RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES
BANGALORE, KARNATAKA
ANNEXURE – II
Proforma for Registration of Subjects for Dissertation
1 / Name and Address of the Candidate / Dr. Vidhya.JPostgraduate student
Dept. Of Oral and MaxilloFacial Pathology
AECS Maaruti College of Dental Sciences and Research Centre,
Bangalore – 560 076
2 / Name of the Institution / AECS Maaruti College of Dental Sciences and Research Centre
3 / Course of Study and Subject / Master of Dental Surgery in
Oral and MaxillaFacial Pathology
4 / Date of Admission to the Course / Aug 2013
5 / Title of the Topic / Immunohistochemical Expression of COX-2 in Normal Oral Mucosa, Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma - A comparative study.
6. Brief resume of intended work:
Introduction:
Cyclooxygenase (COX), officially known as Prostaglandin-Endoperoxide Synthase (PTGS), is an enzyme that is responsible for the formation of important biological mediators called prostanoids, including prostaglandins, prostacyclin and thromboxane. COX converts Arachidonic Acid (AA, an ω-6 PUFA) to Prostaglandin H2 (PGH2), the precursor of the series-2 prostanoids. At present, three COX isoenzymes are known: COX-1, COX-2, and COX-3. COX-1 is considered a constitutive enzyme, being found in most mammalian cells. COX-2, on the other hand, is undetectable in most normal tissues. It is an inducible enzyme, becoming abundant in activated macrophages and other cells at sites of inflammation. More recently, it has been shown to be upregulated in various carcinomas and to have a central role in tumorigenesis. A large amount of experimental data suggests that COX-2 is involved in complex processes related to cancer such as tumorigenic transformation, angiogenesis, immune response modulation, invasion and metastasis.1
The first change suggesting malignant transformation is dysplasia. Dysplasia is defined as ‘A precancerous lesion of stratified squamous epithelium characterized by cellular atypia and loss of normal maturation and stratification short of carcinoma in situ’.2 Oral epithelial dysplasia is the histopathologic marker of a premalignant disorder of the oral mucosa. It may present clinically as leukoplakia, erythroplakia, or leukoerythroplakia.
Oral Squamous Cell Carcinoma is the most common malignant epithelial tissue neoplasm of the oral cavity which is derived from the stratified squamous epithelium. Squamous cell carcinoma can be defined as “A malignant epithelium neoplasm exhibiting squamous differentiation, as characterized by the formation of keratin and / or the presence of intercellular bridges”. 3
Immunohistochemistry is defined as techniques for identifying cellular or tissue constituent (antigens) by means of antigen-antibody interaction.4
Immunohistochemical studies were done by various researchers to observe the immunoreactivity of COX-2 in OED and OSCC. The studies showed positive results in hyperplastic, dysplastic, and neoplastic oral epithelium. It was also observed that COX-2 immunoexpression increased as the severity of the lesion increased. 5
6.1 Need for the Study:
The purpose of this study is to demonstrate immunohistochemical expression of COX-2 in Normal Oral Mucosa, Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma, and to compare its expression among different grades of Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma and to evaluate its role in oral carcinogenesis.
6.2 Review of Literature:
Yuko Higashi et al (2000) investigated expression of COX-2 and Prostaglandin (PG)E2 production in two human skin epidermal cancer cell lines: Cutaneous Squamous Cell Carcinoma (HSC-5) and Eccrine Carcinoma (EcCa). Both COX-2 expression and PGE2 production were significantly enhanced in cancer cell lines compared with the non-tumorigenic human keratinocyte cell line (HaCaT). The results showed increased levels of COX-2 protein expression in skin epidermal cancer cells and increased PGE2 production proportional to COX-2 expression levels in HSC-5 and EcCa; thus, indicating that COX-2 protein expression is enhanced in skin epidermal cancer cells and that COX-2 plays a pivotal role in regulating cell growth.
Shigeto Itoh et al (2003) investigated over expression of COX-2 by immunohistochemistry in surgical specimens from patients with OSCC and evaluated the correlations of COX-2 over expression with clinicopathologic variables. The study revealed that COX-2 over expression was an independent predictor for disease free survival but not for overall survival for the patients. Thus, they suggested that over expression of COX-2 could impact on disease-free survival for patients with Squamous Cell Carcinoma and the selective inhibition of COX-2 is a possible target for therapeutic strategies.
Shadab Mohammad et al (2011), in their immunohistochemical study evaluated the prognostic significance of COX-2 over expression in Oral Squamous Cell Carcinoma (OSCC) patients undergoing chemo radiation therapy and found that COX-2, was over expressed in 15.90% cases and therefore concluded that, COX-2 expression in OSCC can be used as a prognostic marker .
Hitoshi Nagatsuka et al (2012) conducted an immunohistochemical study to investigate the expression patterns of heparanase and COX-2 during progression of OED to carcinoma. They observed that the heparanase and COX-2 messenger RNA and protein were absent in normal oral mucosa but were coexpressed in increasing intensity as OED progressed to OSCC. A strong expression of these enzymes in tumor cells at the advancing front was noticed suggesting their role in local tumor spread.
Helder Antonio Rebelo Pontes et al (2013) performed an immunohistochemical study for immunoexpression of Nuclear factor k- β (NF-k β) and COX-2 protein in OED and OSCC. The results of which showed an increased immunoexpression for NF-kβ in OSCC samples and COX-2 expression in moderate dysplasia. Thus, suggesting that COX-2 may be involved in the early stages of oral carcinogenesis process.
6.2 Objectives of the Study:
1. To evaluate the Immunohistochemical expression of COX-2 in Normal Oral Mucosa, Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma.
2. Comparing the Immunohistochemical expression of COX-2 in
(a) Normal Oral Mucosa, Oral Epithelial Dysplasia and Oral Squamous Cell
Carcinoma.
(b) Different grades of dysplasia (mild, moderate and severe dysplasia).
(c) Different grades of Oral Squamous Cell Carcinoma (well differentiated, moderately
differentiated and poorly differentiated).
7 Materials and Methods:
7.1 Source of Data:
Diagnosed Cases of different grades of Oral Epithelial Dysplasia and Oral Squamous Cell Carcinomas retrieved from the Archives of Department of Oral Pathology, AECS Maaruti College of Dental Sciences and Research center, Bangalore.
7.2 Method of collection of data
7.2.1 Sample Size:
l 10 cases of Normal oral mucosa.
l 30 cases of Oral Epithelial Dysplasia.
l 30 cases of Oral Squamous Cell Carcinoma.
7.2.2 Study Material:
· Formalin-fixed paraffin-embedded blocks of the confirmed cases.
· Cyclooxygenase-2 IHC kit.
7.2.3 Study Method:
All the samples will be immunohistochemically stained with COX-2 and analyzed to observe the membranous, cytoplasmic and nuclear expression. Labeling index will be done to calculate percentage of cells stained in five high power fields and the results will be statistically analyzed.
7.2.4 Study Design:
Retrospective comparative study.
7.3 Inclusion Criteria:
Dysplasia and tumors of any histochemical grade and originating from the oral cavity proper only.
7.4 Exclusion Criteria:
1) Secondary or metastatic tumors.
2) Normal mucosa from patients with any form of tobacco consumption habit.
7.5 Does the study require any investigation or interventions to be conducted on
patients or other humans or animals? If so, please describe briefly.
No.
7.6 Has ethical clearance been obtained from your institution for this study?
Yes.
8 List of Reference
1. Trifan O C, Hla T. Cyclooxygenase-2 modulates cellular growth and promotes tumorigenesis. J Cell Mol Med 2003; 7(3):p.207-222
2. Rastogi V, Puri N, Mishra S, Arora S, Kaur G, Yadav L. An insight to Oral Epithelial Dysplasia. Int J Head Neck Surg 2013; 4(2): p.74-82.
3. Rajendran R, Sivapathasundharam.B. Benign and malignant tumors. Shafer’s textbook of oral pathology, 5th edition 2006; p.143
4. Bankrofft J D and Gamble M. Theory and practice of histological techniques.
Churchill Livingstone publications, 5thedition 2002; p.421-464
5. Pontes H A R, Pontes F S C, Paivafonsea F, Luiz de Carvalho P, Pereira E M, Carvalho de Abreu M et al. Nuclear factor kB and Cyclooxygenase-2 immunoexpression in Oral Dysplasia and Squamous Cell Carcinoma. Annals of diagnostic pathology 2013; 17: p.45-50.
6. Higashi Y, Kanekura T, Kanzaki T. Enhanced expression of Cyclooxygenase (COX)-2 in human skin epidermal cancer cells: evidence for growth suppression by inhibiting cox-2 expression. Int. J. Cancer 2000; 86: p.667-671
7. Pestili de Almeida E M, Piche C, Sirois J, Dore M. Expression of Cyclooxygenase-2 in Naturally Occurring Squamous Cell Carcinomas in Dogs. The Journal of Histochemistry & Cytochemistry 2001; Vol 49(7):p.867-875.
8. Itoh S, Matsui K, Furuta I, Takano Y. Immunohistochemical study on over expression of cyclooxygenase-2 in squamous cell carcinoma of the oral cavity_ its importance as a prognostic predictor. Oral oncology 2003; 39: p.829-835.
9. Mohammad S, Ram H, Gupta P N, Husain N, Bhatt M L B. Over expression of COX-2 in oral squamous cell carcinoma patients undergoing chemo radiotherapy. Natl J Maxillofac Surg. 2011; 2(1):p.17-21.
10. Nagatsuka H, Siar C H, Tsujigiwa H, Naomoto Y, Han P P, Gunduz M et al. Heparanase and cyclooxygenase-2 and protein expression during progression of oral epithelial dysplasia to carcinoma. Annals of diagnostic pathology 2012; 16:p.354-361.
11. en.wikipedia.org/wiki/Cyclooxygenase
9. Signature of the Candidate:10. Remarks of the Guide:
11. Name and Designation of the Guide/s:
11.1. Guide: Dr. UMA SWAMINATHAN
ASSOCIATE PROFESSOR,
Department of Oral and Maxillafacial Pathology,
AECS Maaruti College of Dental Sciences and
Research Centre,
Bangalore – 560 076
11.2. Signature:
11.3. Co-Guide (if any):
11.4. Signature: -
11.5. Head of the Dept.: Dr. P.SHARADA
PROFESSOR & HEAD,
Department of Oral and MaxilloFacial Pathology,
AECS Maaruti College of Dental Sciences and
Research Centre,
Bangalore – 560 076
11.6. Signature:
12. Remarks of the Chairman and Principal:
12.1. Signature:
CONSENT FORM
Paraffin embedded tissue blocks of already diagnosed cases of Oral Epithelial Dysplasia, Oral Squamous Cell Carcinoma and Normal oral mucosa will be retrieved from the Archives of Dept of Oral and Maxillofacial Pathology, AECS Maaruti College of Dental Sciences and Research Centre.
The Department has no objection in issuing the blocks to the student for her dissertation work.
Dr.P.Sharada
Professor and Head
Department of Oral and Maxillofacial Pathology
AECS Maaruti College of Dental Sciences and
Research Centre
Bangalore 560076.
PATIENT CONSENT FORM
Name:
Age:
Sex:
Address:
Biopsy obtained from (site):
I hereby give my consent to the Department of Oral and Maxillofacial Pathology to use the biopsy specimen (Incisional/Excisional) for research purpose.
Signature of the patient
RAJIV GANDHI UNIVERSITY OF
HEALTH SCIENCES
BANGALORE, KARNATAKA
MASTER OF DENTAL SURGERY (MDS)
ORAL AND MAXILLOFACIAL PATHOLOGY
2013
A.E.C.S MAARUTI COLLEGE OF DENTAL SCEINCES
& RESEARCH CENTRE, BANGALORE – 560 076