Histopathological Spectrum of Lower Gastro- Intestinal Colonoscopic Biopsy Lesion With Special Reference To Her-2/Neu Expression In Carcinoma Colon

Introduction : Cancer associated with colon is one of the principal risk factors from decease in women and men. Although importancegrowing aspect of human epidermis receptor2 (Her2)as atherapeutic target is rising butits role as a biomarker in form of predicting indicator indicatorwithin colorectal cancer(CRC)is stilla mystery.Present research isundertakenfor evaluating the of Her2/neu description in cancer of colon. Material & Method : This research comprises 256patientswith spectrum of histopathological treatment ranging from colitis to colorectal carcinoma at our department between 20152017. Her2/neu Immunohistochemistry was done in the colorectal carcinomaand scores based on Ruschoff et al (2012) Her-2 testing in gastric cancer. Result : Out of total number of 256 cases enrolled in our study group, majority belonged to the age group of 40-60 years, with M:F ratio being 1.4:1. Commonest site of lesion occurred in rectum (43.75%) followed by ascending colon and caecum (12.08%). Non neoplastic lesions constituted about two third of all cases, the commonest being inflammatory bowel disease(21.48%). In benign neoplastic lesions of tubular adenoma was thecommonest type, and in malignant commonest type was colorectal adenocarcinoma NOS(64.44%) followed by mucinous adeno carcinoma (22.22%). Because of more prominent membranous staining observed in high grade colorectal cancers, Her2neu expression is found to be an important predictive marker of carcinoma colon, especially the adenocarcinoma,NOS. Conclusion : Like Breast carcinoma, target oriented therapy can be instituted especially in Her 2/neu positive high grade and metastatic tumors.


INTRODUCTION:
A variety of inflammatory disorders affect colon and rectum, accompanying acute and chronic conditions. The non-neoplastic conditions include colitis of various etiology andbowel disease & syndrome diverticular disorder, whereas the neoplastic disease include benign and malignant polyps along with colorectal carcinoma(CRC) [1].
Colorectal carcinoma (CRC) isconsidered to be very frequently happening carcinomas&remains a majorreason of decease [2]. Colon cancer being the third prevalent cancer irrespective of gender predilection and was main factor of both class incidence & death. The rising overall incidence of colorectal cancer globally is around 3 percent per annum [3]. The incidence in rural areas in India are approximately half of that in urban population, less incident of CRC inour country is attributablefordiet which contains more percentage of fibers [4]. Chemotherapy has proven to bean efficient strategy for adjuvant therapy, but is still incapable of preventing recurrence in all patient [5]. In the era of tailored therapy,monoclonal Antibody is now playing a key in the treatment of metastatic CRC [2]. This one goal is the transmitter of human aspect of epidermal growth-2Her2/neu, primarily being associated with breast carcinoma. Although the comprehensive characterization in the molecular histopathogenesisof Colon cancer established ErbB2 enhancement as a effective therapeutic goal but its under-expression was related to its prediction of treatment's outcome [6].
HER1 human epidermal growth factor(EGFR.), (Her2/neu or ErbB2), Her-3 & Her-4 associated with growing aspect of human epidermal receptor family and weremembranous bound G-protein coupled receptor presentupon chromosome 17q21 whichencrypts an intracellular proteins 185KD that lacks a normal ligand. Mitogen activated protein kinase (MAPK) is activated byHer-2 that in turninitiates signal cascades and phosphoinositidyl-3kinase pathways PI3K/AKT those are essential for differentiation and proliferation of cells [7].
The dysregulation in the form of mutations, overexpression by Her2/neu encourages the development & transfer of tumors [8]. In the tumor model system, the geneof Her-2/neu overexpression avidly compare tomutagenesis, improved motility of cell, malignant transformation, metastasis and invasion.Other epithelial malignancies such as carcinoma lung, prostate and bladder have shown overexpression of Her-2/neu in several studies [7]. Thus potential new adjuvant monoclonal antibody can be used which causes direct inhibition of Tyrosine kinase (TRK) activity resulting in proapoptotic, antiangiogenic and anti-invasive effect [9].

AIM& OBJECTIVE :
a) To study the incidence of various lesions of lower gastro-intestinalcolonoscopic biopsy.
b) To study histopathology of lower GI lesions and findout incidence of various nonneoplastic as well as neoplastic lesions. c) To study the Her2/neu expression in carcinoma of colon with respect to age, sex, site, type and grade

Materials and methods :
A study was conducted prospectively in a tertiary care teaching Hospital from July 2015 till Oct 2017, on the lower GI colonoscopic biopsies, sent from the department of Gastroenterology of the same institution. Inclusion criteria included patients of either sex of all age groups with lower gastrointestinal signs and symptom as well as visible mucosal lesions.
The exclusion criteria was patients with perforated viscus, cases with prior chemo and radio therapy andrecent history of myocardial infarction. All histologically confirmed cases of CRC were included for Her2neu study with a follow-up of one year. After obtaining the well informed consent and thorough health experience, endoscopy with a compact forward facing camera endoscope & five to eight colonoscopies from different quadrant of the surface and margins of the mucosal lesions. All clinico-radiologic and endoscopic reports were procured for subsequent histopathologic examination. Full thickness biopsy of mucosa without much submucosal tissue was obtained from the margin (all four quadrants) surface and surrounding mucosa of suspicious lesions, as well as 4-6 biopsy from different segments of colon were taken. Tissue processing was followed byHarris Hematoxylin and eosin (H & E) staining followed by special staining was done, for establishing final diagnosis. All cases of the CRC were subjected to IHC stain of Her2neu using DAKO polyclonal rabbit antihuman cerb-2 oncoprotein.Patients especially with IHC of Her-2/neu score 2+ tumors were enrolled withfluorescent in-situ hybridization (FISH) testing.

PROCEDURE :
IHC is a method based on Antigen-Antibody reaction for identifying antigenic substance in the tissue. The primary antibody binds to specific tissue antigens. The biotinylated secondary antibody is directed to primary antibody. The streptavidin/horse radish peroxidase complex is then applied. Streptavidin then binds to biotin onthe secondary antibody and Horseradish peroxidase (HRP) acts as an indicator enzyme.On addition ofDiaminobenzidine(DAB) substrate, free oxygen radicals are released which oxidize DAB to a brown precipitation. The precipitate gets deposited on the antigen site and can be detected by microscopy [5] The four tier scoring system used by Ruchoff J etal. (2012) [10], for Her2 expression in gastric carcinoma was used in our study for scoring Her2/neu of allthe colorectal carcinoma Table1. Table 1. Guideline for colorectal carcinomatest of Her-2/neu, Ruchoff J et al [10].

Score
Observation Staining 0 No sign of cell staining in < 12 percent Negative One+ Light, scarcely noticeable in > 12 per cent of the cells; Negative Two+ In > 12 percent of cells, low to medium staining;circumferential, basolateral or lateral.

OBSERVATION:
In this study the wide age range was covered from 2 years to 85 years with maximum number of patients who underwent colonoscopy belonged to 41-60 years of age group in both sexes. Sex ratio is 1.4:1, male (58.88%) and female (41.11%). The incidence of disease was more commonly found in rectum (43.7%) in comparison to other sites of lower GI tract.Out of 256 cases the number of patients suffering from colorectal carcinoma was 35.15%, followed by IBD 21.48% , non-neoplastic polyp 14.84 % and neoplastic polyp 5.85%.Among 55 cases of idiopathic collative ulcers, inflammatory bowel syndrome, was 19.6%, disease of crohnwas only 3.4% revealed different stages of colitis ( acute, chronic acute and inactive) together constitute 21.48%. The incidence of Inflammatory polyp (39.47%) is highest among the group of non-neoplastic polypsTable-2.Adenomas are localized in the rectum (46.67%) followed by sigmoid colon (26.6%). Tubular adenomas constituted 80.0%Figure1.tubulovillous adenoma comprised 13.33%and villous adenoma is least with 6.67%. the distribution of adenoma according to different site is depicted inTable 3.  Age sex distribution of colon carcinomas are given inTable4. The different subtypes of CRC included were adenocarcinoma -NOS (64.44%) followed by mucinous signet ring type, basaloid and adenosquamous type. The Histopathological spectrum of carcinoma as per site and type are depicted inTable5. The distribution of Her2 scoring according to site of lesion is provided in Table 6 and Her2/neu scoring according to Histopathological type are given in Table 7. For ninety cases reported into Her-2neu IHC, 6 cases (6.66%) for adenomas were optimistic with a rating of 3+Figure 2, 16 cases (17.77%) of adenomas displaying an equivocal rating of 2+Figure3, two cases (2.22%) of mucinous form displayed an equi-vocal rating of 2 +., whereas 10adeno-carcinoma cases, 5 cases (5.55%) of mucinous carcinoma tworing form signet& single carcinoma basaloid was score 1+Figure 4, and twenty six adenocarcinoma cases, thirteen mucinous scenarios, three cases each of signet ring type, two cases eachof basaloid, & of adeno-squamous carcinomas found -veby rating 0. The score 2+ positive cases were subjected to FISH study.Out of 20 cases of Her2neu 2+ score 4 (33.33%) cases was found to be Her2neu 3+ score in FISH study (33.3%).Both high positive and negative score (18.88 & 40.0%) were found in cancer located in rectum.

DISCUSSION
Colonoscopy biopsy provides useful information for diagnosis of various lesion of lower gastrointestinal tract. In this study, 256 patients were evaluated, most of which were in the 41-60 years age group, mean age being 45 years showing male sex predominance of about 58.88%, while females were 41.11%. The statisticscoincided with that of Deoetal (2001) [11]whose study reported male predominance 64% and mean age of presentation being 48.5 yrs.
Colorectal carcinoma constituted 35.15% majority of which located in rectum (55.55%). The age incidence (>50 years), was similar to Boyle etal. (2002) [19]While cancers of colorectal have reported to be two times higher in males as compared with female and median age being 50 years. CRC in current study was mostly adenocarcinoma NOS type (64.44%) that correlated with Ocha M. etal (2003) [20] showing 98% of colon cancer to be adenocarcinoma, NOS type.
The Her2/neu expression was more found in left sided colon compared to rightsided colon. Strong Her2neu (score 3+) was found in 6.66% cases which were moderately differentiated adenocarcinoma.Score 2+ was observed in 22.22% of colorectalcarcinoma out of which 33.3% were found to be score 3+ on FISH study, were found mostly in high grade colon adenocarcinoma. The subtypes of adenocarcinoma like mucinous, signet ring type, cloacogenic and adenosquamous type showed negative score for Her2neu.
The percentage of positivity of Her2neu in CRC was well correlated with other authors given in Table8.

CONCLUSION
Commonest type of CRC was adenocarcinoma (NOS) followed by mucinous,adenosquamous , basaloid& signet ring form.Because of more prominent membranous staining observed in high grade colorectal cancers like breast carcinoma, Her2neu expression is found to be an important predictive marker of carcinoma colon.
Thus in a nutshell this study concludes that both conventionalas non-conventional adenocarcinoma among colorectal cancer especially those by lymphovascular inflation metastasis, Her-2 / new expression must be evaluated in form of a different therapeutic approach. Due to much stained membranesin higher grades of CRC (grade II, grade III).Treatment of Herceptinmay assists individuals suffering from diseaseespecially in adenocarcinoma,NOS category. Other poorly differentiated carcinoma (grade III) like mucinous and signet ring cell type donot well respond to Herceptin therapy.