EHA-TSH Haematology Tutorial on Lymphoma Session 1: The History of Lymphoma Classification and 2016 WHO revision ‘Learn from the past, look to the future, but live in the present’: Classification of Lymphoid neoplasms, WHO 2016 Update

Speaker:Ayşegül Üner Hacettepe University School of Medicine ‒ No disclosures Learning objectives

‒ Learn about the history of lymphoma classification ‒ To learn what is new in the WHO 2016 classification of lymphoid neoplasms Why Classify?

‒ Classification is essential for medicine – diseases can be described, defined and named. ‒ Disease entities should be distinct with non- overlapping features ‒ Consensus on definitions and terminology is necessary so that pathogenetic mechanisms behind diseases are better understood, clinical trials could be carried out and comparisons are feasible ‒ Any proposed classification should be biologically rational ‒ It should be clinically useful

Although pathologists took the primary responsibility for developing the classification of Hematological Malignancies, the advise of clinicians was obtained to ensure its usefulness and acceptance in practice Lymphoid neoplasms

Hodgkin’s Lymphoma NHL

NLPHL B-cell neoplasms T/NK-cell neoplasms

Classical HL NSHL Precursor B- Mature B- Precursor T- Mature T- MCHL cell cell cell cell Lymphocyte-rich neoplasms neoplasms neoplasms neoplasms Lymphocyte depleted Thomas Hodgkin was the first lecturer in Morbid anatomy and curator of its museum at Guy’s Hospital

S.A.Geller & C.R. Taylor, Virchows Arch (2013)

Lukes & Collins Gall & Mallory WF BNLI 1832 1966 1978 Classification of malignant L&C Included presumed NCI-lymphomas divided lymphomas cell of origin (B, T and into low, intermediate histiocytic) and and high grade groups morphology BNLI-Bennett et al

Thomas Hodgkin Rappaport Lennert

Report of 7 lymphoma 1940 Armed Forces Institute of 1974 Kiel Classification 1982 cases Pathology fascicle Concept of tumor grading Samuel Wilks used the published-based on introduced, low grade ‘- term ‘Hodgkin’s Disease in morphology: architecture cytic’ and higher grade ‘- 1865 and cell size blastic’

Macroscopic Microscopic Cell of origin- features features immunology ‘Nowhere in pathology has a chaos of names so clouded clear concepts as in the subject lymphoid tumors’

R.A. Willis WHO Classification WHO Classification of Hematopoietic and of Hematopoietic and 1994 Lymphoid Tissue 2008 Lymphoid Tissue ? Based on REAL Revision of the 4th Classification edition of the classification

WHO Classification REAL Classification of Hematopoietic and Lymphoid Tissue ILSG brought together 2001 Revised classification 2016 experts from around the globe Defined ‘real’ disease entities based on clinical features, morphology, immunophenotype and genetic information

Clinical, morphologic, phenotypic and genetic features Development WGS of numerous WES in antibodies GEP hematopoetic 1975 used in 1985 2010 malignancies immunophenotyping

Monoclonal Antibody PCR and NGS Technology Sequencing 1980’ 2000’ 2010’ s s s

Immuno- Targeted- chemotherapy chemotherapy therapies WHO Classification 2016 • New genomic technologies, clinical and morphologic observations has provided new insights into the biology of lymphomas • Data from multicenter clinical trials WHO Classification- 2016 • Recommendations for handling patients with early lesions • New definite and provisional entities defined • Modifications and renaming of some former entities Nomenclature influenced by clinical behaviour • Refinement of diagnostic criteria Molecular changes Diagnostic, prognostic and therapeutic implications

Early lesions of malignant lymphomas

‒ Can we apply the multistep carcinogenesis model to lymphomas? ‒ Is there a benign lymphoma? Monoclonal B-cell Lymphocytosis

• Precursor to CLL • <5x109/L monoclonal B cells without LAP, organomegaly or extramedullary disease • CLL-type-75% of the cases • Atypical CLL-type • Non-CLL-type • Minority of the patients will progress to overt lymphoid malignancy WHO 2016 • Low-count MBL (<0.5x109/L) vs High- count MBL Low count MBL – no specific follow-up recommended as it has very limited potential for progression to CLL Patients with high- count MBL require periodic evaluation

Rawstron AC et al Cytometry Part B (Clinical Cytometry) 78B (Suppl. 1):S19, 2010 CLL • No disease defining mutations • Number of mutations have been identified-at low frequency • Some mutations are associated with poor prognosis TP53 NOTCH1 SF3B1 BIRC3 In Situ Follicular Neoplasia (WHO 2016) (Follicular Lymphoma in situ WHO 2008) • Incidental finding; Intrafollicular BCL-2 positive centrocytes and centroblasts in an otherwise normal lymph node • Must be differentiated from partial involvement by FL • Staging work-up is necessary to exclude systemic FL • Few (<5%) patients progress to disseminated FL- markers to predict progression are not well defined What is new in Follicular lymphoma- WHO 2016 • Intestinal follicular lymphoma • Duodenal-type • Testicular FL • Childhood • Cytologically grade 3a FL • Bcl-2 negative • Diffuse FL • Pediatric-type FL Diffuse (appearing) FL • Frequently presents as a localised inguinal mass • Grade 1-2/3 morphology • BCL2 gene rearrangement negative • 1p36 deletion (not specific can be also seen in conventional FL) Pediatric-type Follicular Lymphoma (WHO 2016)

• It became a definite entity in WHO 2016 (was a provisional entity in WHO 2008) • Similar lymphoma can be seen in adult age group • Median age 15-18 • M:F 10:1 Pediatric-type Follicular Lymphoma (WHO 2016) • Large expansile follicles • No-diffuse areas • “Blastoid” morphology • Usually Grade 3, exceptionally Grade 1- 2 • No BM involvement Pediatric-type Follicular Lymphoma (WHO 2016) ‒ Differential diagnosis from grade 3 FL is necessary ‒ BCL 2, BCL6 or MYC rearrangements are not seen although Bcl-2 protein expression can be seen ‒ Usually involves head/neck region ‒ Stage 1 disease ‒ Excision alone can be curative IRF4-associated Large B-cell Lymphoma • Rare -0.05% of LBCL • Head&neck region and GIS • Wide age range (4-79) median age of 12 • F:M (9:11) • Usually stage I-II (84%) • Excellent prognosis (5 year survival 100%) • Nodular-diffuse growth of medium-large cells • BCL-6 and MUM-1 expression is a clue for the diagnosis Salaveria et al, Blood, 2011 Mantle Cell Lymphoma

• In-situ mantle cell neoplasia • Should be differentiated from mantle zone pattern of MCL • Leukemic non-nodal MCL • Indolent • IGHV-mutated SOX11- B cells Swerdlow S et al Blood 127;2381, 2016 Molecular alterations are included in the diagnostic algorithm Lymphoplasmacytic lymphoma MYD88 L265P mutation • İdentified in 90% of LPL/ Waldenstrom makroglobulinemi- • IgM MGUS • Not present in plasma cell myeloma • Can be seen in some othe NHL – Some of the other low grade B cell lymphomas – DLBCL, non-GC (%30), leg-type (%50), cases involving immune- priviliged such as testis, CNS etc CXCR4 mutations • LPL (30%) & IgM MGUS (20%) • Not seen in IgG or IgA MGUS Molecular alterations are included in the Hairy cell leukemia diagnostic algorithm • BRAF V600E mutation • Non present in Hairy cell leukemia-variant • MAP2K1 (MEK1) mutation • In cases which do not carry BRAF mutation and those which use IGHV4- 34 • 50% of Hairy cell leukemia-variant Changes in Low grade B-cell Neoplasms- WHO 2016 CLL HCL Even if there is cytopenias BRAF V600E and MAP2K1 >5x109/L CLL cells necessary for diagnosis LPL Proliferation centers’ importance MYD88 L265P Clinically relevant mutations FCL identified Mutations better defined MBL In situ lesions-changed from Low-count/high-count lymphoma to neoplasia MCL Localized forms and diffuse forms defined Genetic profile better delineated İndolent types Pediatric-type FL In situ lesions-changed from Became a definite entity, broader lymphoma to neoplasia age Diffuse Large B cell Lymphoma

Rosenwald A, et al: NEJM 346:1937, 2002 COO CD10 (+)

GC Bcl-6 (+) (-)

MUM1 Non- (+) (-) GC Non- GC GC

Hans CP. Blood; 103:275-282, 2004 Scott et al. JCO 2015 COO-WHO 2016

• Identification of the COO subtype is required • pathogenesis of GC and ABC subtypes are different • Requiring different therapeutic approaches • IHC surrogates can be used (ease and expense) until gene-expression technology can be implemented in clinical practice ‘Double expressor’ DLBCL • 19-34% of cases • Neoplastic cells MYC >40% and BCL-2 >50% of • Prognostically relevant but DE is not considered a separate entity High Grade B cell Lymphomas WHO 2016 • Aggressive High Grade B cell cases which should not be classified as BL or DLBCL • Two subcategories HGBCL with MYC and BCL2 and /or BCL6 rearrangements (so-called DH and TH) HGBCL, NOS • No evidence of DH or TH by genetic analysis • Should appear blastoid or Burkitt-like DHL/THL BCL2/MYC DHL BCL6/MYC DHL Majority show GC- ABC phenotype more phenotype (CD10+) frequent Proliferation index Immunoblastic generally high (>80%) morphology However, in about 20% Frequent extranodal of the cases it is low involvement Bcl-2 expression can be seen FISH

BCL-2 BCL-6 MYC Burkitt-like lymphoma with 11q aberration (WHO 2016 provisional) ‒ Resembles Burkitt lymphoma morphologically and phenotypically ‒ 11q alteration instead of MYC rearrangement ‒ More complex karyotypes, higher degree of cytological pleomorphism Swerdlow SH et al Blood 127: 2375, 2016 EBV positive diffuse large B-cell lymphoma of the elderly-WHO 2008 • ‘EBV+ clonal B-cell lymphoid proliferation that occurs in patients >50 years and without any known immunodeficiency or prior lymphoma’ • Rare cases of similar lymphoma may occur in younger individuals • Well-defined disorders that may be EBV+ are excluded from this category Nakamura S, Jaffe E, Swerdlow S. EBV positive diffuse large B-cell of the elderly. In: S. Swerdlow et al., eds.WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. Lyon, France: IARC, 2008:243–244. EBV+ DLBCL, NOS WHO 2016 • EBV+ DLBCL is recognized in younger patients with a broader morphological spectrum and better survival than initially thought • ‘NOS’ is to differentiate this from more specific entities such as LG etc • EBV+ mucocutaneous ulcer has been segragated from EBV+ DLBCL as a provisional entity due to its self limited growth potential and response to conservative management EBV+ Mucocutaneous Ulcer

Volume 34, 2010 ﰒ Am J Surg Pathol EBV+ mucocutaneous ulcer • ‘EBV+ circumscribed, ulcerative lesions involving oropharyngeal mucosa, skin, and gastrointestinal tract associated with various types of IS’ • Self-limited, indolent course, generally responding well to conservative management. • Patients with age-related or iatrogenic immunosuppression

Dojcinov et al, Am J Surg Pathol, 2010 Changes in High grade B-cell Neoplasms- WHO 2016 DLBCL EBV+ mucocutaneous COO ulcer ‘Double-expressor’ İatrogenic IS or age related Genetic landscape better ımmune senecense delineated Burkitt Lymphoma EBV+ DLBCL, NOS TCF3 and ID3 mutations in Can be seen at any age 70% Should be differentiated Burkitt-like lymphoma from EBV-related specific with 11q aberration entitites Changes in High grade B-cell Neoplasms- WHO 2016 HGBCL with MYC and BCL2 and /or BCL6 rearrangements (so- called DH and TH) HGBCL, NOS

Mature T/NK cell lymphomas

Account for 10-15% of lymphomas Diagnosis not easy!!! Morphologic and phenotypic variability Frequent extranodal presentation Neoplastic cells are frequently accompanied by reactive cell population Genetic tests are necessary for demonstration of clonality and neoplastic nature of the proliferation What is new in T- NK- cell neoplasms? • ALCL- a new definite and a provisional entity • Lymphomas derived from follicular TH cells better defined • New genetic information for PTCL,NOS • Better understanding of EBV-associated lymphoproliferative disorders • Name changes for some previously defined entities

CD 30+ mature T cell lymphomas Savage KJ et al, Blood 111, 2008 Anaplastic Large Cell Lymphoma

WHO 2008 WHO 2016 Anaplastic Large Cell ALCL, ALK+ Lymphoma ALCL, ALK- ALCL, ALK+ Breast implant ALCL, ALK- associated ALCL Anaplastic Large Cell Lymphoma, ALK- • Provisionel entity in WHO 2008 Classification, became a real entity in the WHO 2016 update • ‘CD30-positive mature T cell neoplasm which is morphologically similar to ALK-positive ALCL but with no ALK expression’ Savage KJ et al, Blood 111, 2008 Parilla Castellar E.R. et al, Blood, 2014 Breast implant associated ALCL

Thompson et al. Haematologica, 2010 Nodal T cell Lymphomas with TFH Phenotype TFH markers • Angioimmunoblastic T CD279/PD1, CD10, cell Lymphoma BCL6, CXCL13, ICOS, • Follicular T cell SAP and CXCR5. Lymphoma • Peripheral T cell Genetics Lymphoma, NOS with IDH2, TET2, DNMT3A,

TFH Phenotype CD28, RHOA t(5;9) ITK-SYK Follicular T-cell Lymphoma PTCL, NOS, Follicular variant

• Derived from TFH cells • Follicular/nodular growth pattern • Expansion of FDC, arborising blood vessels and polymorphic cellular background characteristic for AITL is not observed • RS-like cells (EBV+/-) can be present • Localised disease • ITK-SYK translocation t(5;9)(q33;q22) • Unique gene expression signatures were identified for major PTCL entities • 14% of PTCL, NOS cases were re-classified as AİTL • ALK(–) ALCL is a distinct entity • Tumor microenvironment plays an important role in prognosis of AITL

Javeed Iqbal et al. Blood 2014;123:2915-2923 PTCL, NOS molecular subgroups

Intestinal T-cell Lymphomas

Enteropathy- associated TCL Monomorphic epitheliotropic EATL, Type I (WHO 2008) intestinal TCL (MEITL) EATL, Type II (WHO 2008) • Associated with celiac disease • No association with celiac • Seen in individuals of northern disease European origin • İncreased in incidence in Asians and Hispanic population • Morphology: Polymorphic • Morphology: Monomorphic • Adjacent mucosa; epitheliotropism, villus • Phenotype: CD8, CD56 and atrophy, crypt hyperplasia MATK+, mostly derived form gd T-cells (STAT5B mutations) Indolent T-cell LPD of the GI Tract • Most common in small intestine and colon; less often in stomach and oral mucosa • Morphology • Low proliferative index • No destruction of glands • No cytologic atypia • Mostly CD8+ • Conservative management

Perry A et al Blood 2013 Gastrointestinal indolent T-cell lymphoproliferative disorder

Ganapathi KA et al Haematologica, 2014 Cutaneous T-cell Lymphomas • Primary cutaneous acral CD8+ TCL Derived from CD8+ cytotoxic T cells • Primary cutaneous gd TCL • Primary cutaneous CD4+ small/ medium T-cell LPD (provisional entity in the 2008 classification) TFH phenotype Recurrent mutations seen in nodal TFH lymphoma were not identified Indolent clinical behavior Conservative local management Mature T- and NK-cell neoplasms- New provisionel entities in WHO 2016

• Indolent T-cell lymphoproliferative disorder (LPD) of the GI tract • Primary cutaneous acral CD8+ TCL • Follicular T-cell lymphoma

• Peripheral T cell lymphoma with TFH phenotype • Breast implant–associated ALCL Name changes

WHO 2016 WHO 2008 • Systemic EBV+ T-cell • Systemic EBV+ T-cell lymphoma of childhood lymphoproliferative disorder of childhood • Hydroa vacciniforme–like • Hydroa vacciniforme-like lymphoproliferative disorder lymphoma • Monomorphic epitheliotropic • EATCL type 2 intestinal T-cell lymphoma • Primary cutaneous CD4+ • Primary cutaneous CD4+ small/medium T-cell small/medium T-cell lymphoproliferative disorder lymphoma Summary

‒ Refinement of definitions, diagnostic criteria and terminology ‒ Early lymphoid lesions and those with indolent clinical behavior better delineated ‒ Impact of location, age of the patient and infectious agents ‒ Relevance of phenotypic and molecular information in subtyping of various entities ‒ Impact of NGS is reflected in classification Future

‒ Ongoing research is providing insights and also raising questions for future discovery ‒ Potential targets for new therapies will be better defined and implemented ‒ New Classifications /updates incorporating newly acquired information is at the horizon……..