MOLECULAR AND CLINICAL ONCOLOGY 7: , 2017

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1 MOLECULAR AND CLINICAL ONCOLOGY 7: , 2017 Evlution of epiderml growth fctor receptor serum levels nd their ssocition with clinicopthologicl chrcteristics in ptients with colorectl cncer MEHMET KARABULUT 1, CIGDEM USUL AFSAR 2, HALIL ALIS 1, EBRU ORAN 1, SENEM KARABULUT 3, CEVHER AKARSU 1, NURI ALPER SAHBAZ 1, ALPEN YAHYA GÜMÜSOGLU 1, ELIF BILGIN 4 nd NURI FARUK AYKAN 5 1 Clinic of Generl Surgery, Istnbul Bkırkoy Dr Sdi Konuk Eduction nd Reserch Hospitl, Istnbul; 2 Clinic of Medicl Oncology, Bkirkoy Acibdem Hospitl, Istnbul; 3 Clinic of Medicl Oncology, Istnbul Bkirkoy Dr Sdi Konuk Eduction nd Reserch Hospitl, Istnbul; Deprtments of 4 Bsic Oncology nd 5 Medicl Oncology, Institute of Oncology, Istnbul University, Istnbul, Turkey Received June 16, 2016; Accepted Mrch 9, 2017 DOI: /mco Abstrct. Colorectl cncer (CRC) is mjor public helth concern nd one of the leding cuses of cncer relted mortlity worldwide. The im of the present study ws to determine the serum epiderml growth fctor receptor (segfr) levels in helthy volunteers nd ptients with CRC, to determine the ssocition between tumor mrker levels nd clinicopthologicl findings, nd investigte its prognostic vlue. A totl of 140 ptients with CRC were enrolled in the present study. Pre tretment segfr levels were determined using ELISA. A totl of 40 ge nd sex mtched helthy controls were included in the nlysis. The medin ge of ptients ws 60 yers (rnge, yers); the mjority of the tumor locliztion ws to the colon (n=81, 58%). The medin follow up time ws 14 months, while 43 (31%) ptients experienced disese progression nd 31 (22%) succumbed to the disese. A totl of 81 ptients (58%) were in the erly stges of disese (stge II nd III), nd 42% of the ptients hd stge IV disese. The estimted 2 yer overll nd 1 yer progression free survivl rtes for the whole ptient group were 70% [95% confidence intervl (CI): ] nd 26.2% (95% CI: ), respectively. The number of ptients who received neodjuvnt tretment ws 37. Of the ptients who were dministered pllitive tretment, 24 received oxlipltin, wheres 22 received irinotecn nd 9 received fluorourcil/cpecitbine. A totl of 36 nd 15 of the ptients who received trgeted therpy were dministered bevcizumb nd cetuximb, respectively. Of Correspondence to: Dr Cigdem Usul Afsr, Clinic of Medicl Oncology, Bkırkoy Acıbdem Hospitl, 1 Hlit Ziy Usklıgil Cd., Zeytinlik Mhllesi, Istnbul, Turkey E mil: cigdemusul@yhoo.com Key words: epiderml growth fctor receptor, serum, dignostic, clinicopthologicl chrcteristics, colorectl cncer the 55 ptients with metsttic disese who received pllitive chemotherpy (CTx), 31% were CTx responsive. The bseline medin segfr levels were significntly higher in ptients with CRC compred with the helthy control group (P=0.002). In ddition, estblished clinicl vribles, including no surgicl resection, metsttic stge, higher pthologicl tumor stge, poorer regression score (3 4) nd higher lctte dehydrogense levels, were found to be ssocited with higher segfr levels (P=0.03, P=0.009, P=0.05, P=0.05 nd P=0.05, respectively). The results of the present study did not revel sttisticlly significnt ssocitions between segfr concentrtions nd overll nd progression free survivl rtes. In conclusion, segfr concentrtions my be dignostic mrkers in ptients with CRC; however, their predictive nd prognostic vlues were not determined. Introduction Colorectl cncer (CRC) is common nd lethl disese. CRC incidence nd mortlity rtes vry mrkedly worldwide. Globlly, CRC is the third most commonly dignosed cncer in men nd the second in women, with n estimted >1.2 million new cses nd 608,700 CRC relted deths in 2008 (1). Specific genetic chnges re considered to drive the trnsformtion from norml colonic epithelium to invsive cncer, nd these genetic muttions my be inherited or cquired (2). CRC represents n idel model for the study of the moleculr pthogenesis of cncer, due to the ccessibility of tissue for biopsy nd the cler progression from norml colonic epithelium to invsive cncer vi n intermedite precursor, the denomtous polyp (2). Severl blood biomrkers hve been investigted in CRC, including circulting microrna, endothelil cell specific molecule 1, neutrophil/lymphocyte rtio, pltelet/lymphocyte rtio, α 2 mcroglobulin, KRAS nd epiderml growth fctor receptor (EGFR) (3 9). EGFR is 170 kd glycoprotein tht belongs to the trnsmembrne tyrosine kinse receptor fmily, nd hs

2 788 KARABULUT et l: SERUM EGFR LEVELS IN COLORECTAL CANCER been detected in wide vriety of cncer types (10). The ctivtion of EGFR hs multiple consequences, such s cell growth, differentition nd prolifertion; it lso promotes mlignnt trnsformtion, ngiogenesis nd metsttic dissemintion (10). EGFR hs been reported to be overexpressed in the mjority (50 80%) of colorectl tumors, nd its expression hs been demonstrted to be ssocited with poor outcome in ptients with stge IV disese (11 14). Mourtzikou et l (15) identified tht serum (s)egfr levels were significntly lower in the ptient group when compred with those in helthy control individuls. In this previous study, there ws no significnt ssocition between tumor node metstsis (TNM) stge, histologicl grde, performnce sttus nd EGFR expression (15). Few studies hve reported n ssocition between histologicl grde nd EGFR overexpression (16,17), wheres number of investigtors consider the clinicopthologicl chrcteristics of colon crcinom not to be ffected by EGFR expression (18,19). However, in certin studies, higher segfr level t bseline ws ssocited with the best objective response nd my be considered significnt predictor of outcome in ptients with dvnced CRC (9). The present study imed to determine the segfr levels in helthy volunteers nd ptients with CRC, to determine the ssocition between the levels of this tumor mrker nd clinicopthologicl findings, nd to investigte its prognostic vlue. Ptients nd methods Study design nd eligibility criteri. The serum smples of 140 consecutive ptients with CRC who were referred to Istnbul University Institute of Oncology nd Bkirkoy Dr. Sdi Konuk Trining nd Reserch Hospitl (Istnbul, Turkey) between My 2011 nd August 2014 were obtined. The medin ge of the ptients ws 60 yers (rnge, yers). All the ptients were stged using the seventh edition of the Americn Joint Committee on Cncer TNM system (20) on rdiologicl nd pthologicl bsis. All the ptients were treted with multidisciplinry pproch. Ptients with colon cncer who hd undergone surgery including segmentl colon resection were treted with djuvnt chemotherpy (CTx) ccording to their stge. Ptients with rectl cncer, who received neodjuvnt rdiochemotherpy (RCTx) or rdiotherpy (RT), hd undergone low nterior resection or bdominoperinel resection. Certin ptients underwent pllitive surgery nd stge IV ptients received pllitive CTx, with or without trgeted therpy (bevcizumb or cetuximb). The pretretment evlution included detiled clinicl history nd physicl exmintion, with series of biochemistry tests nd complete blood cell count. Selection for tretment required n Estern Coopertive Oncology Group (ECOG) performnce sttus score of 0 2 (21), nd pproprite bone mrrow (hemoglobin >9 g/dl, bsolute neutrophil count >1,500/µl nd pltelet count >100,000/µl), crdic, renl nd heptic function. Ptients were treted with vrious CTx regimens, including single gent or combintion therpy. Regimens of single or combintion CTx were selected ccording to the performnce sttus of the ptients nd extension of disese. Ptients received one of the following tretment regimens: Simplified LV5FU2 (leucovorin 400 mg/m 2, followed by 5 fluorourcil s 400 mg/m 2 bolus nd 2,400 mg/m 2 infusion over 46 h every 2 weeks), cpecitbine (1,000 mg/m 2, twice dily, orl dministrtion, for 14 dys of ech 21 dy cycle), modified FOLFOX regimen (simplified LV5FU2 regimen plus oxlipltin 85 mg/m 2 every 2 weeks), FOLFIRI (simplified LV5FU2 regimen plus irinotecn 180 mg/m 2 every 2 weeks), XELOX (cpecitbine 1,000 mg/m 2, twice dily, orl dministrtion, for 14 dys plus oxlipltin 130 mg/m 2 every 3 weeks), or XELIRI (cpecitbine 1,000 mg/m 2, twice dily, orl dministrtion, for 14 dys plus irinotecn 240 mg m 2 every 3 weeks). Bevcizumb ws given t dose schedule of either 5 mg/kg every 2 weeks or 7.5 mg/kg every 3 weeks. Cetuximb 500 mg/m 2 ws dministered intrvenously every 2 weeks. All the ptients underwent pretretment imging of primry tumors using mgnetic resonnce imging (MRI) or computed tomogrphy (CT) scn. For ptients with evluble imging studies prior to nd following tretment, the rdiologicl response ws evluted ccording to the Response Evlution Criteri in Solid Tumors (version 1.1) (22) nd clssified s follows: Complete response (CR), prtil response (PR), stble disese (SD) or progressive disese (PD). The tumor response fter 2 months of CTx ws used for sttisticl nlysis. Follow up progrms for metsttic disese consisted of clinicl nd lbortory progrms nd CT scn or MRI, depending on which imging methods were used t bseline, nd performed t 8 week intervls during CTx or every 12 weeks for ptients receiving no nticncer tretment. Ptients with either CR or PR were clssified s responders, nd ptients with n SD or PD were considered non responders. The present study ws pproved by the Institutionl Review Bord (IRB) of Istnbul University, Institute of Oncology (Istnbul, Turkey). Bseline demogrphic, clinicl nd lbortory dt, including ge, sex, performnce sttus, tumor mrker levels, KRAS muttion sttus nd tretment detils, were obtined retrospectively for ll ptients using uniform dtbse templtes to ensure consistent dt collection. The ptient comorbidities included crdic nd metbolic diseses. The control group consisted of 40 ge nd sex mtched helthy femles with no previous history of mlignncy or utoimmune disorders. Blood smples were obtined from ptients with CRC t first dmission, prior to the dministrtion of ny therpy. Blood smples of helthy controls were collected in dry tubes nd the ser seprted from cellulr elements by centrifugtion (t 1,431 x g for 10 min) within 30 min following collection. Blood smples were stored t 80 C prior to nlysis. All the smples were collected under the pprovl of the IRB nd ll the ptients provided written informed consent. Mesurement of segfr levels. An EGFR ELISA kit (Shnghi Yehu Biologicl Technology Co. Ltd, Shnghi, Chin), which uses double ntibody sndwich ELISA to determine the level of humn EGFR in smples, ws used ccording to the mnufcturer's protocol. Serum smples nd stndrds were dded to the wells, which were pre coted with humn EGFR monoclonl ntibody. Streptvidin horserdish peroxidse ws dded to form immune complexes nd llowed to incubte t 37 C for 1 h. Unbound mteril ws wshed wy, nd chro-

3 MOLECULAR AND CLINICAL ONCOLOGY 7: , Tble I. Ptient clinicopthologicl chrcteristics. Chrcteristics No. of ptients Totl 140 Age, yers, medin (rnge) 60 (24 84) Sex, mle/femle 96/44 Performnce sttus, 0/1/2/3 68/61/7/1 Smoking, yes/no 61/66 Alcohol intke, yes/no 26/99 Comorbidities yes/no 56/79 Obstruction, yes/no 17/123 Surgery type Colectomy 56 Low nterior resection 36 Abdominoperinel resection 13 Pllitive surgery 11 pt stge b, 0/1/2/3/4 9/2/12/45/10 pn stge b, 0/1/2 42/18/14 Stge of disese, 2/3/4 17/64/59 Site of lesion, colon/rectum 81/59 Response to CTx c, CR/PR/SD/PD/unknown 2/15/10/24/4 Trgeted therpy, bevcizumb/cetuximb 36/15 Metstsis, yes/no d 59/81 Ptients with unknown dt were not included in the nlysis. b 81 non metsttic disese ptients with unknown dt were not included in the nlysis. c In 59 ptients with metsttic CRC. d Stge II nd III. CR, complete response; PR, prtil response; SD, stble disese; PD, progressive disese; CTx, djuvnt chemotherpy. Tble II. Histopthologicl chrcteristics nd lbortory prmeters. Vribles No. of ptients Histology, denocrcinom/mucinous 129/11 Grde, 1/2/3 8/56/6 Angiolymphtic invsion b, yes/no 30/18 Vsculr invsion b, yes/no 16/30 Perineurl invsion b, yes/no 18/28 Regression score c, 1/2/3/4 1/12/4/8 KRAS muttion sttus d, mutnt/wild type 24/28 Lctte dehydrogense, IU/ml Norml (<450) 97 High (>450) 16 Albumin, g/dl Norml (>4) 54 Low (<4) 58 Crcinoembryonic ntigen, ng/ml Norml (<5) 78 High (>5) 17 Crbohydrte ntigen 19 9, U/ml Norml (<38) 81 High (>38) 28 Ptients with unknown dt were not included in the nlysis. b 81 non metsttic disese ptients with unknown dt were not included in the nlysis. c 37 ptients with rectl cncer who received neodjuvnt tretment. d 59 ptients with metsttic colorectl cncer. mogen solution ws dded nd incubted t 37 C for 10 min in the drk for the conversion of the colorless solution to blue solution, the intensity of which is proportionl to the mount of EGFR in the smple. Upon the ddition of the cidic stop solution, the color ws converted to yellow. The colored rection product ws mesured using n utomted ELISA reder (ChroMte 4300; Awreness Technology, Inc., Plm City, FL, USA) t 450 nm. The results were expressed s ng/ml. Sttisticl nlysis. SPSS for Windows version 21.0 (IBM Corp., Armonk, NY, USA) ws employed for dt nlysis. Continuous vribles were ctegorized using medin vlues s cut off point. The Chi squre test or one wy nlysis of vrince were used for group comprison of ctegoricl vribles, nd the Mnn Whitney U test or Kruskll Wllis test were used for comprison of continuous vribles. The Spermn's rnk order correltion ws used for correltion nlysis. Progression free survivl (PFS) ws clculted from the dte of dmission to the dte of first rdiologicl progression, with or without elevted serum tumor mrker. Overll survivl (OS) ws clculted from the dte of first dmission to the clinic to disese ssocited mortlity or dte of lst contct with the ptient or ny fmily member. The Kpln Meier method ws used for the estimtion of survivl distribution, nd vritions in PFS nd OS were ssessed using the log rnk test. All sttisticl tests were two sided nd P 0.05 ws considered to indicte sttisticlly significnt difference. Results In totl, 140 ptients who were pthologiclly dignosed with CRC between My 2011 nd August 2014 were included in the present study. The bseline demogrphic nd histopthologicl/lbortory chrcteristics of ptients re presented in Tbles I nd II. The medin ge of the ptients ws 60 yers (rnge, yers). Mles constituted the mjority of the group (n=96, 69%). A totl of 43 of the ptients hd fmily history of cncer, including 12 with history of lung cncer nd 14 with history of CRC. The tumor locliztion ws to the rectum in 42% (n=59) nd the colon in 58% (n=81) of the ptients (right colon, n=17; heptic flexure, n=5; trnsverse colon, n=5; descending colon, n=13; splenic flexure, n=1; sigmoid colon, n=37; recto sigmoid junction, n=6; nd multiple synchronous colon tumors, n=3). The most frequent metsttic sites were the liver (n=40, 67.8%) nd the peritoneum (n=17, 28.8%). The rtes of synchronous (n=34) nd metchronous metstses (n=25) were 57.6 nd 42.4%, respectively. Of the 37 ptients with rectl cncer, 28 received fluoropyrimidine bsed RCTx, wheres 9 received short course RT. A totl of 71 ptients who hd djuvnt CTx received one

4 790 KARABULUT et l: SERUM EGFR LEVELS IN COLORECTAL CANCER Tble III. Serum mrker levels in ptients with colorectl cncer nd helthy controls. Ptients (n=140) Controls (n=40) Mrker Medin Rnge Medin Rnge P vlue segfr level (ng/ml) , , EGFR, serum epiderml growth fctor receptor. Figure 1. The vlues of serum EGFR ssys in ptients with colorectl cncer nd controls (P=0.002). EGFR, epiderml growth fctor receptor. of the following tretment regimens: Simplified LV5FU2 or cpecitbine (n=14), mfolfox (n=26) or XELOX (n=31). Oxlipltin nd irinotecn bsed combintion CTx regimens nd single gent fluoropyrimidine were used in 24, 22 nd 9 ptients, respectively. Bevcizumb ws dministered to 36 ptients, wheres 15 ptients received cetuximb s trgeted gent. A response to CTx ws observed in 31% of the 55 metsttic ptients who received pllitive CTx. The levels of segfr in ptients with CRC nd helthy controls re presented in Tble III. The bseline segfr levels were significntly higher compred with the control group ( vs ng/ml, respectively; P=0.002; Fig. 1). The ssocitions between the levels of segfr nd clinicopthologicl fctors re presented in Tbles IV nd V. No surgicl resection, metsttic sttus, higher pthologicl tumor stge, poorer regression score (3 4) nd higher lctte dehydrogense (LDH) levels were significntly ssocited with higher segfr concentrtions (ll P vlues <0.05). The medin follow up time ws 14.0 months (rnge, 1 34 months), 43 ptients (31%) experienced disese progression, nd 31 ptients (22%) succumbed to the disese. The medin PFS nd OS of the whole group were 7.3±1.0 months [95% confidence intervl (CI): 5 9 months] nd 26.9±1.1 months (95% CI: months), respectively. The 1 yer PFS rte ws 26.2% (95% CI: ); the 1 nd 2 yer OS rtes were 82.7% (95% CI: ) nd 70.0% (95% CI: ), respectively. Univrite nlyses were used to evlute the impct of clinicl fctors nd biomrkers on prognosis. The Kpln Meier method nd the log rnk test were performed for univrite nlysis of PFS nd OS. A significnt ssocition ws observed between other clinicopthologicl vribles, including presence of metstsis (P 0.05), no surgicl resection (P=0.01), CTx unresponsiveness (P=0.001), high serum levels of crcinoembryonic ntigen (CEA) (P=0.04) nd crbohydrte ntigen (CA) 19 9 (P=0.03), nd poorer PFS (Tbles VI nd VII). There were significnt ssocitions between other clinicopthologicl vribles, including the locliztion to the rectum (P=0.03), presence of metstsis (P<0.001), vsculr invsion (P=0.02), perineurl invsion (P=0.03), poor grde (P=0.02), low performnce sttus (P=0.04), no surgicl resection (P<0.001), CTx unresponsiveness (P=0.002), high serum levels of LDH (P=0.02), CEA (P<0.001) nd CA 19 9 (P<0.001), low serum levels of lbumin (P=0.02) nd poor OS (Tbles VIII X). However, segfr levels reveled no significntly dverse ssocition with PFS nd OS (P=0.12 nd P=0.11, respectively; Tbles VII nd X; Figs. 2 nd 3).

5 MOLECULAR AND CLINICAL ONCOLOGY 7: , Tble IV. Results of comprisons between the serum ssys nd vrious demogrphic nd disese chrcteristics. Vribles n Medin EGFR, ng/ml (rnge) P vlue Age, yers 0.33 < , ( ,230.81) , ( ,615.28) Sex 0.81 Mle 96 1, ( ,230.81) Femle 44 1, ( ,615.28) PS , ( ,143.55) , ( ,230.81) Smoking 0.54 Yes 61 1, ( ,615.28) No 66 1, ( ,230.81) Alcohol intke 0.87 Yes 26 1, ( ,116.45) No 99 1, ( ,230.81) Comorbidity 0.35 Yes ( ) No 79 1, ( ,615.28) Obstruction 0.38 Yes 17 1, ( ,230.81) No 123 1, ( ,141.99) Surgery 0.03 b Yes 116 1, ( ,230.81) No 24 2, ( ,643.89) pt stge 0.05 b ( ,169.16) , ( ,615.28) pn stge ( ,069.96) , ( ,615.28) Metstsis b Yes 59 2, ( ,230.81) No 81 1, ( ,615.28) Response to CTx 0.76 Yes (CR + PR) 17 1, ( ,116.45) No (SD + PD) 34 2, ( ,230.81) Trgeted therpy 0.37 Bevcizumb 36 1, ( ,116.45) Cetuximb 15 2, ( ,643.89) Site of lesion 0.56 Colon 81 1, ( ,069.96) Rectum 59 1, ( ,230.81) Stge II nd III. b P EGFR, epiderml growth fctor receptor; CTx, djuvnt chemotherpy; CR, complete response; PR, prtil response; SD, stble disese; PD, progressive disese; PS, performnce sttus. Discussion CRC is mjor public helth concern, with continuously incresing incidence rtes (23). In previous yers, notble steps forwrd in the moleculr chrcteriztion of dvnced CRC hve been tken. A multiplicity of serum mrkers hve been proposed for erly dignosis of CRC, estimtion of the disese extent nd monitoring ptient tretment (24,25). EGFR hs been detected in wide vriety of cncer types, for some of which its overexpression hs been suggested to

6 792 KARABULUT et l: SERUM EGFR LEVELS IN COLORECTAL CANCER Tble V. Results of comprisons between the serum ssys nd vrious histopthologicl fetures nd lbortory prmeters. Vribles n Medin EGFR, ng/ml (rnge) P vlue Histology 0.39 Adenocrcinom 129 1, ( ,615.28) Mucinous 11 2, ( ,230.81) Grde 0.51 Good ( ,747.00) Intermedite ( ,450.66) Poor 6 1, ( ,615.28) Angiolymphtic invsion 0.33 Yes 30 1, ( ,615.28) No ( ,143.55) Vsculr invsion 0.23 Yes 30 1, ( ,615.28) No ( ,615.28) Perineurl invsion 0.19 Yes 18 1, ( ,615.28) No ( ,143.55) Regression score ( ,462.00) , ( ,069.96) KRAS muttion sttus 0.63 Mutnt 24 2, ( ,643.89) Wild type 28 2, ( ,615.28) LDH Norml 97 1, ( ,230.81) High 16 2, ( ,643.89) Albumin 0.83 Norml ( ,230.81) Low 58 2, ( ,615.28) CEA 0.56 Norml 78 1, ( ,615.28) High 17 1, ( ,493.59) CA Norml 81 1, ( ,230.81) High 28 2, ( ,615.28) P LDH, lctte dehydrogense; CEA, crcinoembryonic ntigen; CA, crbohydrte ntigen be fctor ssocited with poor prognosis nd more ggressive clinicl progression (10). EGFR expression hs been demonstrted to be ssocited with poor outcome in ptients with stge IV CRC (11 14). However, segfr levels nd their dignostic, prognostic nd predictive roles in CRC hve not been investigted in detil. For non smll cell lung crcinom ptients, higher segfr levels hve been found to be significntly ssocited with higher OS, nd the pre tretment segfr levels constituted n independent prognostic fctor (26). For dvnced CRC, in the mjority of the studies, the clinicopthologicl chrcteristics of colon crcinom re not ffected by EGFR expression (18,19); however, in certin studies, higher segfr level t bseline ws ssocited with the best objective response nd my be considered significnt predictor of outcome in ptients with dvnced CRC (9). In the present study, the bseline segfr level ws significntly higher compred with the control group ( vs ng ml; P=0.002), wheres no surgicl resection, metsttic stge, higher pthologicl tumor stge, poorer regression sttus (3 4) nd higher LDH levels were found to be correlted with higher segfr concentrtions

7 MOLECULAR AND CLINICAL ONCOLOGY 7: , Tble VI. Univrite nlyses of progression free survivl ccording to ptient nd disese chrcteristics. Progression free survivl (months) Vribles Event no./totl no. Medin survivl (±SE) 1 yer survivl, % (±SE) P vlue All ptients 43/ (1.0) 26.2 (6.8) Age, yers 0.45 <50 6/ (2.2) Not reched 50 37/ (1.1) 25.0 (7.2) Sex 0.46 Mle 29/ (1.1) 28.6 (8.5) Femle 14/ (2.1) Not reched PS / (2.1) Not reched / (1.2) 24.1 (7.9) Obstruction 0.43 Yes 6/ (1.9) Not reched No 33/ (1.1) 24.2 (7.5) Surgery 0.01 b Yes 32/ (1.2) 31.3 (8.2) No 11/ (1.3) Not reched pt stge / (3.2) Not reched 3 4 8/ (6.0) Not reched pn stge / (3.2) Not reched 1 2 6/ (3.7) Not reched Metstsis 0.05 b Yes 33/ (0.9) 21.9 (7.3) No 10/81 NR Not reched Response to CTx b Yes (CR + PR) 4/ (2.3) Not reched No (SD + PD) 27/ (0.6) Not reched Trgeted therpy 0.06 Bevcizumb 21/ (1.2) 28.6 (9.9) Cetuximb 4/ (1.2) Not reched Site of lesion 0.18 Colon 19/ (1.4) 33.3 (11.1) Rectum 24/ (1.3) 20.8 (8.3) Histology 0.79 Adenocrcinom 37/ (2.6) 24.3 (7.1) Mucinous 5/ (1.1) Not reched Grde 0.79 Good 1/8 NR 9.0 (0.0) Intermedite 13/56 NR 7.5 (2.2) Poor 2/6 NR 5.5 (2.5) Regression score / (6.5) Not reched 3 4 0/ (0.0) Not reched KRAS muttion sttus 0.14 Mutnt 14/ (1.2) Not reched Wild type 14/ (1.7) Not reched Stge II nd III. b P SE, stndrd error; CTx, djuvnt chemotherpy; CR, complete response; PR, prtil response; SD, stble disese; PD, progressive disese.

8 794 KARABULUT et l: SERUM EGFR LEVELS IN COLORECTAL CANCER Tble VII. Univrite nlyses of progression free survivl ccording to lbortory prmeters. Progression free survivl (months) Vribles Event no./totl no. Medin survivl (±SE) 1 yer survivl, % (±SE) P vlue LDH Norml 27/ (1.1) 25.9 (8.4) 0.14 High 5/ (5.0) NR Albumin Norml 12/ (1.6) 26.3 (10.7) 0.57 Low 19/ (2.1) 41.7 (14.2) CEA Norml 16/ (1.5) 43.8 (12.4) 0.04 High 9/ (2.1) NR CA19 9 Norml 18/ (1.3) 38.9 (11.5) 0.03 High 19/ (1.7) 21.1 (9.4) EGFR <Medin 17/ (1.3) 31.3 (11.6) 0.12 >Medin 26/ (1.1) 23.1 (8.3) P NR, not reched; LDH, lctte dehydrogense; CEA, crcinoembryonic ntigen; CA, crbohydrte ntigen; EGFR, epiderml growth fctor receptor; SE, stndrd error. Figure 2. Progression free survivl curves in ptients with colorectl cncer ccording to serum EGFR levels (P=0.12). EGFR, epiderml growth fctor receptor. Figure 3. Overll survivl curves in ptients with colorectl cncer ccording to serum EGFR levels (P=0.11). EGFR, epiderml growth fctor receptor. (ll P vlues <0.05). However, segfr levels exhibited no significntly dverse ssocition with PFS nd OS (P=0.12 nd P=0.11, respectively). A previous study by Mourtzikou et l (15) reveled tht segfr levels were significntly lower in the ptient group when compred with those in helthy control individuls; however, these uthors collected blood smples from 20 ptients with CRC t preopertive stte nd from 30 ptients undergoing chemotherpy, which my hve ffected the study results. In nother study performed by Zmpino et l (9), the greter the segfr level t bseline, the lower the risk of no clinicl

9 MOLECULAR AND CLINICAL ONCOLOGY 7: , Tble VIII. Univrite nlyses of overll survivl ccording to ptient nd disese chrcteristics. Overll survivl (months) Medin survivl 1 yer survivl, % Vribles Event no./totl no. (± stndrd error) (± stndrd error) P vlue All ptients 31/ (1.1) 82.7 (3.3) Age, yers 0.30 <50 4/ (1.4) 90.9 (6.1) 50 27/ (1.2) 81.1 (3.8) Sex 0.76 Mle 20/ (1.3) 83.3 (4.0) Femle 11/ (1.9) 81.5 (5.9) PS 0 9/ (1.7) 87.5 (4.2) / (0.9) 77.3 (5.2) Obstruction 0.50 Yes 5/ (2.0) 81.1 (9.9) No 23/ (1.3) 83.1 (3.6) Surgery Yes 20/ (1.1) 88.0 (3.1) No 11/ (2.0) 56.9 (10.4) pt stge /23 NR (0.0) 3 4 3/55 NR 98.2 (1.8) pn stge / (0.7) 97.6 (2.4) 1 2 2/ (1.2) (0.0) Metstsis Yes 27/ (1.4) 61.1 (6.8) No 4/81 NR 97.5 (1.7) Response to CTx Yes (CR + PR) 2/ (1.6) 93.3 (6.4) No (SD + PD) 19/ (1.4) 47.6 (9.4) Trgeted therpy 0.55 Bevcizumb 13/ (1.7) 69.9 (8.6) Cetuximb 7/ (2.8) 52.5 (13.1) Site of lesion Colon 8/ (1.2) 91.0 (3.8) Rectum 23/ (1.6) 76.6 (4.9) 0.02 b <0.001 b <0.001 b Stge II nd III. b P NR, not reched; CTx, djuvnt chemotherpy; CR, complete response; PR, prtil response; SD, stble disese; PD, progressive disese b 0.03 b response; furthermore, higher segfr t bseline ws ssocited with the best objective response to EGFR trgeted therpy nd my be considered s significnt predictor of outcome in ptients with dvnced CRC. In conclusion, CRC is mjor public helth concern nd its incidence rtes continue to increse. Reserch into the biology of CRC hs identified lrge number of tumor mrkers tht provide dignostic, prognostic or predictive informtion. The present study demonstrted tht segfr concentrtions my be dignostic mrkers in ptients with CRC. However, their predictive nd prognostic vlues were not determined.

10 796 KARABULUT et l: SERUM EGFR LEVELS IN COLORECTAL CANCER Tble IX. Univrite nlyses of overll survivl ccording to histopthologicl chrcteristics. Overll survivl (months) Vribles Event no./totl no. Medin survivl (±SE) 1 yer survivl, % (±SE) P vlue Histology 0.48 Adenocrcinom 28/ (1.1) 84.4 (3.3) Mucinous 3/ (2.7) 70.7 (14.3) Grde 0.02 Good 0/8 NR (0.0) Intermedite 6/56 NR 90.7 (4.0) Poor 3/6 NR 66.7 (19.2) Angiolymphtic invsion 0.25 Yes 3/30 NR 96.6 (3.4) No 0/18 NR (0.0) Vsculr invsion 0.02 Yes 3/16 NR 93.3 (6.4) No 0/30 NR (0.0) Perineurl invsion 0.03 Yes 3/18 NR 94.1 (5.7) No 0/28 NR (0.0) Regression score /12 NR 91.7 (8.0) 3 4 0/13 NR (0.0) KRAS muttion sttus 0.25 Mutnt 13/ (2.0) 52.6 (10.3) Wild type 8/ (2.1) 75.8 (9.7) P NR, not reched; SE, stndrd error. Tble X. Univrite nlyses of overll survivl ccording to lbortory prmeters. Overll survivl (months) Vribles Event no./totl no. Medin survivl (±SE) 1 yer survivl, % (±SE) P vlue LDH Norml 21/ (0.9) 84.6 (3.8) 0.02 High 7/ (3.8) 62.5 (12.1) Albumin Norml 7/ (1.0) 89.8 (4.3) 0.02 Low 20/ (1.9) 73.7 (5.8) CEA Norml 7/ (0.6) 95.7 (2.5) <0.001 High 6/ (2.6) 68.0 (12.2) CA19 9 Norml 10/ (0.7) 93.4 (2.9) <0.001 High 13/ (2.8) 61.5 (9.7) EGFR <Medin 12/ (1.4) 84.7 (4.5) 0.11 >Medin 19/ (1.2) 80.6 (4.9) P SE, stndrd error; EGFR, epiderml growth fctor receptor; CA, crbohydrte ntigen; CEA, crcinoembryonic ntigen; LDH, lctte dehydrogense.

11 MOLECULAR AND CLINICAL ONCOLOGY 7: , References 1. Jeml A, Bry F, Center MM, Ferly J, Wrd E nd Formn D: Globl cncer sttistics. CA Cncer J Clin 61: 69, Ionov Y, Peindo MA, Mlkhosyn S, Shibt D nd Perucho M: Ubiquitous somtic muttions in simple repeted sequences revel new mechnism for colonic crcinogenesis. Nture 363: , Nonk R, Miyke Y, Ht T, Kgw Y, Kto T, Osw H, Nishimur J, Ikeng M, Murt K, Uemur M, et l: Circulting mir 103 nd mir 720 s novel serum biomrkers for ptients with colorectl cncer. Int J Oncol 47: , Jing H, Fu XG nd Chen YT: Serum level of endothelil cell specific molecule 1 nd prognosis of colorectl cncer. Genet Mol Res 14: , Ji J, Zheng X, Chen Y, Wng L, Lin L, Ye X, Chen Y, Chen D nd Dettke M: Stge dependent chnges of preopertive neutrophil to lymphocyte rtio nd pltelet to lymphocyte rtio in colorectl cncer. Tumour Biol 36: , Šunderić M, Šedivá A, Robjc D, Miljuš G, Gemeiner P, Nedić O nd Ktrlík J: Lectin bsed protein microrry nlysis of differences in serum lph 2 mcroglobulin glycosyltion between ptients with colorectl cncer nd persoons without cncer. Biotechnol Appl Biochem 63: , Li Y, Fu XH, Yun JQ, Yng ZY, Mo C, Dong XM, Tng JL nd Wng SY: Colorectl cncer: Using blood smples nd tumor tissue to detect K rs muttions. Expert Rev Anticncer Ther 15: , Tkhshi N, Ymd Y, Furut K, Ngshim K, Kubo A, Sski Y, Shoji H, Honm Y, Iws S, Okit N, et l: Assocition between serum lignds nd the skin toxicity of nti epiderml growth fctor receptor ntibody in metsttic colorectl cncer. Cncer Sci 106: , Zmpino MG, Mgni E, Sntoro L, Zorzino L, Dell'Orto P, Sonzogni A, Fzio N, Monfrdini L, Chipp A, Biffi R nd de Brud F: Epiderml growth fctor receptor serum (segfr) level my predict response in ptients with EGFR positive dvnced colorectl cncer treted with gefitinib? Cncer Chemother Phrmcol 63: , Spno JP, Lgorce C, Atln D, Milno G, Domont J, Benmouzig R, Attr A, Benichou J, Mrtin A, Morere JF, et l: Impct of EGFR expression on colorectl cncer ptient prognosis nd survivl. Ann Oncol 16: , Cheirsilp A, Rungvejvorchi P, Krlk A, Sngprkrn S, Pummi S nd Sngrjrng S: Determintion of epiderml growth fctor receptor (EGFR) in ptients with colorectl cncer. Cncer Ther 5: , Arteg CL: The Epiderml Growth Fctor Receptor: From Mutnt Oncogene in Non humn Cncers to Therpeutic Trget in Humn Neoplsi. J Clin Oncol 19 (18 Suppl): 32S 40S, Yrden Y: The EGFR fmily nd its lignds in humn cncer. signlling mechnisms nd therpeutic opportunities. Eur J Cncer 37 (Suppl 4): S3 S8, Goldstein NS nd Armin M: Epiderml growth fctor receptor immunohistochemicl rectivity in ptients with Americn Joint Committee on Cncer Stge IV colon denocrcinom: Implictions for stndrdized scoring system. Cncer 92: , Mourtzikou A, Stmouli M, Kroupis C, Christodoulou S, Skondr M, Kstni A, Pectsides D, Athnss G nd Dims C: Evlution of crcinoembryonic ntigen (CEA), epiderml growth fctor receptor (EGFR), epithelil cell dhesion molecule EpCAM (GA733 2), nd crbohydrte ntigen 19 9 (CA 19 9) levels in colorectl cncer ptients nd correltion with clinicopthologicl chrcteristics. Clin Lb 58: , Guo GF, Ci YC, Zhng B, Xu RH, Qiu HJ, Xi LP, Jing WQ, Hu PL, Chen XX, Zhou FF nd Wng F: Overexpression of SGLT1 nd EGFR in colorectl cncers showing correltion with the prognosis. Med Oncol 28 (Suppl 1): S197 S203, McKy JA, Murry LJ, Currn S, Ross VG, Clrk C, Murry GI, Cssidy J nd McLeod HL: Evlution of the epiderml growth fctor receptor (EGFR) in colorectl tumors nd lymph node metstses. Eur J Cncer 38: , Mohhmdi G, Jmilhmdi K, Lry S nd Ghffrzdegn K: Expression of membrnousepidermlgrowthfctorreceptor in colorectl denocrcinom nd its correltion with clinico pthologicl fetures. Pk J Biol Sci 14: , Abd El All HS, Mishriky AM nd Mohmed FA: Epiderml growth fctor receptor in colorectl crcinom: Correltion with clinico pthologicl prognostic fctors. Colorectl Dis 10: , AJCC (Americn Joint Committee on Cncer) Cncer Stging Mnul, 7th edition, Edge SB, Byrd DR, Compton CC (Eds), et l: Springer, New York. pp143, Krnofsky DA nd Burchenl JH: The Clinicl Evlution of Chemotherpeutic Agents in Cncer. In: McLeod CM (Ed), Evlution of Chemotherpeutic Agents. Columbi Univ Press, New York, pp196, Schwrtz LH, Litière S, de Vries E, Ford R, Gwyther S, Mndrekr S, Shnkr L, Bogerts J, Chen A, Dncey J, et l: RECIST 1.1 Updte nd clrifiction: From the RECIST committee. Eur J Cncer 62: , Siegel RL, Fedew SA, Miller KD, Goding Suer A, Pinheiro PS, Mrtinez Tyson D nd Jeml A: Cncer sttistics for Hispnics/Ltinos, Cncer J Clin 65: Sturgeon CM, Duffy MJ, Stenmn UH, Lilj H, Brünner N, Chn DW, Bbin R, Bst RC Jr, Dowell B, Estev FJ, et l: Ntionl Acdemy of Clinicl Biochemistry lbortory medicine prctice guidelines for use of tumor mrkers in testiculr, prostte, colorectl, brest, nd ovrin cncers. Clin Chem 54: e11 e79, Duffy MJ, vndlen A, Hglund C, Hnsson L, Holinski Feder E, Klpdor R, Lmerz R, Peltomki P, Sturgeon C nd Topolcn O: Tumour mrkers in colorectl cncer: Europen Group on Tumour Mrkers (EGTM) guidelines for clinicl use. Eur J Cncer 43: , Romero Ventos EY, Blnco Prieto S, González Piñeiro AL, Rodríguez Berrocl FJ, Piñeiro Corrles G nd Páez de l Cden M: Pretretment levels of the serum biomrkers CEA, CYFRA 21 1, SCC nd the soluble EGFR nd its lignds EGF, TGF lph, HB EGF in the prediction of outcome in erlotinib treted non smll cell lung cncer ptients. Springerplus 4: 171, 2015.