| Abstract | Uvod: Karcinom prostate (KaP) najčešće je dijagnosticirani karcinom u odraslih muškaraca i danas je drugi po redu vodeći uzrok smrti u muškaraca. Kao i kod drugih tumora intenzivno se izučavaju biomarkeri korisni u ranom otkrivanju i dijagnozi karcinoma prostate, prognozi, terapiji ili u određivanju molekularnih putova karcinogeneze. Prijeoperacijski serumski PSA, patološki gradus po Gleasonu, pozitivni kirurški rubovi (s ekstraprostatičnom ekstenzijom i bez nje) i kapsularna incizija značajni su prediktori kliničkog i biokemijskog povrata bolesti. Ipak pravu prognostičku važnost pozitivnih rubova preostaje još definirati. U karcinomima prostate metaloproteinaze -2 i -9 (MMP-2 i -9) su molekularni biomarkeri koji upućuju na njegov invazivni i metastatski potencijal. Promjene u ekspresiji i funkciji adhezijskih molekula kao što je CD44s koreliraju s progresijom malignih tumora. CD44s regrutira i nakuplja MMP-9 na staničnoj membrani što omogućuje ovoj kinazi da indirektno potiče angiogenezu i invaziju. CD44s je, također, prepoznat kao marker matičnih tumorskih stanica koje u konceptu rane metastaze igraju veliku ulogu u samoobnavljanju stanica, pripremanju premetastatskog gnijezda epitelno-mezenhimnoj tranziciji i otpornosti na apoptozu. Spoznaja o različitoj ekspresiji CD44s biljega te metaloproteinaza u području invazije može pomoći u prepoznavanju bolesnika s agresivnijim oblicima tumora prostate koje bi trebalo, osim kirurški, dodatno liječiti.
Ciljevi su ove studije bili utvrđivanje međusobne povezanosti biomarkera kao što su dob, PSA, Gleasonov zbroj, stupanj bolesti kod bolesnika liječenih radikalnom prostatektomijom te utjecaja ovih biomarkera na biokemijski povrat bolesti kod prostatektomiranih bolesnika uz usporedbu grupe s pozitivnim i onih s negativnim rubovima resekcije. Također je cilj bio utvrditi i usporediti tkivnu ekspresiju metaloproteinaza -2 i - 9 i CD44s biljega u glavnoj masi tumora i tumoru na pozitivnom rubu, uz usporedbu ove ekspresije sa spomenutim kliničko-patološkim parametrima.
Pacijenti i metode: Istraživanje je randomizirano retrospektivno i obuhvaća razdoblje od 2001. do 2006. godine. Iz arhive medicinske dokumentacije na Klinici za urologiju Kliničkog bolničkog centra Rijeka odabran je 121 bolesnik s acinarnim tipom adenokarcinoma prostate te su prikupljeni odgovarajući klinički podatci. Iz arhivskog materijala Zavoda za patologiju Medicinskog fakulteta u Rijeci izdvojio se tumorski materijal odabranih bolesnika dobiven radikalnom prostatektomijom. Tkivna ekspresija metaloproteinaza -2 i -9 i CD44s biljega te proliferacijski marker Ki67 odreĐivali su se standardnom Avidin-Biotin imunoperoksidaza tehnikom na tkivnim mikroarejima (TMA). Ekspresija MMP-a i CD44s, procjenjivala se semikvantitativno, uzimajući u obzir postotak i intenzitet obojenja tumorskih stanica. Dobiveni podatci obraĐeni su statističkim metodama i usporeĐeni s novijom svjetskom literaturom.
Rezultati: Dob bolesnika nije bila u korelaciji s ostalim patohistološkim parametrima, pa tako niti u korelaciji s ekspresijom molekularnih markera. Također se niti jedan patohistološki parametar samostalno nije izdvojio kao koristan i značajan dijagnostički pokazatelj. Analizom prosječnog zbroja i usporedbom skupina s pozitivnim i negativnim rubom ustanovljen je značajno veći prosječni Gleasonov zbroj u skupini s pozitivnim rubom (p=0,0032), dok se PSA u obje skupine značajno ne razlikuje. Značajna razlika između skupina postoji samo u frekvencijama stupnjeva pT2a, pT3a te pT3b. MMP-2 i -9 obojenje u tumorskim stanicama je citoplazmatsko i variralo je u intenzitetu i postotku. U svim tumorima bojenje je bilo pozitivno na matriks metaloproteinaze. Veća ekspresija MMP-2 nađena je u tumorima slabije diferencijacije, odnosno višeg Gleasonova zbroja. MMP-2 ekspresija u području rubova bila je značajno veća od ekspresije u glavnoj tumorskoj masi (p=0,0301). MMP-9 pokazuje značajno veću ekspresiju u grupi tumora s pozitivnim rubom resekcije (p=0,0121) i tumorima većim od 1 cm (p=0,038), a povišena ekspresija je statistički značajno povezana s biokemijskim povratom bolesti u grupi bolesnika s tumorima negativnog ruba resekcije (p=0,0207). CD44s molekula pokazuje gubitak ekspresije na tumorskim stanicama s porastom Gleasonova zbroja, s diskontinuiranim membranskim bojenjem i fokalnom nepravilnom distribucijom u uzorku. CD44s i MMP-9 međusobno su inverzno ovisni biomarkeri (p=0,0185). Ki67 indeks proliferacije značajno je povećan u tumorima u skupini pacijenata s biokemijskim povratom bolesti (p=0,0172).
Zaključak: Naš cilj određivanja markera koji se razlikuje u ekspresiji na rubu i u tumorskoj masi, u konačnici je postignut. Uspjeli smo dokazati jaču ekspresiju MMP-2 upravo na rubu tumorske mase, ali i pokazati korisnost određivanja MMP-9 koji se jače eksprimirao u skupini tumora s pozitivnim rubom. Naše istraživanje također je dokazalo korelaciju MMP-9 i CD44s, a određivanje Ki67 indeksa proliferacije u tumorima pokazalo je prognostički značaj. |
| Abstract (english) | Objectives: Prostate cancer is the most frequently diagnosed cancer in elderly men, and it is the second cause of death in man today. As well as in other tumors, biomarkers useful in the early detection and diagnosis of prostate cancer, prognosis and therapy, or in determining molecular ways of carcinogenesis are intensively examined.
Pre-operative serum PSA, pathological grade according to Gleason, positive surgical margins (with and without extraprostatic extension) and capsular incision are significant in predicting clinical and biochemical recurrence of the disease. However, a real prognostic importance of positive margins should still be defined. In prostate carcinomas, MMP-2 and MMP-9 are molecular biomarkers which indicate their invasive and metastatic potential. Changes in the expression and function of adhesion molecules, as for example CD44s, correlate with a progression of malignant tumors. CD44s recruits and accumulates MMP-9 on the cell membrane, enabling this kinaze to stimulate indirectly angiogenesis and invasion.
CD44s is also recognized as a marker of parent tumor cells playing, in early metastasis a significant role in the renewal of the cells, preparation of pre-metastatic,
epithelic-mesenchimal transition and resistance to apoptosis. Understanding a different expression of CD44s marks and metalloproteinasis in the field of invasion may help in recognizing patients with more aggressive forms of prostate tumors who should be treated surgically and also additionally.The aim of this study was the identification of a mutual connection of biomarkers, as the patients age, PSA, Gleason score, the stage of the disease in patients treated by radical prostatectomy and the influence of these biomarkers on the biochemical recurrence of the disease in prostatectomy patients, and comparing groups of patients with positive as well as those with negative resection margins. The aim was also to determine and compare tissue expression of metalloproteinasis-2 and -9 and CD44s markers in the main tumor mass and the tumor on the positive margin, and comparing this expression with the mentioned clinic pathological parameters.
Patients, material and methods: The study is retrospective and it covers the period from 2000 to 2006. From the files of the medical documentation of the Clinic of urology of the Clinical Hospital Center Rijeka, 121 patients with acinous type of prostate adenocarcinoma were chosen and adequate clinical data were collected. From the files of the Department of Pathology of the School of medicine in Rijeka, tumor material was taken from chosen patients received by radical prostatectomy.
Tissue expression of metalloproteinesis-2 and -9 and CD44s marks and a proliferative marker Ki67 were identified by immunohistochemic standard Avidin-Biotin immunoperoksidose technique on tissue microareas (TMA). The expression of MMP and CD44s was evaluated semiquantitively, taking into consideration the percentage and intensity of tumor cells coloring. The obtained data were analyzed using statistical methods and then compared with more recent world literature.
Results: The age of the patient was not in correlation with other pathohistologic parameters and so neither in correlation with the expression of molecular markers. Also, not a single pathologic parameter was isolated as a useful and significant diagnostic tool. By analyzing an average score and comparing groups with a positive and negative margin, a significantly larger Gleason score was found in groups with a positive margin (p=0.0032), while PSA in both groups is not significantly different. A significant difference among groups exists only in frequencies of stages pT2A, pT3A and pT3B. MMP-2 and -9 coloring in tumor cells are cytoplasmic and varied in the intensity and percentage, and we did not find negative tumors. Increased expression of MMP-2 was found in tumors of poor differentiation and higher Gleason sum. MMP-2 expression at the edges was significantly higher than the expression in the main tumor mass (p=0,0301). MMP-9 shows a notably higher expression in a group of tumors with a positive group resection edge (p=0,0121).
Tumors larger than 1 cm. (p=0,038) and elevated expression were significantly assiociated with biochemical recurrence in the group of patients with tumors of a negative edge of the resection (p=0,0207). CD44s shows a loss of expression of molecules on tumors cells with increased Gleason sum of discontinuous membrane staining and focal irregular distribution in the sample. CD44s and MMP-9 together are inversely dependent biomarkers (p=0,0185). Ki67 proliferation index was significantly increased in tumors in the group of patients with biochemical reccurence (p=0,01272).
Conclusion: This study confirmed the influence and prognostic significance of the investigated biomarkers in prostate carcinomas in patients treated by a radical prostatectomy, especially in a tumor group with a positive margin of surgical resection which presents a problem in the choice of a postoperative treatment of the patient. |
