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

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Dr Mohan Z Mani

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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 : 2023 | Month : June | Volume : 17 | Issue : 6 | Page : EC06 - EC09 Full Version

Expression of PD-L1 in Microsatellite Instability High Tumours: A Retrospective Study


Published: June 1, 2023 | DOI: https://doi.org/10.7860/JCDR/2023/63627.18029
Dinisha Einstien, Gayathri Devi Thanigaimani, KS Mouleeswaran, A Prathiba, G Sarumathy, John Vergilin

1. Associate Professor, Department of Pathology, Panimalar Medical College Hospital and Research Institute, Chennai, Tamil Nadu, India. 2. Associate Professor, Department of Pathology, Panimalar Medical College Hospital and Research Institute, Chennai, Tamil Nadu, India. 3. Associate Professor, Department of Pathology, Tagore Medical College and Hospital, Chennai, Tamil Nadu, India. 4. Associate Professor, Department of Pathology, Panimalar Medical College Hospital and Research Institute, Chennai, Tamil Nadu, India. 5. Associate Professor, Department of Pathology, Panimalar Medical College Hospital and Research Institute, Chennai, Tamil Nadu, India. 6. Registrar, Department of Pathology, Gleneagles Global Health City, Chennai, Tamil Nadu, India.

Correspondence Address :
Dr. A Prathiba,
Associate Professor, Department of Pathology, Panimalar Medical College Hospital and Research Institute, Varadharajapuram, Outer Ring Road, Poonamallee, Chennai-600123, Tamil Nadu, India.
E-mail: acs.path@gmail.com

Abstract

Introduction: Microsatellite Instability (MSI) is the hallmark of Lynch syndrome/Constitutional Mismatch Repair Deficiency (CMMRD) and is also found in many sporadic cancers like colorectal cancer, endometrial, gastric, small intestine, urothelial, central nervous system and sebaceous gland neoplasms. MSI is a predictive biomarker for immunotherapy and Immunohistochemistry (IHC) antibodies against four Mismatch Repair (MMR) proteins: MutL Homolog 1 (MLH1), MutS Homolog 2 (MSH2), MutS Homolog 6 (MSH6) and Postmeiotic Segregation increased 2 (PMS2) can identify the MSI status of the tumour. In addition to MSI, immune checkpoint Programmed Cell Death Protein 1 (PD-1) expression and its ligand PD-L1 are biomarkers that can predict response to immunotherapy. Considering this increasing interest to identify deficient MMR (dMMR) status in different cancers, authors have assessed the expression of PD-L1 and status of MSI in various cancer types.

Aim: To evaluate the expression of PD-L1 in MSI-high status tumours.

Materials and Methods: This retrospective cross-sectional study was done in the Department of Pathology, Panimalar Medical College Hospital and Research Institute, Chennai, Tamil Nadu, India for a period of six months from July 2022 to December 2022. A total of 151 cases were identified for the period of three years from January 2020 to December 2022. The slides and blocks were retrieved from the archives. Tumour sections from the paraffin embedded tissues were deparaffinised and antigen retrieval was done. IHC using four antibodies (MLH1, MH2, MSH6 and PMS2) was performed on these slides to assess the MSI status. The slides were reviewed and were further subjected to PD-L1 IHC. PD-L1 expression on tumour cells was compared with the MSI status of different cancer types. The p-value was calculated using t-test and p<0.5 was considered statistically significant. Statistical analysis was done using International Business Machines (IBM) Statistical Package for the Social Sciences (SPSS) software version 21.0.

Results: A total of 151 cases were included in the present study. A positive nuclear stain for the four MMR proteins denote expression of wild type MMR proteins, hence MSI low status. A loss of nuclear expression denotes mutation of MMR proteins and hence MSI high (MSI-H) status. The MSI was high in nine out of 48 cases (18.75%) of colon cancer, three out of 15 cases (20%) of endometrial cancers, three out of 20 cases (15%) of gastric cancers. Rest were one out of sixteen cases of ovarian cancer, one out of two malignant melanoma and one out of three glioblastoma. Out of these, PD-L1 was positive in seven of the 18 MSI-H cases (38.88% of MSI-H cases). The MSI was low/stable in the remaining 133 cases. The p-value of significance was 0.03 (statistically significant).

Conclusion: This study shows a significant association of MSI-H with PD-L1 expression in tumours. Further large scale studies can help in assessing the role of PD-L1 as an effective therapeutic biomarker in MSI-high status patients who can benefit from targeted therapy.

Keywords

Lynch syndrome, Mismatch repair deficiency, Mismatch repair proteins, Programmed cell death protein ligand 1

The accumulation of dMMR mutations in monomorphic microsatellites as short tandem repeats which are prone to mismatch errors is termed as MSI. Although, MSI is the hallmark of Lynch syndrome and CMMRD, it has been found in many sporadic cancers belonging to this spectrum, such as gastric, small intestinal, colorectal, endometrial, urothelial, central nervous system and sebaceous gland neoplasms (1),(2). Various studies suggest that MSI is a predictive biomarker for immunotherapy (3),(4). MSI can be identified by the use of IHC and molecular tests like Polymerase Chain Reaction (PCR) and Next-Generation Sequencing (NGS). IHC uses antibodies against four MMR proteins: MLH1, MSH2, MSH6 and PMS2. The immune checkpoint PD-1 expression and its ligand PD-L1 are also biomarkers that can predict response to immunotherapy (5),(6) and can be identified by the use of IHC on tumour sections. Considering this increase in the interest to identify dMMR status in different cancers, authors have assessed the expression of PD-L1 and status of MSI in various cancer types. This can provide valuable information to identify patients with MSI high status and PD-L1 expression, who may be potential candidates for targeted therapy. Therefore, the aim of the study was to assess the MSI status of different cancer types and to assess the distribution of PD-L1 expression of these cancers with the MSI status.

Material and Methods

This retrospective cross-sectional study was done in the Department of Pathology, Panimalar Medical College Hospital and Research Institute, Chennai, Tamil Nadu, India. for a period of six months from July 2022 to December 2022. A total of 151 cancer cases were identified for the period of three years from January 2020 to December 2022. The slides and blocks were retrieved from the archives.

Inclusion criteria: Biopsies and resection specimens of cancer cases with adequate tissue; blocks with adequate representation of tumour. Tumours of the gastrointestinal tract, lung, thyroid, breast, endometrium, ovary, cervix, malignant melanoma and glioblastoma were included in the present study.

Exclusion criteria: Inadequate tissue for sectioning; inadequate representation of tumour; cytoplasmic staining of MSI by tumour cells; positive PD-L1 expression by immune cells in normal mucosa, ulcerated areas and adenoma; pale cytoplasmic staining of PD-L1 were excluded from the study.

Study Procedure

The IHC using four antibodies {Mouse monoclonal antibody MLH1 (Clone: GM011), Rabbit monoclonal antibody MSH2 (clone: RED2), Rabbit monoclonal antibody MSH6 (Clone: EP49) and Rabbit monoclonal antibody PMS2 (clone: EP51)} was performed on these slides to assess the MSI status. The slides were reviewed and were further subjected to PD-L1 IHC (Clone SP263). The tumour sections from paraffin embedded tissues were taken on polylysine coated slides, incubated overnight at 37oC and deparaffinised. After antigen retrieval in Tris-EDTA buffer, blocking was done with 3% hydrogen peroxide. Primary antibody was added followed by target binder and Horseradish peroxidase. The slides are then treated with Diaminobenzidine (DAB) chromogen, stained with haematoxylin and mounted with DPX (Dibutylphthalate Polystyrene Xylene). A positive nuclear stain for the four MMR proteins denote expression of wild type MMR proteins, hence MSI low status. A loss of nuclear expression denotes mutation of MMR proteins and hence MSI high (MSI-H) status. The percentage of PD-L1 positive tumour cells out of total tumour cells was estimated. Membranous staining of >1% of the tumour cells was considered as positive. CPS was calculated by dividing the total number of PD-L1 positive cells by the total number of viable tumour cells. Score >1% was considered to be positive (7).

Statistical Analysis

The PD-L1 expression on tumour cells was compared with the MSI status of different cancer types. Statistical analysis was done using IBM SPSS software version 21.0. The p-value was calculated using t-test and p <0.5 was considered statistically significant.

Results

A total of 151 cases were included in the present study, out of which 87 (57.6%) were males and 64 (42.4%) were females. The mean age was 53.5 years (26-81 years). MSI-H was observed in 18 cases (11.92% of cases). The mean age was 66.5 years (42-91 years), out of which 11 were males and seven were females. The MSI was high in nine out of 48 cases (18.75%) of colon cancer (Table/Fig 1),(Table/Fig 2), three out of 15 cases (20%) of endometrial cancers, three out of 20 cases (15%) of gastric cancers (Table/Fig 3),(Table/Fig 4).

7Rest includes one out of sixteen cases of ovarian cancer, one out of two malignant melanomas and one out of three glioblastoma. The MSI was low in 133 cases (Table/Fig 5),(Table/Fig 6),(Table/Fig 7),(Table/Fig 8). PD-L1 was positive in seven of the 18 MSI-H cases (38.88% of MSI-H cases), which included five cases of colon cancer, one gastric cancer and one endometrial cancer. The p-value of significance was 0.03 (statistically significant).

Among the 18 MSI-H cases, 12 were moderately differentiated adenocarcinoma, four were poorly differentiated adenocarcinoma, one malignant melanoma and one glioblastoma. PD-L1 positivity was seen in 11 cases- one squamous cell carcinoma (lung), one neuroendocrine tumour (gastric), five poorly differentiated adenocarcinoma (one gastric, two colon, one lung and one endometrium), one Grade-2 infiltrating ductal carcinoma breast (Table/Fig 9) and three were moderately differentiated adenocarcinoma (colon).

The MSI was low/stable in the remaining 133 cases. The mean age was 62 years (26-98 years). Among these, 79 were males and 54 were females. PD-L1 was positive in 4 cases (3% of MSI low/stable cases), including two cases of lung cancer, one case each of gastric and breast cancer. The distribution of MSI status and PD-L1 among different cancers is summarised in (Table/Fig 10). The tumour cell score of PD-L1 was >10% in five out of seven cases of MSI-H. Rest of the cases had tumour cell score of <10%. The CPS was >10% in two cases of MSI-H. The CPS was positive in one MSI low/stable case and it was <10%.

Discussion

Microsatellites are short tandem repeats composed of repeating DNA sequences and found throughout the genome, in both the coding and non coding regions. They are highly polymorphic in different individuals but are of the same length in germline DNA and somatic DNA of tumour in the patient. Microsatellites are highly susceptible to DNA mismatch errors during the DNA replication process or any iatrogenic damage (8). These errors are usually rectified by the DNA MMR mechanism. The four critical genes involved in this process are MLH1, MSH2, MSH6 and PMS2. MSI is a defect in these DNA MMR genes which results in genetic hypermutability. dMMR occurs when any of these genes are inactivated either by germline and/or somatic mutations or epigenetic silencing (9). Mutations resulting in defective MSH6 can be compensated in the heterodimer by MSH3, and PMS2 by PMS1 or MLH3. MLH1 and MSH2 gene mutations cause proteolytic degradation of PMS2 and MSH6, whereas PMS2 or MSH6 mutations may not result in proteolytic degradation MLH1 or MSH2. A tumour with large number of clustered mutations in microsatellites and harbouring MSI is called a dMMR tumour. Hence, MSI is a marker of dMMR. IHC or PCR tests can be used to recognise dMMR/MSI in sporadic cancers which belong to the CMMRD spectrum like gastric, colorectal, small intestinal, endometrial, urothelial, central nervous system and sebaceous gland tumours (10),(11),(12). There is no sufficient data available for cancers which are not part of this spectrum.

The IHC uses antibodies against the four MMR proteins: MLH1, MSH2, MSH6 and PMS2 for predicting the MSI status of patients. The MMR proteins are expressed in the cell nuclei. As explained earlier, MLH1 mutations result in IHC loss of MLH1 as well as PMS2, while MSH2 mutation shows IHC loss of MSH2 and also MSH6. Therefore, PMS2, IHC can identify cases harbouring MLH1 or PMS2 defects, while MSH6 can identify cases with MSH2 or MSH6 defects. But, standalone MLH1 and MSH2 IHC cannot recognise cases with PMS2 or MSH6 defects (13),(14). Preanalytical errors such as inadequate tissue fixation may result in false negative staining or aberrant patterns like cytoplasmic, dot-like or perinuclear staining. Hence, it is mandatory to include an internal positive control like normal mucosa (15). Positive immunostaining in the presence of MMR deficiency can occur due to catalytically inactive but antigenic intact missense mutant MMR proteins or lack of PMS2 or MSH6 substituted by MLH3/PMS1 or MSH3 respectively. Therefore, it is recommended to use all the four IHC antibodies and whenever there is a doubt in the IHC, MSI-PCR should be done for confirmation (16). Indeterminate IHC results such as loss of only one heterodimer subunit also warrants confirmation by MSI-PCR. MSI is loss of stability in ≥2 of the microsatellite markers. MSI can also be assessed by Next Generation Sequencing (NGS) (17). The host antitumour immune function is negatively regulated by immune checkpoints which are critical for the suppression of the host antitumour immune reactivity. These immune checkpoints are the targets of tumour’s ability to escape immunosurveillance. This forms the basis for the targeted treatment of human cancers using immune checkpoint inhibitors. T and B lymphocytes, Natural Killer (NK) cells and Tumour-Infiltrating Lymphocytes (TILs) express an inhibitory co-receptor PD-1 (18). In the Tumour Microenvironment (TIME), PD-1 binds with the PD-L1 and inactivates the TILs resulting in immune resistance of the tumour (19). The PD-1/PD-L1 pathway is a major negative modulator of immune response. Inhibition of this pathway by administration of monoclonal antibodies (mAbs) reactivating the Cytotoxic T Lymphocytes (CTLs) can be used to treat human cancers such as melanoma (20), renal cell carcinoma (21), and Non Small Cell Lung Cancer (NSCLC) (22).

Expression of PD-L1 by tumour cells is being used as a biomarker for targeted immunotherapy, while dMMR helps in predicting the response of tumours to PD-1 blockade (4),(23). There are reports of various clinical studies showing relatively more “sensitive” response of dMMR/MSI-H colorectal carcinoma patients to anti-PD-1/PD-L1 mAbs therapy when compared to proficient MMR (pMMR)/MSI-L cases (24),(25). The immune cell positivity for PD-L1 staining was found to be significantly higher in dMMR tumours than pMMR tumours (26). The expression of PD-L1 on tumour cells has been found to be independent of MSI and EBV in cases of gastric carcinoma with lymphoid stroma (27). There is significant correlation between high TMB and PD-L1 status in melanoma (28); MSI cases are very rare in NSCLC, but the percentage of PD-L1 positive cases is very high, although it has been demonstrated that PD-L1 and TMB-high are independent in such cases. MSI-H/dMMR has been found in 1.16% lung cancer patients, most of which are Squamous Cell Carcinoma (SCC) (29). MSI-H/dMMR is of limited prognostic value in triple negative breast cancer as their incidence is extremely low (30). Studies are available which show higher PD-L1 expression in aggressive thyroid cancers suggesting that targeted therapy can help these cases (31). Clinicians can predict response to anti PD-L1 therapy in their patients by evaluating the MSI and PD-L1 expression in the tumour and thereby select suitable patients for this therapy. This helps in identifying candidates who can benefit from anti-PD-1/PD-L1 therapy (32).

Limitation(s)

This study shows a significant association of MSI-H with PD-L1 expression in tumours. The limitations of the present study are the limited number of MSI-high cases available for comparison with PD-L1 and lack of confirmation of MSI status with PCR or NGS studies. Larger scale studies will help to establish a more definitive correlation between MSI status and PD-L1 expression in a variety of tumours.

Conclusion

This study shows a significant association of MSI-H with PD-L1 expression in tumours. This can provide valuable information to identify patients with MSI-high status and PD-L1 expression, who may be potential candidates for targeted therapy. However, more extensive and large scale studies are required to standardise and recommend definitive cut-off values for PD-1/PD-L1 expression and also to study the response of tumours harbouring MSI to immune checkpoint blockade therapy. This can be a big breakthrough for cancer treatment worldwide.

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DOI and Others

DOI: 10.7860/JCDR/2023/63627.18029

Date of Submission: Feb 24, 2023
Date of Peer Review: Mar 29, 2023
Date of Acceptance: May 08, 2023
Date of Publishing: Jun 01, 2023

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? No
• For any images presented appropriate consent has been obtained from the subjects. No

PLAGIARISM CHECKING METHODS:
• Plagiarism X-checker: Mar 01, 2023
• Manual Googling: May 01, 2023
• iThenticate Software: May 04, 2023 (17%)

ETYMOLOGY: Author Origin

EMENDATIONS: 8

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