunohistohemijska bojenja na FKBPL, VEGF-A, ERα i CD34 su urađena, prema
uputstvima proizvođača, u laboratoriji Instituta za patologiju, Medicinskog fakulteta u Beogradu.
Intenziteti ekspresije FKBPL i VEGF-A su izvedena dvostruko slepom analizom od strane dva
patologa, kao i morfometrijskom metodom određivanja procenta površine žlezdanog tkiva sa
pozitivnom reakcijom antitela. Nivo ERα je određen u okviru dvostruko slepe analize određivanjem
intenziteta skalom 0 - 3, procentualne zastupljenosti ERα pozitivnih epitelnih ćelija skalom 0 - 5 i
Allered-ovim skorom koji je suma vrednosti prethodne dve skale i u rasponu je 0 - 8. Krvni sudovi
obeleženi antitelom CD34 su analizirani određivanjem vaskularne gustine morfometrijskom
metodom, na delu uzorka. Morfometrijska merenja su rađena softverom za analizu slike Fiji: an
open-source platform for biological-image analysis.
Rezultati: Intenzitet ekspresije FKBPL je pokazao statisticki značajno (p<0.001) niže vrednosti u
endometrioidnom karcinomu nego u benignoj hiperplaziji endometrijuma, umerenu pozitivnu
korelaciju (p<0.05) sa sva tri parametra ekspresije ERα, i umerenu negativnu korelaciju (p<0.05) sa
nivoom ekspresije VEGF-A na nivou celog uzorka...
ANALYSIS OF THE EXPRESSION OF ANTIANGIOGENIC FKBPL PROTEIN,
VASCULAR ENDOTHELIAL GROWTH FACTOR AND ESTROGEN RECEPTOR IN
UTERINE ENDOMETRIOID CARCINOMA AND BENIGN ENDOMETRIAL
HYPERPLASIA
Abstract
Background: FKBPL (FK506 binding protein-like), is a divergent member of the immunophilin
family, FK506 binding proteins, and is currently a widely studied negative regulator of tumor
growth, angiogenesis, and metastasis, with a perspective in the development of targeted therapy in
gynecological oncology. A higher level of FKBPL expression in breast cancer patients has been
proven to be an independent prognostic factor of longer survival, as well as a longer relapse-free
interval in the group with high estrogen receptor expression, while a lower level of FKBPL
expression is associated with a worse survival prognosis. It has been shown that increased
expression of FKBPL in patients with breast cancer is accompanied by lower expression of ER,
greater dependence of tumor growth on estrogen stimula...tion, and greater sensitivity of tumors to
endocrine therapy. In the framework of preclinical studies, the inhibitory effect of FKBPL and its
derivatives on the growth of breast cancer stem cells has been proven. The expression of FKBPL
was also examined on the cultures of ovarian cancer stem cells and the targeted therapy based on
FKBPL showed an antiangiogenic effect, which leads to the slower growth of ovarian cancer stem
cell colonies, by inhibiting the migration of endothelial cells. The gene expression of FKBPL in the
stroma of endometrial carcinoma has been proven, while currently, the only available source on the
immunohistochemical expression of FKBPL in the endometrium and endometrial carcinoma is the
encyclopedic human protein database. According to available and relevant sources, there are no
publications offering a comprehensive analysis on the subject of expression of the anti-angiogenic
FKBPL protein in endometrial cancers has been performed, with a detailed review of the key
prognostic parameters.
Objectives: The objectives of this study were to examine the expression level of FKBPL, VEGF -A,
and ER, as well as the individual association of the expression level of VEGF-A and ER with the
expression level of FKBPL, in endometrioid carcinoma and benign endometrial hyperplasia; to
examine the association of FKBPL expression levels with vascular density in endometrioid
carcinoma and benign endometrial hyperplasia; and to explore the association of FKBPL expression
levels in endometrioid endometrial carcinoma with the depth of myometrial invasion, presence of
lymphovascular invasion, histological tumor grade and clinical stage of the disease.
Methods: The study population consisted of 90 cases of endometrioid carcinoma and 40 cases of
benign endometrial hyperplasia. Paraffin molds were obtained from the archives of the pathology
service, and clinical and histological data were obtained from patient documentation at the Zemun
Clinical Hospital Center. Immunohistochemical stainings for FKBPL, VEGF-A, ERα, and CD34
were performed, according to the manufacturer's instructions, in the laboratory of the Institute of
Pathology, Faculty of Medicine in Belgrade. Expression intensities of FKBPL and VEGF-A were
performed by double-blind analysis by two pathologists, as well as by the morphometric method of
determining the percentage of the glandular tissue surface with a positive antibody reaction. The
level of ERα was determined in a double-blind analysis by determining the intensity on a scale of 0-
3, the percentage of ERα-positive epithelial cells on a scale of 0-5, and Allered's score, which is the
sum of the values of the previous two scales and is in the range of O-8. Blood vessels labeled with
the CD34 antibody were analyzed by determining the vascular density using the morphometric
method, on part of the sample. All morphometric measurements were made with Fiji image analysis
software: an open-source platform for biological-image analysis.
Results: FKBPL expression intensity showed statistically significantly (p<0.001) lower values in
endometrioid carcinoma than in benign endometrial hyperplasia, moderate positive correlation
(p<0.05) with all three parameters of ERα expression, and moderate negative correlation (p<0.05)
with the level of VEGF-A expression observed analyzing the whole sample...