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

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Dr Mohan Z Mani

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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."



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Sri Devaraj Urs Medical College
Sri Devaraj Urs Academy of Higher Education and Research , Kolar, Karnataka
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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.
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Professor and Head
Department of Pediatric Dentistry
Saraswati Dental College
Lucknow
On Sep 2018




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




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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 : 2022 | Month : May | Volume : 16 | Issue : 5 | Page : EC15 - EC18 Full Version

Immunohistochemical Characterisation of Cell of Origin in Diffuse Large B-cell Lymphoma and its Association with the Double Expressor Phenotype: A Retrospective Study


Published: May 1, 2022 | DOI: https://doi.org/10.7860/JCDR/2022/52051.16311
Prachi, Hemamalini Aiyer, Gaurav Sharma, Suparno Chakrabarti, Sarita Rani Jaiswal

1. Senior Resident, Department of Pathology, Dharamshila Narayana Superspeciality Hospital, New Delhi, India. 2. Head and Senior Consultant, Department of Pathology, Dharamshila Narayana Superspeciality Hospital, New Delhi, India. 3. Senior Consultant, Department of Pathology, Dharamshila Narayana Superspeciality Hospital, New Delhi, India. 4. Head and Senior Consultant, Department of Haemato-oncology and BMT, Dharamshila Narayana Superspeciality Hospital, New Delhi, India. 5. Senior Consultant, Dharamshila Narayana Superspeciality Hospital, New Delhi, India.

Correspondence Address :
Dr. Prachi,
Resident, Department of Pathology, Dharamshila Narayana Superspeciality Hospital, New Delhi, India.
E-mail: prachipath123@gmail.com

Abstract

Introduction: Diffuse Large B-cell Lymphoma (DLBCL) is the most common type of Non Hodgkin Lymphoma (NHL) and is categorised into the Germinal Centre B-cell (GCB) and Activated B-cell (ABC/Non GCB) subtypes as per the Cell Of Origin (COO) model with the help of gene expression profiling/immunohistochemistry. The non GCB subtype has been found to associate with the Double Expressor (DE) phenotype (i.e., co-expression of BCL-2 and c-myc by IHC and this association was substantiated and proven to be statistically significant in the present study.

Aim: To categorise all DLBCL cases into GCB and Non GCB and further into DE and Non-DE by using Hans and Choi IHC alogrithm.

Materials and Methods: A retrospective study of 50 patients was carried out with the help of archival material filed in the Department of Anatomic Pathology at Dharamshila Narayana Superspeciality Hospital, New Delhi, India from 1st January 2019 to 30th June 2020. The study was approved by Instituitional Ethics Committee (IEC). Chi-square test and Yates correction were used for statistical analysis.

Results: The study cohort was divided into two Groups- group-A with nodal presentation and group-B with extranodal presentation. By using the Hans and Choi IHC algorithms, the cases were categorised into the GCB and non GCB subtypes in both the groups. The DE phenotype was determined in each case by the co-expression of c-myc (>40%) and BCL-2 (>50%) by IHC. By using the statistical chi-square test analysis and Yates correction, the association of DE was found to be statistically significant with non GCB type lymphomas and non DE with GCB lymphomas with p-value being 0.0005 and 0.00168, respectively.

Conclusion: Due to the heterogeneity inherent in DLBCL, prediction of the DE phenotype and the COO by IHC is a sensitive tool and helps in the prognostication and therapeutic triage of patients. Hence, in all the cases diagnosed as DLBCL, a detailed morphological and IHC work up is mandatory to determine prognosis and possibly tailor therapy according to COO and DE phenotype.

Keywords

B-cell lymphoma, Cell of origin model, Double expressor, Hans and choi algorithm, Immunohistochemistry

The DLBCL is the most common type of aggressive NHL, representing approximately 24% of all newly diagnosed cases of NHL. It represents a heterogenous group of diseases having variable outcomes that are differentially characterised by clinical features and most recently by recurring mutations (1),(2). Clinically, patients present with rapidly enlarging lymphadenopathy, along with constitutional symptoms. There is a high frequency of extranodal disease, if there is generalised lymphadenopathy (1),(2).

The “COO” model divides DLBCL into the GCB type and non GCB type or ABC type depending upon gene expression pattern. The gene expression of the germinal centre type fits with the normal germinal centre derived B cell and in the subtype non GCB DLBCL or ABC lymphoma, the gene expression profiling more closely fits in with a normal ABC (1),(2). COO model also determines the prognosis of the two biological subtypes of DLBCL that should be treated differently. C-myc is a proto-oncogene on chromosome 8q24 and encodes a transcription factor, which leads to cellular survival and proliferation, if dysregulated. BCL-2 is an oncogene with an anti-apoptotic property (1),(2).

Myc rearrangement t(8;14) is prototypically associated with Burkitt lymphoma but is also associated with 12-15% of DLBCL. BCL-2 rearrangement t(14;18) is also important as it leads to a drug resistant phenotype with increase in cancer cell survival. Thus a concurrent rearrangement of BCL-2 and c-myc, which are present in approximately 5-7% of DLBCL, leads to a clinically resistant form of DLBCL, termed as a double hit lymphoma and is associated with a poor prognosis (3). The revised WHO classification recognises the co-expression of myc (>40%) and BCL-2 (>50%) proteins as a new adverse prognostic marker within DLBCL, Not Otherwise Specified (NOS) as “DE lymphoma” (4),(5),(6).

Another variant of the double hit lymphoma presents with co-rearrangement of c-myc and BCL-6 genes. All the three genes BCL-2, myc and BCL-6 are simultaneously rearranged in the phenotype termed as “Triple-Hit Lymphoma (THL)” (6). Patient who have the double hit rearrangement usually have protein over-expression and therefore have the DE phenotype. However, the converse is not always true (7). Double hit lymphoma occurs more commonly in GCB type of DLBCL, while DE lymphoma is commoner in the non GCB type of DLBCL. DEs are detected by IHC staining and double hit lymphomas are detected by Fluorescence In Situ Hybridization (FISH) (7).

The most common upfront treatment for DE lymphomas is Chemoimmunotherapy (CI) with R-CHOP (Rituximab, Cyclophosphamide, Doxorubicin, Vincristine and Prednisolone) which leads to a cure in 50-60% of patients. Patients, who develop disease that is refractory to upfront treatment or relapse after achieving remission, are treated with Autologous Stem Cell Transplantation (ASCT) and they generally have a poor outcome.

The majority of patients with DE lymphoma relapse after R-CHOP (8), hence such patients should be treated with ASCT. For patients unable to undergo transplantation, the median survival is six months. Chimeric Antigen Receptor T-cell Therapy (CART) is promising for patients with aggressive BCL that do not respond to other treatment modalities.

The main aim and objective of present study was that all high grade DLBL should be categorised into their molecular subtypes i.e., GCB lymphomas and non GCB lymphomas and further into DE and non DE phenotypes by using Hans and Choi Alogrithm (IHC Alogrithm) [9,10], as it aids in risk stratification, prognostication and for appropriate treatment, as it varies for each subtype. Therefore, all patients of DLBCL should be assessed for the DE phenotype, which aids in risk stratification of the patient and further helps in optimising the treatment strategy.

Material and Methods

This was a retrospective study conducted at Dharamshila Narayana Superspeciality Hospital, New Delhi, India from 1st January 2019 to 30th June 2020, to validate a model to predict the DE phenotype based on COO subtype. The study was approved by Institutional Ethics Committee (IEC), (name of committee-Dharamshila Narayana Superspeciality Hospital) vide registration no. ECR/226/Inst/DL/2019. Written consent was obtained.

Inclusion criteria: The study includes all high grade DLBCL cases.

Exclusion criteria: Excludes other high grade non Hodgkin lymphomas.

All newly diagnosed DLBCL patients were divided into group A including patients with nodal involvement and group B including patients with extranodal involvement. Based on “COO” both the groups were further divided into GCB type and non GCB type of DLBCL. With the help of IHC using the Hans or Choi algorithm IHC staining for BCL-2 and c-myc was used to determine the DE phenotype and its association with the COO subtypes of DLBCL. Present study included 50 patients, who presented with generalised lymphadenopathy and/or extranodal disease along with other associated constitutional symptoms. A biopsy was performed in each case and sent in 10% neutral buffered formalin to the Department of Surgical Pathology for further processing and evaluation. Formalin fixed paraffin embedded tissue sections were cut and mounted on slides and stained with routine Haematoxylin and Eosin (H&E) stain.

Further, IHC stains were applied to assess the ‘COO’ and DE phenotypes based on the percentage of staining of the cells and were analysed. In all cases IHC was performed using the automated Roche Ventana Benchmark XT IHC Autostainer (Ventana Medical Systems Inc. Tucson, Arizona) with the aid of the UltraView Universal DAB Detection Kit. The antibodies and clones employed were- CD3 (PS-1), CD5 (4C-7), CD10 (56C6), CD20 (L-26), CD30 (BerH2), BCL-2 (EP-36), BCL-6 (EP278), C-myc (EP121), MUM-1 (EP190), Ki-67 (30-9) and FOXP-1 (EP137) provided by Pathnsitu Biotechnologies Pvt Ltd and GCET-1(ab68880 from ABCAM (Cambridge, UK). Co expression of c-myc (>/=40%) and BCL-2 (>50%) in neoplastic cells by IHC staining was considered as DE phenotype.

Statistical Analysis

The statistical analysis used in present study was Chi-square test and Yates Correction to calculate the p-value and also to determine the significance of different parameters.

Results

The median age at presentation was 49 years (20-81 years) with a male to female ratio of 2.3:1 (35 males and 15 females). Group A with nodal disease included 28 patients and group B with extranodal disease included 22 patients, constituting 56% and 44%, respectively of the study cohort. Histological sections examined from biopsy specimens showed diffuse infiltration by predominantly large atypical lymphoid cells with prominent nucleoli and mitotic activity. All cases were positive for the pan B-cell marker CD20 and a few showed positivity for CD30. Eight of the 28 patients in Group A were categorised as GCB type and 20 (71.4%) were categorised as non GCB type by immunohistochemistry, using the Hans and Choi algorithm. Out of the eight GCB type, only 1 (12.5%) had a DE phenotype, while 7 (87.5%) were non DE type. Out of the 20 non GCB type, 15 (75%) had a DE phenotype and 5 (25%) were of the non DE type. The features of immunostaining can be appreciated in (Table/Fig 1),(Table/Fig 2),(Table/Fig 3),(Table/Fig 4),(Table/Fig 5),(Table/Fig 6),(Table/Fig 7),(Table/Fig 8),(Table/Fig 9),(Table/Fig 10).

Four of the 22 patients in Group B (18.2%) were categorised as GCB type and 18 (81.8%) as non GCB type. Out of the four GCB type, none showed a DE phenotype. However, 10 of the 18 non GCB type (55.6%) were DE type and 8 (44.4%) were non DE type. The DE phenotype was present in 26/50 (52%) and the NDE phenotype was expressed in 24/50 (48%) of cases included in the present study. While in the non GCB type DE phenotype was expressed in 25 cases (65.8%) out of 38 cases, only one of the 12 GCB cases expressed the DE phenotype (8.3) (Table/Fig 11),(Table/Fig 12),(Table/Fig 13).

Thus, in the nodal group the non GCB subtype analysed with the DE phenotype and the association was found to be statistically significant (p-0.009). In the extranodal group, though the non GCB subtype showed the DE phenotype more commonly than the NDE phenotype, the association was not found to be statistically significant (p-0.1434). Overall the DE phenotype analysed with the non GCB subtype and the NDE phenotype with the GCB subtype. The association was found to be statistically significant.

Thus, the DE phenotype associated with the non GCB subtype and the NDE phenotype with the GCB subtype in this study. The study has used the IHC algorithm (Hans and Choi) to do the molecular subtyping of all lymphoma cases. The co-expression of c-myc (>40%) and BCL-2 (>50%) proteins in DLBCL, NOS cases or any of the above one with BCL-6 (>30%) is considered as “DE lymphoma” (Table/Fig 14),(Table/Fig 15) (9),(10).

Discussion

According to the 2016, WHO classification, diagnosis of all cases of DLBCL, NOS should include COO, (GCB vs. ABC or non GCB, if an IHC algorithm is used), because of their different molecular features, biologic behaviour, prognosis and treatment (3). This distinction is important because better outcomes are seen in patients with the germinal-center subtype than with the non germinal-center subtype.

We therefore now have a “COO” model showing that there are at least two genetic biologic types of DLBCL i.e., GCB and non GCB subtype that explains the prognosis and is targeted differently. This insight has led to recent trials that test new treatments in one genetic subtype vs the other. We are hopefully very close to understanding whether these two subtypes of DLBCL by using two different IHC algorithm (Hans CP et al., and Choi WWL et al.,) (9),(10) and should be treated differently. The most challenging profile to detect in routine practice is COO. The gold standard for identifying germinal-centre DLBCL versus non germinal-centre DLBCL was first defined as gene-expression profiling patterns in frozen tumour material. However, gene-expression profiling is not routinely available nor is it considered a standard test. The most common approach is IHC using different algorithms, such as the Hans, Choi or Tally algorithms, to determine whether a lymphoma is germinal-centre DLBCL or non germinal-centre DLBCL. As compared to gene expression profiling in frozen material, there is an error rate of approximately 20% with immunohistochemical algorithms shown below (11). The present study has used the Hans/Choi Algorithms to determine the COO.

The presence of both the myc and BCL-2 rearrangements defines Double-Hit Lymphoma (DHL). This phenotype is very proliferative and drug-resistant, and it is associated with a poor prognosis. Another variant of DHL is co-rearrangement of myc and the BCL-6 gene. Rarely, all three genes BCL-2, Bcl-6 and C-myc, patients who have the double-hit rearrangement usually have protein overexpression, and therefore have the DE phenotype. However, the converse is not always true: dual-expressor protein overexpression is not always associated with an underlying double-hit re-arrangement. Complicating the picture is that most DHLs occur in the setting of a germinal-centre DLBCL, whereas most DE lymphomas occur in non-germinal-centre DLBCL as demonstrated in this study which shows a strong association between the DE phenotype and the Non GCB COO subtype by IHC (12),(13). myc and BCL-6 are simultaneously rearranged in a phenotype termed THL. Both double-hit and THL s have a poor prognosis with standard treatment (14). The starting point in a discussion about prognostic variables in DLBL is the observation that some patients can be cured and others cannot. Several studies have investigated the biologic underpinnings for these different outcomes. A pioneering genetic evaluation of DLBCL, published by Alizadeh AA et al., showed two subtypes: germinal center and non germinal center (also known as ABC) (15).

IHC staining to identify protein expression of myc also showed that there are lymphomas in which myc and BCL-2 genes are overexpressed at a protein level, without the genetic rearrangements. This profile has been referred to as the “DE” phenotype in DLBCL in the revised 2016 World Health Organisation (WHO) classification of lymphoid neoplasms. The WHO classification defines overexpression
as greater than 40% c-myc-expressing cells and greater than 50% BCL-2-expressing cells by IHC (3). As shown in a study by Hu S et al., patients with DE DLBCL have worse outcomes than patients in whom these proteins are not overexpressed; in general, only onethird of patients have long-term disease control with R-CHOP (16).

DHL is relatively uncommon, occurring in approximately 5-7% of patients with DLBCL. However, DE lymphomas may be present in as many as one-third of patients with DLBCL and serve to identify a significant subset of cases with a worse prognosis. The DHL’s can be detected with FISH or standard cytogenetic analysis. The DE lymphomas are diagnosed by IHC. The DE phenotype was not given a unique category, but was recognised by the WHO as a poor prognostic sign within DLBCL (3).

When patients with aggressive B-cell lymphomas relapse or become refractory to therapy, standard options are limited. For patients unable to undergo transplant, or for those who relapse after a stem cell transplant, the median survival is approximately six months. Despite the many trials that have tried to improve upon this dismal statistic, there are no breakthroughs at this time. Chimeric Antigen Receptor T-cell therapy (CART) is exciting. This treatment is still in the early phases of research and associated with toxicity, but it is promising for patients with aggressive B-cell lymphomas that do not respond to other therapies. There are also a number of new biologic and targeted agents that are promising, and finding which patients may respond to a particular treatment is a high priority (17).

Limitation(s)

The limitation of present study was that, there was an approximately 20% error rate with IHC algorithm. The most sensitive is always gene expression profiling/m-RNA based technique, which is not usually available in a limited setting facility. Hence, Immunohistochemistry is routinely done to categorise all the high grade B-cell lymphomas into their molecular subtypes as an alternative solution.

Conclusion

The distinction of GCB versus ABC-DLBCL has led to differences in primary treatment by emerging new-targeted therapy. The current standard of care for most patients is R-CHOP, which has improved dramatically the outcome of DLBCL. However, for patients who fails to respond to R-CHOP, the choice of therapy is very likely to be influenced by the COO and the molecular pathways used by the tumours for survival and proliferation.

Although there are no strict recommendations on how to select cases for FISH analysis, a reasonable approach is to perform FISH analysis for myc, BCL-2 and/or BCL-6 in cases with aggressive clinical presentation, GCB phenotype, and double expression of myc and BCL-2 which has been shown to associate with the non GCB subtype as demonstrated by this study, thereby mandating a comprehensive and detailed morphologic and IHC work-up in all cases of high GBL in order to determine prognostic and therapeutically relevant and biologically distinct phenotypes.

Acknowledgement

The authors thank the entire technical staff, Suresh Chand Sharma, Barfi Ram Chaudhary and Nidhi Chaudhary from the Department of Pathology and entire technical team of Department of Haematooncology and Bone marrow transplantation unit, Dharamshila Narayana Superspeciality Hospital, New Delhi, India.

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DOI and Others

DOI: 10.7860/JCDR/2022/52051.16311

Date of Submission: Aug 21, 2021
Date of Peer Review: Dec 26, 2021
Date of Acceptance: Jan 21, 2022
Date of Publishing: May 01, 2022

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 24, 2021
• Manual Googling: Jan 14, 2022
• iThenticate Software: Jan 17, 2022 (7%)

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