Journal of Clinical and Diagnostic Research, ISSN - 0973 - 709X

Users Online : 48049

AbstractMaterial and MethodsResultsDiscussionConclusionAcknowledgementReferencesDOI and Others
Article in PDF How to Cite Citation Manager Readers' Comments (0) Audio Visual Article Statistics Link to PUBMED Print this Article Send to a Friend
Advertisers Access Statistics Resources

Dr Mohan Z Mani

"Thank you very much for having published my article in record time.I would like to compliment you and your entire staff for your promptness, courtesy, and willingness to be customer friendly, which is quite unusual.I was given your reference by a colleague in pathology,and was able to directly phone your editorial office for clarifications.I would particularly like to thank the publication managers and the Assistant Editor who were following up my article. I would also like to thank you for adjusting the money I paid initially into payment for my modified article,and refunding the balance.
I wish all success to your journal and look forward to sending you any suitable similar article in future"



Dr Mohan Z Mani,
Professor & Head,
Department of Dermatolgy,
Believers Church Medical College,
Thiruvalla, Kerala
On Sep 2018




Prof. Somashekhar Nimbalkar

"Over the last few years, we have published our research regularly in Journal of Clinical and Diagnostic Research. Having published in more than 20 high impact journals over the last five years including several high impact ones and reviewing articles for even more journals across my fields of interest, we value our published work in JCDR for their high standards in publishing scientific articles. The ease of submission, the rapid reviews in under a month, the high quality of their reviewers and keen attention to the final process of proofs and publication, ensure that there are no mistakes in the final article. We have been asked clarifications on several occasions and have been happy to provide them and it exemplifies the commitment to quality of the team at JCDR."



Prof. Somashekhar Nimbalkar
Head, Department of Pediatrics, Pramukhswami Medical College, Karamsad
Chairman, Research Group, Charutar Arogya Mandal, Karamsad
National Joint Coordinator - Advanced IAP NNF NRP Program
Ex-Member, Governing Body, National Neonatology Forum, New Delhi
Ex-President - National Neonatology Forum Gujarat State Chapter
Department of Pediatrics, Pramukhswami Medical College, Karamsad, Anand, Gujarat.
On Sep 2018




Dr. Kalyani R

"Journal of Clinical and Diagnostic Research is at present a well-known Indian originated scientific journal which started with a humble beginning. I have been associated with this journal since many years. I appreciate the Editor, Dr. Hemant Jain, for his constant effort in bringing up this journal to the present status right from the scratch. The journal is multidisciplinary. It encourages in publishing the scientific articles from postgraduates and also the beginners who start their career. At the same time the journal also caters for the high quality articles from specialty and super-specialty researchers. Hence it provides a platform for the scientist and researchers to publish. The other aspect of it is, the readers get the information regarding the most recent developments in science which can be used for teaching, research, treating patients and to some extent take preventive measures against certain diseases. The journal is contributing immensely to the society at national and international level."



Dr Kalyani R
Professor and Head
Department of Pathology
Sri Devaraj Urs Medical College
Sri Devaraj Urs Academy of Higher Education and Research , Kolar, Karnataka
On Sep 2018




Dr. Saumya Navit

"As a peer-reviewed journal, the Journal of Clinical and Diagnostic Research provides an opportunity to researchers, scientists and budding professionals to explore the developments in the field of medicine and dentistry and their varied specialities, thus extending our view on biological diversities of living species in relation to medicine.
‘Knowledge is treasure of a wise man.’ The free access of this journal provides an immense scope of learning for the both the old and the young in field of medicine and dentistry as well. The multidisciplinary nature of the journal makes it a better platform to absorb all that is being researched and developed. The publication process is systematic and professional. Online submission, publication and peer reviewing makes it a user-friendly journal.
As an experienced dentist and an academician, I proudly recommend this journal to the dental fraternity as a good quality open access platform for rapid communication of their cutting-edge research progress and discovery.
I wish JCDR a great success and I hope that journal will soar higher with the passing time."



Dr Saumya Navit
Professor and Head
Department of Pediatric Dentistry
Saraswati Dental College
Lucknow
On Sep 2018




Dr. Arunava Biswas

"My sincere attachment with JCDR as an author as well as reviewer is a learning experience . Their systematic approach in publication of article in various categories is really praiseworthy.
Their prompt and timely response to review's query and the manner in which they have set the reviewing process helps in extracting the best possible scientific writings for publication.
It's a honour and pride to be a part of the JCDR team. My very best wishes to JCDR and hope it will sparkle up above the sky as a high indexed journal in near future."



Dr. Arunava Biswas
MD, DM (Clinical Pharmacology)
Assistant Professor
Department of Pharmacology
Calcutta National Medical College & Hospital , Kolkata




Dr. C.S. Ramesh Babu
" Journal of Clinical and Diagnostic Research (JCDR) is a multi-specialty medical and dental journal publishing high quality research articles in almost all branches of medicine. The quality of printing of figures and tables is excellent and comparable to any International journal. An added advantage is nominal publication charges and monthly issue of the journal and more chances of an article being accepted for publication. Moreover being a multi-specialty journal an article concerning a particular specialty has a wider reach of readers of other related specialties also. As an author and reviewer for several years I find this Journal most suitable and highly recommend this Journal."
Best regards,
C.S. Ramesh Babu,
Associate Professor of Anatomy,
Muzaffarnagar Medical College,
Muzaffarnagar.
On Aug 2018




Dr. Arundhathi. S
"Journal of Clinical and Diagnostic Research (JCDR) is a reputed peer reviewed journal and is constantly involved in publishing high quality research articles related to medicine. Its been a great pleasure to be associated with this esteemed journal as a reviewer and as an author for a couple of years. The editorial board consists of many dedicated and reputed experts as its members and they are doing an appreciable work in guiding budding researchers. JCDR is doing a commendable job in scientific research by promoting excellent quality research & review articles and case reports & series. The reviewers provide appropriate suggestions that improve the quality of articles. I strongly recommend my fraternity to encourage JCDR by contributing their valuable research work in this widely accepted, user friendly journal. I hope my collaboration with JCDR will continue for a long time".



Dr. Arundhathi. S
MBBS, MD (Pathology),
Sanjay Gandhi institute of trauma and orthopedics,
Bengaluru.
On Aug 2018




Dr. Mamta Gupta,
"It gives me great pleasure to be associated with JCDR, since last 2-3 years. Since then I have authored, co-authored and reviewed about 25 articles in JCDR. I thank JCDR for giving me an opportunity to improve my own skills as an author and a reviewer.
It 's a multispecialty journal, publishing high quality articles. It gives a platform to the authors to publish their research work which can be available for everyone across the globe to read. The best thing about JCDR is that the full articles of all medical specialties are available as pdf/html for reading free of cost or without institutional subscription, which is not there for other journals. For those who have problem in writing manuscript or do statistical work, JCDR comes for their rescue.
The journal has a monthly publication and the articles are published quite fast. In time compared to other journals. The on-line first publication is also a great advantage and facility to review one's own articles before going to print. The response to any query and permission if required, is quite fast; this is quite commendable. I have a very good experience about seeking quick permission for quoting a photograph (Fig.) from a JCDR article for my chapter authored in an E book. I never thought it would be so easy. No hassles.
Reviewing articles is no less a pain staking process and requires in depth perception, knowledge about the topic for review. It requires time and concentration, yet I enjoy doing it. The JCDR website especially for the reviewers is quite user friendly. My suggestions for improving the journal is, more strict review process, so that only high quality articles are published. I find a a good number of articles in Obst. Gynae, hence, a new journal for this specialty titled JCDR-OG can be started. May be a bimonthly or quarterly publication to begin with. Only selected articles should find a place in it.
An yearly reward for the best article authored can also incentivize the authors. Though the process of finding the best article will be not be very easy. I do not know how reviewing process can be improved. If an article is being reviewed by two reviewers, then opinion of one can be communicated to the other or the final opinion of the editor can be communicated to the reviewer if requested for. This will help one’s reviewing skills.
My best wishes to Dr. Hemant Jain and all the editorial staff of JCDR for their untiring efforts to bring out this journal. I strongly recommend medical fraternity to publish their valuable research work in this esteemed journal, JCDR".



Dr. Mamta Gupta
Consultant
(Ex HOD Obs &Gynae, Hindu Rao Hospital and associated NDMC Medical College, Delhi)
Aug 2018




Dr. Rajendra Kumar Ghritlaharey

"I wish to thank Dr. Hemant Jain, Editor-in-Chief Journal of Clinical and Diagnostic Research (JCDR), for asking me to write up few words.
Writing is the representation of language in a textual medium i e; into the words and sentences on paper. Quality medical manuscript writing in particular, demands not only a high-quality research, but also requires accurate and concise communication of findings and conclusions, with adherence to particular journal guidelines. In medical field whether working in teaching, private, or in corporate institution, everyone wants to excel in his / her own field and get recognised by making manuscripts publication.


Authors are the souls of any journal, and deserve much respect. To publish a journal manuscripts are needed from authors. Authors have a great responsibility for producing facts of their work in terms of number and results truthfully and an individual honesty is expected from authors in this regards. Both ways its true "No authors-No manuscripts-No journals" and "No journals–No manuscripts–No authors". Reviewing a manuscript is also a very responsible and important task of any peer-reviewed journal and to be taken seriously. It needs knowledge on the subject, sincerity, honesty and determination. Although the process of reviewing a manuscript is a time consuming task butit is expected to give one's best remarks within the time frame of the journal.
Salient features of the JCDR: It is a biomedical, multidisciplinary (including all medical and dental specialities), e-journal, with wide scope and extensive author support. At the same time, a free text of manuscript is available in HTML and PDF format. There is fast growing authorship and readership with JCDR as this can be judged by the number of articles published in it i e; in Feb 2007 of its first issue, it contained 5 articles only, and now in its recent volume published in April 2011, it contained 67 manuscripts. This e-journal is fulfilling the commitments and objectives sincerely, (as stated by Editor-in-chief in his preface to first edition) i e; to encourage physicians through the internet, especially from the developing countries who witness a spectrum of disease and acquire a wealth of knowledge to publish their experiences to benefit the medical community in patients care. I also feel that many of us have work of substance, newer ideas, adequate clinical materials but poor in medical writing and hesitation to submit the work and need help. JCDR provides authors help in this regards.
Timely publication of journal: Publication of manuscripts and bringing out the issue in time is one of the positive aspects of JCDR and is possible with strong support team in terms of peer reviewers, proof reading, language check, computer operators, etc. This is one of the great reasons for authors to submit their work with JCDR. Another best part of JCDR is "Online first Publications" facilities available for the authors. This facility not only provides the prompt publications of the manuscripts but at the same time also early availability of the manuscripts for the readers.
Indexation and online availability: Indexation transforms the journal in some sense from its local ownership to the worldwide professional community and to the public.JCDR is indexed with Embase & EMbiology, Google Scholar, Index Copernicus, Chemical Abstracts Service, Journal seek Database, Indian Science Abstracts, to name few of them. Manuscriptspublished in JCDR are available on major search engines ie; google, yahoo, msn.
In the era of fast growing newer technologies, and in computer and internet friendly environment the manuscripts preparation, submission, review, revision, etc and all can be done and checked with a click from all corer of the world, at any time. Of course there is always a scope for improvement in every field and none is perfect. To progress, one needs to identify the areas of one's weakness and to strengthen them.
It is well said that "happy beginning is half done" and it fits perfectly with JCDR. It has grown considerably and I feel it has already grown up from its infancy to adolescence, achieving the status of standard online e-journal form Indian continent since its inception in Feb 2007. This had been made possible due to the efforts and the hard work put in it. The way the JCDR is improving with every new volume, with good quality original manuscripts, makes it a quality journal for readers. I must thank and congratulate Dr Hemant Jain, Editor-in-Chief JCDR and his team for their sincere efforts, dedication, and determination for making JCDR a fast growing journal.
Every one of us: authors, reviewers, editors, and publisher are responsible for enhancing the stature of the journal. I wish for a great success for JCDR."



Thanking you
With sincere regards
Dr. Rajendra Kumar Ghritlaharey, M.S., M. Ch., FAIS
Associate Professor,
Department of Paediatric Surgery, Gandhi Medical College & Associated
Kamla Nehru & Hamidia Hospitals Bhopal, Madhya Pradesh 462 001 (India)
E-mail: drrajendrak1@rediffmail.com
On May 11,2011




Dr. Shankar P.R.

"On looking back through my Gmail archives after being requested by the journal to write a short editorial about my experiences of publishing with the Journal of Clinical and Diagnostic Research (JCDR), I came across an e-mail from Dr. Hemant Jain, Editor, in March 2007, which introduced the new electronic journal. The main features of the journal which were outlined in the e-mail were extensive author support, cash rewards, the peer review process, and other salient features of the journal.
Over a span of over four years, we (I and my colleagues) have published around 25 articles in the journal. In this editorial, I plan to briefly discuss my experiences of publishing with JCDR and the strengths of the journal and to finally address the areas for improvement.
My experiences of publishing with JCDR: Overall, my experiences of publishing withJCDR have been positive. The best point about the journal is that it responds to queries from the author. This may seem to be simple and not too much to ask for, but unfortunately, many journals in the subcontinent and from many developing countries do not respond or they respond with a long delay to the queries from the authors 1. The reasons could be many, including lack of optimal secretarial and other support. Another problem with many journals is the slowness of the review process. Editorial processing and peer review can take anywhere between a year to two years with some journals. Also, some journals do not keep the contributors informed about the progress of the review process. Due to the long review process, the articles can lose their relevance and topicality. A major benefit with JCDR is the timeliness and promptness of its response. In Dr Jain's e-mail which was sent to me in 2007, before the introduction of the Pre-publishing system, he had stated that he had received my submission and that he would get back to me within seven days and he did!
Most of the manuscripts are published within 3 to 4 months of their submission if they are found to be suitable after the review process. JCDR is published bimonthly and the accepted articles were usually published in the next issue. Recently, due to the increased volume of the submissions, the review process has become slower and it ?? Section can take from 4 to 6 months for the articles to be reviewed. The journal has an extensive author support system and it has recently introduced a paid expedited review process. The journal also mentions the average time for processing the manuscript under different submission systems - regular submission and expedited review.
Strengths of the journal: The journal has an online first facility in which the accepted manuscripts may be published on the website before being included in a regular issue of the journal. This cuts down the time between their acceptance and the publication. The journal is indexed in many databases, though not in PubMed. The editorial board should now take steps to index the journal in PubMed. The journal has a system of notifying readers through e-mail when a new issue is released. Also, the articles are available in both the HTML and the PDF formats. I especially like the new and colorful page format of the journal. Also, the access statistics of the articles are available. The prepublication and the manuscript tracking system are also helpful for the authors.
Areas for improvement: In certain cases, I felt that the peer review process of the manuscripts was not up to international standards and that it should be strengthened. Also, the number of manuscripts in an issue is high and it may be difficult for readers to go through all of them. The journal can consider tightening of the peer review process and increasing the quality standards for the acceptance of the manuscripts. I faced occasional problems with the online manuscript submission (Pre-publishing) system, which have to be addressed.
Overall, the publishing process with JCDR has been smooth, quick and relatively hassle free and I can recommend other authors to consider the journal as an outlet for their work."



Dr. P. Ravi Shankar
KIST Medical College, P.O. Box 14142, Kathmandu, Nepal.
E-mail: ravi.dr.shankar@gmail.com
On April 2011
Anuradha

Dear team JCDR, I would like to thank you for the very professional and polite service provided by everyone at JCDR. While i have been in the field of writing and editing for sometime, this has been my first attempt in publishing a scientific paper.Thank you for hand-holding me through the process.


Dr. Anuradha
E-mail: anuradha2nittur@gmail.com
On Jan 2020

Important Notice

Original article / research
Year : 2021 | Month : August | Volume : 15 | Issue : 8 | Page : EC01 - EC04 Full Version

Expression of B-type RAF V600E in Thyroid Carcinoma and its Association with Histological Type


Published: August 1, 2021 | DOI: https://doi.org/10.7860/JCDR/2021/46250.15184
Sonia Kumari, Turuvekere Narayanrao Suresh, SM Azeem Mohiyuddin

1. Junior Resident, Department of Pathology, Sri Devaraj Urs Medical College, Kolar, Karnataka, India. 2. Professor, Department of Pathology, Sri Devaraj Urs Medical College, Kolar, Karnataka, India. 3. Professor, Department of ENT, Sri Devaraj Urs Medical College, Kolar, Karnataka, India.

Correspondence Address :
Dr. Turuvekere Narayanrao Suresh,
Professor, Department of Pathology, Sri Devaraj Urs Medical College, Tamaka, Kolar, Karnataka, India.
E-mail: sureshstn@rediffmail.com

Abstract

Introduction: Thyroid cancer is one of the most common cancers amongst all endocrine cancers. Incidence of thyroid malignancy is about 3-4% of all malignancy in India and 80% of thyroid malignancy belongs to Papillary Thyroid Carcinoma (PTC). Factors affecting the prognosis of PTC include patient’s gender, age, histological findings, tumour size, lymph node metastasis, extrathyroidal extension, and remote metastasis. Presence of B-type RAF V600E (BRAF V600E) mutation in thyroid carcinoma patients tends to present with more aggressive clinicopathological behaviours of PTC, prompting more aggressive radioiodine treatment.

Aim: To find out the frequency of occurrence and expression of BRAF V600E mutation by Immunohistochemistry (IHC) in thyroid cancer and its association with histological type and Tumour Nodes Metastases (TNM) Staging.

Materials and Methods: The present observational retrospective study included patients treated for thyroid carcinoma between January 2014 to February 2019 at RL Jalappa hospital and Research centre, Kolar, Karnataka, India. The IHC was done with rabbit monoclonal anti-BRAF V600E antibody IgG Clone RM8 (VE1). Clinical records, Fine Needle Aspiration Cytology (FNAC) diagnosis were analysed for 45 thyroid carcinoma cases. Immunopositivity was scored positive when unambiguous clear cytoplasmic staining for the antibody was observed in tumour cells. Categorical data was presented in the form of frequencies and proportions and continuous data was presented as mean and standard deviation. The t-test were applied to find out the difference in means among the groups. The p-value <0.05 was considered statistically significant.

Results: Out of total 45 cases, 18 were classical PTC, 18 were Follicular Variant of Papillary Thyroid Cancer (FV-PTC), 4 were micropapillary carcinomas, 3 were Follicular carcinomas, 1 was Oncocytic variant of PTC and 1 undifferentiated thyroid carcinoma. Out of total 45 cases, 33 cases (73%) were found to be BRAF V600E positive and 12 (26%) were negative for BRAF V600E. Out of the total 33 BRAF V600E positive cases, 19 cases showed strong staining, 14 cases showed moderately positive staining.

Conclusion: The PTC is the most frequent type of thyroid carcinoma amongst all the sub-types. BRAF V600E expression is commonly seen in higher tumour size (T3, T4) and classical PTC. Tumours with extrathyroidal extension and capsular invasion showed strong positivity.

Keywords

Histopathology, Immunohistochemistry, Molecular pathology

Thyroid cancer is one of the most common cancers amongst all endocrine cancers. Incidence of thyroid malignancy is about 3-4% of all malignancy in India and 80% of thyroid malignancy belongs to PTC (1). Factors affecting the prognosis of PTC include patient’s gender, age, histological findings, tumour size, lymph node metastasis, extrathyroidal extension and remote metastasis. There are numerous variants of Raf kinase. The most periodically found genetic aberration in PTC is BRAF V600E mutation. Literature states that BRAF gene mutations affect the factors that predict extrathyroidal extension, lymph node metastasis, disease recurrence, higher tumour stages 3 and 4 (2),(3). Role of BRAF mutations in the pathogenesis of tumours in malignant melanoma, ovarian tumours and colorectal carcinoma has been studied (2),(4).

The BRAF V600E mutation has a vital role in infancy stage of tumour where it induces high proliferation of the tumour cells. It is also involved in the aggressiveness and tumour undifferentiation (5). Retrospectively analysis of patients who were on higher doses of radioiodine treatments were found to be positive for BRAF V600E (6). This likely reflects that BRAF V600E patients tends to present with more aggressive clinicopathological behaviours of PTC, prompting more aggressive radioiodine treatment. BRAF studies are reported mainly from western countries and only limited publications are from India (3),(7). Gold standard for detection of BRAF V600E mutation detection is molecular genetics study using Polymerase Chain Reaction (PCR) and Deoxyribonucleic Acid (DNA) sequencing. Only few IHC identification methods of BRAF V600E mutation using formalin fixed paraffin embedded tissue sections is reported in the literature (8),(9),(10),(11),(12),(13) in which the histological type was not correlated with BRAF expression.

This study was conducted to assess the frequency of occurrence and expression of BRAF V600E mutation by IHC in thyroid cancer and its association with histological type.

Material and Methods

The present observational retrospective study was conducted at RL Jalappa Hospital and Research Centre, Kolar, Karnataka, India from January 2014 to February 2019. Case analysis was done between May to July 2019.

All the methods in the study were in accordance with the standards of the Ethical Committee on human experimentation with Ethical clearance number SDUMC/KLR/IEC/44/2019-2020 for starting the study. Ethical clearance was also obtained for publishing this study with the Ethical clearance number SDUMC/KLR/IEC/81/2020-2021.

Sample size calculation: The sample size was calculated using the following formula:

n= Z2 (p×q) / d2

Where, Z=1.96; it is standard deviation score for 95% set interval
p= assumed or estimated proportion (77%)=0.77 (14).
q=1-P (1-0.77)=0.23
d=allowable error (20% of P)=0.15

The sample size came to be 31 subjects at 95% confidence limit and 20% allowable error after assuming the percentage of BRAF positive expression in thyroid carcinoma in 77% of subjects in a similar study done by AbdElmageed ZY et al., (14). However, in this study 45 cases were studied.

Inclusion criteria: All patients who underwent thyroidectomy from January 2014 to February 2019 with or without therapeutic neck dissection for thyroid carcinoma were subjected to histopathological examination.

Exclusion criteria: Cases showing extensive tumour necrosis without sufficient viable tumour cells and patients who received chemotherapy were excluded from the study.

Study Procedure

The IHC for the BRAF VE1 antibody was performed on formalin-fixed, paraffin-embedded 5 μm thick sections which were mounted on super frost slides and dried in an oven at 60°C for one hour to ensure optimal adhesion. Melanoma and colorectal carcinoma slides were used as positive control. Non biotin polymer-based Horseradish Peroxidase (HRP) detection system was used. Antigen retrieval was performed in microwave at power 10 for 6 minutes in Citrate buffer. Endogenous peroxidase was inhibited by adding 0.3% H2O2 to slides and incubating for 10 minutes. Sections were incubated with primary antibody (Rabbit antihuman BRAF V600E monoclonal antibody; clone VE1 Bio SB, California USA) diluted at 1:70 for 120 minutes in a humid chamber followed by three washes with Tris-Buffered Saline (TBS) at pH 7.6, and then incubated for 120 minutes with prediluted secondary antibody. Next, the slides were washed again with TBS buffer. Finally, 3,3’-diaminobenzidine chromogenic solution (enhancer) was applied for three minutes, and the slides were counter stained with haematoxylin for five seconds. IHC staining assessment was done as per Spring bioscience method (Table/Fig 1) (10),(14). Weak (1+) or no staining was considered as negative (Table/Fig 2). Positive staining was considered when cytoplasmic staining of the tumour cells was moderate 2+ (Table/Fig 3) to strong 3+ (Table/Fig 4).

Statistical Analysis

Categorical data was represented in the form of frequencies and proportions. Continuous data was represented as mean and standard deviation. Chi-square test was used as test of significance. The p-value <0.05 was considered statistically significant. Data was analysed after entering into Microsoft Excel sheet using IBM Statistical Package for Social Sciences (SPSS) (trial version 23) and PRIMER statistical software.

Results

Majority of the patients were females with a male to female ratio of 1:5 having a mean age of 40.56±11.86 years (ranging from 19 to 67 years). Majority of the cases 38 (84.4%) were BETHESDA category V on FNAC, while 7 (15.5%) cases belonged to BETHESDA category IV. Clinically, majority of the nodules 34 (75%) were solitary thyroid swellings, rest of the cases presented with diffuse swellings. Out of total 45 cases, 18 (40%) were classical PTC, 18 (40%) were FV-PTC, 4 (8.89%) were micropapillary carcinomas, 3 (6.67%) were follicular carcinomas, 1 (2.22%) was oncocytic variant of PTC and 1 (2.22%) undifferentiated thyroid carcinoma. Out of total 45 cases studied; 33 cases were positive for BRAF V600E antibody and 12 cases were negative. BRAF positivity is seen more frequently in classical PTC that is 17 out of 18 cases (94%), as compared to FV-PTC that is 10 out of 18 (55%) cases (Table/Fig 5). Out of total 33 BRAF positive cases, strong expression of BRAF (3+ staining) was observed in 19 (58%) cases (Table/Fig 4) while 14 (42%) cases showed 2+ staining (Table/Fig 3). Among BRAF positive cases, it was found that most of the patients (55%) were females of the age group of 40 years and above.

Majority of positive cases i.e., 22 (66.67%) were T3 Stage. All 4 cases of T4 showed BRAF positivity (Table/Fig 6).

Extrathyroidal extension/capsular invasion was seen in six cases. Out of these six cases, 5 (83%) showed 3+ BRAF staining. Cervical lymph node dissection was done for 9 (20%) cases out of which 7 (15%) cases showed lymph node metastasis. Out of 7 cases with lymph node metastasis, 6 (86%) showed strong BRAF V600E (3+) staining in thyroid tumour tissue and 1 case (14%) was negative for BRAF staining.

Discussion

The most common genetic alteration reported in PTC is BRAF V600E mutation and found to be associated with an aggressive phenotype. BRAF mutations reported in PTC ranges from 5-90% (2). This variation may be due to temporal differences in PTC pathogenesis and these differences may be due to different procedures (frozen tissue versus paraffin embedded tissue) and different techniques of detection (microdissection, direct sequencing, pyro sequencing, and microdissection) and different sensitivity and specificity. IHC staining is a technique that is widely used in diagnostic pathology laboratories and in contrast to molecular techniques, it is rapid and economical. It also provides the ability to easily quantify the BRAF V600E mutation and has the added advantage of allowing visualisation of individual antigen-bearing tumour cells and assessing for tumour homogeneity and heterogeneity.

Mutated BRAF protein occurs early in carcinogenesis of PTC with no significant difference between macro-carcinomas and micro-carcinomas (15). It is also associated with diverse aggressive subtypes of PTC, more commonly seen in tall cell variants, oncocytic variants including Warthin tumour like variant (2),(3),(6),(7). In contrast, an exclusively follicular growth was predictive for the absence of mutated BRAF protein. Similarly, in present study BRAF positivity is seen more frequently in classical PTC cases (94%) as compared to FV-PTC (55%). Researchers found that solid variants and even focal presence of solid growth pattern is inversely related to BRAF protein expression because of different BRAF mutation in solid tumours being on exon 15 of the BRAF gene which is different from the typical T1799A BRAF mutation (16).

In this study, BRAF mutation is more commonly seen in >40-year females (55%). In this study, total 33 (73%) of the cases were positive for BRAF V600E mutation. BRAF mutation was seen in aggressive tumour type and high BRAF V600E intensity staining in tumour with extrathyroidal extension in six cases, out of which 5 (83%) of cases showed strong 3+ staining. It was also found to be associated with vascular invasion in 36 (80%) of the cases similar to the findings by Zagzag J et al., (12). Similar to other studies association of BRAF V600E positivity with higher tumour stage T3 and T4 was observed in this study (Table/Fig 7) (3),(10),(11),(12),(14),(17).

The expression of BRAF mutation will be of high clinical interest especially in therapy-resistant cases of PTC, as targeted therapy inhibiting mutated BRAF V600E protein is under the clinical trial investigation as FDA approved BRAF inhibitor vemurafenib (18),(19),(20). Other additional advantages of BRAF detection by IHC include usage on small tumours, micro-metastasis in a lymph node. BRAF tests have got a higher sensitivity of 81.25% when tested for preoperative immunocytochemistry. Hence, the potential of this diagnostic test can be utilised for indeterminate thyroid nodule (21). An advantage of IHC over genetic analysis alone is that translated protein products and post-translational regulatory modifications such as activating phosphorylation of kinases can be qualitatively assessed (22).

Patients whose tumours displayed the molecular high-risk group of BRAF V600E positive IHC had quadruple the risk of recurrence compared to the low-risk group of BRAF V600E negative cases (22). Meta-analysis showed BRAF independently predicted PTC recurrence (23).

Targeted therapy for BRAF positive thyroid carcinoma patients: Vemurafenib and dabrafenib are the BRAF V600E inhibitors that function as ATP-Competitive inhibitors. These targeted therapies are highly suggested in recurrent or metastatic radio-iodine refractory PTC patients to reduce the rate of progression of disease and increasing the disease free survival (20).

Limitation(s)

In view of small sample size, multicentric study with large sample size is suggested for further validation of prognostic role of BRAF in thyroid carcinoma.

Conclusion

The BRAF V600E expression is strongly associated with higher tumour size (T3, T4) and classical PTC. It is also associated with extrathyroidal extension and vascular invasion showing strong positivity. In view of strong BRAF expression in classical PTC, BRAF V600E immunostaining may be applied as “Rule In” diagnostic and prognostic marker in FNAC of suspicious thyroid nodules.

Acknowledgement

The authors would also like to extend gratitude to Dr. Kalyani R, Professor and Head, Department of Pathology and Dr. PN Sreeramulu, Dean and Professor of Surgery, Sri Devaraj Urs Medical College, Kolar, Tamaka, Karnataka for encouragement to complete this study.

References

1.
Yeole BB. Descriptive epidemiology of thyroid cancer in greater Bombay. Indian J Cancer. 1998;35:57-64. [crossref] [PubMed]
2.
Ritterhouse LL, Barletta JA. BRAF V600E mutation-specific antibody: A review. Semin Diagn Pathol. 2015;32:400-08. [crossref] [PubMed]
3.
Chakraborty A, Narkar A, Mukhopadhyaya R, Kane S, D’Cruz A, Rajan MG. BRAF V600E mutation in papillary thyroid carcinoma: significant association with node metastases and extra thyroidal invasion. Endocr Pathol. 2012;23:83-93. [crossref] [PubMed]
4.
Ghasemi M, Larijani VL, Emadian O, Yazdani J, Sajadianfar A, Abediankenari S. Immunohistochemical investigation of mutant BRAF V600E in common pigmented skin neoplasms, study on a sample of Iranian Patients. Iran J Pathol. 2019;14:08-16. [crossref] [PubMed]
5.
McKelvie PA, Chan F, Yu Y. The prognostic significance of the BRAF V600E mutation in papillary thyroid carcinoma detected by mutation- Specific immunohistochemistry. Pathology. 2013;45:637-44. [crossref] [PubMed]
6.
Kim TY, Kim WB, Rhee YS, Song JY, Kim JM, Gong G, et al. The BRAF mutation is useful for prediction of clinical recurrence in low-risk patients with conventional papillary thyroid carcinoma. Clinical Endocrinology. 2006;65:364-68. [crossref] [PubMed]
7.
Krishnamurthy A, Ramshankar V, Murherkar K, Vidyarani S, Raghunandhan GC, Das A. Role and relevance of BRAF mutations in risk stratifying patients of papillary thyroid cancers along with a review of literature. Indian J Cancer. 2017;54:372-78. [crossref] [PubMed]
8.
Bullock M, O’Neill C, Chou A, Clarkson A, Dodds T, Toon C, et al. Utilisation of a MAB for BRAF(V600E) detection in papillary thyroid carcinoma. Endocr Relat Cancer. 2012;19:779-84. [crossref] [PubMed]
9.
Crescenzi A, Guidobaldi L, Nasrollah N. Immunohistochemistry for BRAF(V600E) antibody VE1 performed in core needle biopsy samples identifies mutated papillary thyroid cancers. Horm Metab Res. 2014;46:370-74. [crossref] [PubMed]
10.
Koperek O, Kornauth C, Capper D. Immunohistochemical detection of the BRAF V600E-mutated protein in papillary thyroid carcinoma. Am J Surg Pathol. 2012;36:844-50. [crossref] [PubMed]
11.
McKelvie AP, Chan F, Yu Y, Waring P, Gresshoff I, Farell S, et al. The prognostic significance of the BRAF V600E mutation in papillary thyroid by mutation-specific immunohistochemistry. Pathology. 2013;45:637-44. [crossref] [PubMed]
12.
Zagzag J, Pollack A, Dultz L, Dhar S, Ogilvie JB, Heller KS, et al. Clinical utility of immunohistochemistry for the detection of the BRAF v600e mutation in papillary thyroid carcinoma. Surgery. 2013;154:1199-204. [crossref] [PubMed]
13.
Routhier CA, Mochel MC, Lynch K, Dias-Santagata D, Louis DN, Hoang MP. Comparison of monoclonal antibodies for immunohistochemical detection of BRAF V600E mutation in malignant melanoma, pulmonary carcinoma, gastrointestinal carcinoma, thyroid carcinoma, and gliomas. Human Pathol. 2013;44:2563-70. [crossref] [PubMed]
14.
AbdElmageed ZY, Sholl AB, Tsumagari K, Al-Qurayshi Z, Basolo F, Moroz K, et al. Immunohistochemistry as an accurate tool for evaluating BRAF-V600E mutation in 130 samples of papillary thyroid cancer. Surgery. 2017;161:1122-28. [crossref] [PubMed]
15.
Capper D, Preusser M, Habel A, Sahm F, Ackermann U, Schindler G. Assessment of BRAF V600E mutation status by immunohistochemistry with a mutation-specific monoclonal antibody. Acta Neuropathol. 2011;122:11-19. [crossref] [PubMed]
16.
Trovisco V, Soares P, Soares R, Magalhães J, Sá-Couto P, Sobrinho-Simões M. A new BRAF gene mutation detected in a case of a solid variant of papillary thyroid carcinoma. Hum Pathol. 2005;36:694-97. [crossref] [PubMed]
17.
Li F, Chen G, Sheng C, Gusdon AM, Huang Y, Lv Z. BRAF V600E mutation in papillary thyroid microcarcinoma: A meta-analysis. Endocr Relat Cancer. 2015;22:159-68. [crossref] [PubMed]
18.
Lin AJ, Samson P, De Wees T, Henke L, Baranski T, Schwarz J, et al. A molecular approach combined with American Thyroid Association classification better stratifies recurrence risk of classic histology papillary thyroid cancer. Cancer Med. 2019;8:437-46. [crossref] [PubMed]
19.
Kim BK, Cabanillas EM, Lazar JA, Williams DM, Sanders LD, Ilagan LJ, et al. Clinical responses to vemurafenibin patients with metastatic papillary thyroid cancer harbouring BRAF(V600E) mutation. Thyroid. 2013;23:1277-83. [crossref] [PubMed]
20.
Rosove MH, Peddi PF, Glaspy JA. BRAF V600E inhibition in anaplastic thyroid cancer. N Engl J Med. 2013;368:684-85. [crossref] [PubMed]
21.
Laha D, Nilubol N, Boufraqech M. New therapies for advanced thyroid cancer. Front Endocrinol. 2020;22;11:82. [crossref] [PubMed]
22.
Sudarsa IW, Pualillin EDK, Adiputra PAT, Manuaba IBTW. Immunocytochemistry test of protein BRAF expression for diagnosis of well differentiated thyroid carcinoma. Case Rep Oncol. 2018;11:843-49. [crossref] [PubMed]
23.
Li X, Kwon H. The impact of BRAF mutation on the recurrence of papillary thyroid carcinoma: A meta-analysis. Cancers (Basel). 2020;12:2056. [crossref] [PubMed]

DOI and Others

10.7860/JCDR/2021/46250.15184

Date of Submission: Aug 09, 2020
Date of Peer Review: Sep 25, 2020
Date of Acceptance: Apr 22, 2021
Date of Publishing: Aug 01, 2021

AUTHOR DECLARATION:
• Financial or Other Competing Interests: None
• Was Ethics Committee Approval obtained for this study? Yes
• Was informed consent obtained from the subjects involved in the study? Yes
• For any images presented appropriate consent has been obtained from the subjects. Yes

PLAGIARISM CHECKING METHODS:
• Plagiarism X-checker: Aug 17, 2020
• Manual Googling: Mar 04, 2021
• iThenticate Software: May 21, 2021 (19%)

ETYMOLOGY: Author Origin

JCDR is now Monthly and more widely Indexed .
  • Emerging Sources Citation Index (Web of Science, thomsonreuters)
  • Index Copernicus ICV 2017: 134.54
  • Academic Search Complete Database
  • Directory of Open Access Journals (DOAJ)
  • Embase
  • EBSCOhost
  • Google Scholar
  • HINARI Access to Research in Health Programme
  • Indian Science Abstracts (ISA)
  • Journal seek Database
  • Google
  • Popline (reproductive health literature)
  • www.omnimedicalsearch.com