Identifikacija karboniliranih proteina u različitim stadijima kolorektalnog karcinoma

doctoral thesis

• Title (english): Identification of carbonylated proteins in different stages of colorectal cancer
• Author: Marijana Popović
• Mentor: Irena Drmić-Hofman (mentor)
• Mentor: Marijana Hranjec (mentor)
• Committee member: Tatjana Gazivoda Kraljević (predsjednik povjerenstva)
• Committee member: Silvana Raić-Malić (član povjerenstva)
• Committee member: Ivančica Delaš (član povjerenstva)
• Granter: University of Zagreb; Faculty of Chemical Engineering and Technology; Zagreb
• Defense date and country: 2016-05-05, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry
• Universal decimal classification (UDC): 54 - Chemistry. Crystallography. Mineralogy ; 577 - Biochemistry. Molecular biology. Biophysics

Abstract

Kolorektalni karcinom jedna je od najčešćih zloćudnih bolesti u oba spola, a većina slučajeva je sporadična i povezana s raznim čimbenicima rizika iz okoliša. Tkivo debelog crijeva i rektuma konstantno je izloženo prisutnosti reaktivnih kisikovih (ROS) i dušikovih (RNS) vrsta koji potječu od različitih spojeva, poput oksidiranih otpadaka hrane i toksina. Identifikacija oksidiranih (karboniliranih) proteina omogućava detekciju upravo onih proteina koji su podložni inaktivaciji pomoću ROS/RNS, što je temeljni korak u povezivanju patoloških obilježja s oksidativnim/nitrozativnim promjenama. U ovoj studiji analizirali smo ireverzibilna oksidativna oštećenja (karbonilaciju) proteoma tumorskog tkiva u različitim stadijima bolesti (Dukes A, B i C) i pripadajuće zdrave sluznice 18 pacijenata (6 pacijenata po stadiju) pomoću dvodimenzionalne diferencijalne gel-elektroforeze i spektrometrije masa. Također, određena je i razina malondialdehida kao sekundarnog čimbenika proteinske karbonilacije i pokazatelja ukupne lipidne peroksidacije. Rezultati pokazuju da postoje značajne razlike oksidativnog stanja proteina i lipida u različitim stadijima karcinoma debelog crijeva te da se povećavaju s porastom stadija karcinoma. Ukupno su identificirana 102 proteina s razlikama u karbonilaciji. Nije pronađen ni jedan karbonilirani protein koji se odnosio samo na stadij A kolorektalnog karcinoma, 16 karboniliranih proteina odnosilo se samo na stadij B, a njih 46 samo na stadij C. Sedam karboniliranih proteina zajednički su svim analiziranim stadijima, 33 proteina stadiju B i C, dok zajedničkih karboniliranih proteina nije bilo između stadija A i B te stadija A i C. Identificirani karbonilirani proteini pripadaju staničnim signalnim putevima koji su povezani s nastankom karcinoma, antioksidativnom enzimskom sustavu, metaboličkim procesima (ponajviše oksidativnoj fosforilaciji) te bi mogli biti razmatrani kao mogući novi biomarkeri kolorektalnog karcinoma. Ukoliko bi se daljnjim istraživanjima na većem broju uzoraka i na drugim tipovima tumorima potvrdilo da razlike u nastanku malignih fenotipova mogu biti izravna funkcionalna posljedica oksidativnog oštećenja relevantnih proteina, to bi moglo doprinijeti boljem razumijevanju etiologije kolorektalnog karcinoma te bi moglo biti polazište za razvoj novih preventivnih i terapijskih strategija.

Abstract (english)

Colorectal cancer is one of the most frequent disease in both genders, and is mostly sporadic, associated with a variety of environmental risk factors. The gastrointestinal tract, especially the colon, is constantly exposed to presence of reactive oxygen (ROS) and nitrogen (RNS) species derived from different compounds, like oxidized food debris and toxins. Identification of proteins modified by specifically oxidative/nitrosative injury in affected tissues allows determination of proteins more prone to inactivation by ROS/RNS, which is a fundamental step in linking well-established pathological hallmarks to oxidative/nitrosative changes. We have identified oxidatively damaged (carbonylated) proteins in different stages of colorectal cancer (Dukes A, B and C) compared to non-tumor tissue of the same patient with two dimensional differential gel-electrophoresis and mass spectrometry. In total, 102 carbonylated proteins were identified. There were no changes in protein carbonylation related only to stage A, 16 carbonylated proteins were related only to stage B and 46 proteins related only to stage C. Seven carbonylated proteins were mutual to stages A, B and C, 33 mutual proteins to stages B and C, while there were no mutual carbonylated proteins related to stages A and B, and stages A and C. In total, tissues of 18 patients were analysed, six patients per each stage. The level of malondialdehyde, secondary products during lipid peroxidation and oxidative agens for protein carbonylation generation, was also determined. Our study showed there are significant differences in protein and lipid oxidative damage that differs between tumor and non-tumor tissue in different stages of colorectal cancer and is increasing through the cancer stages. Carbonylated proteins identified in this study belong to cancer-related signaling pathways, antioxidative enzymatic system, metabolic pathways (oxidative phosphorylation the most), and might be potential new biomarkers of colorectal cancer. If validated on other larger number of samples and different tumor types, this finding that differences in emergence of the malignant phenotypes can be a direct functional consequence of oxidative damage to relevant proteins, would make large contribution to better understanding of the ethiology of colorectal cancer and could be the starting point for novel preventive and therapeutic strategies.

• Keywords: dvodimenzionalna diferencijalna gel-elektroforeza; ireverzibilna oksidacija proteina; karbonilacija; karcinom debelog crijeva; LTQ Orbitrap spektrometrija masa oksidativni stres; RNS; ROS
• Keywords (english): two dimensional differential gel-electrophoresis; irreversible protein oxidation; carbonylation; colorectal cancer; LTQ Orbitrap mass spectrometry; oxidative stress; RNS; ROS
• Language: croatian
• URN:NBN: urn:nbn:hr:149:178783
• Promotion: 2016
• Project: Number: 141-0000000-0080; Title: PROFIL GLIKOPROTEINA U NORMALNIM I TUMORSKIM STANICAMA; Title: DIFFERENCES IN GLYCOPROTEIN PROFILE BETWEEN NORMAL AND TUMOR CELLS; Leader: Irena Drmić-Hofman; Jurisdiction: Croatia; Funder: MZOS; Funding stream: ZP
• Study programme: Title: Engineering Chemistry - Doctoral Course; Study programme type: university; Study level: postgraduate; Academic / professional title: doktor/doktorica znanosti, područje prirodnih znanosti, polje kemija (doktor/doktorica znanosti, područje prirodnih znanosti, polje kemija)
• Type of resource: Text
• Extent: 112 str. ; 30 cm
• File origin: Born digital
• Access conditions: Closed access
• Created on: 2023-12-11 10:27:05