Updating the 2008 WHO Classification of Small B cell lymphomas Elias Campo

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Updating the 2008 WHO Classification of Small B cell lymphomas Elias Campo Hospital Clinic, University of Barcelona Barcelona, Spain

Disclosure of Relevant Financial Relationships Dr. ELIAS CAMPO declares he has no conflict(s) of interest to disclose.

Chronic Lymphocytic Leukemia/ Small Lymphocytic Lymphoma Maintain current definitions (> 5 x10 9 /L monoclonal lymphocytes with the CLL phenotype) Extramedullary tissue involvement and cytopenias allow for lower number of atypical lymphocytes SLL is the same disease but restricted to tissues of non-leukemic (< 5 x10 9 /L) patients without cytopenias 20 15 10 5 0 B clonal (x10 9 /L) Lymphocytes (x10 9 /L) WHO 2008

Monoclonal B-cell Lymphocytosis Definitions and Subtypes Monoclonal B-cells 5000 x 10 9 /L Absence of other lymphoproliferative disorders or autoimmune disease CLL-like (70%) Atypical CLL* (15%) Non-CLL* (15%) * Cytogenetic studies are recommended Low-count MBL Low lymphocyte and B cell counts (usually <50/μl) Different IGHV/No stereotyped BCR Very low risk of progression (if any) No indication to monitor even if detected incidentally Clinical MBL High lymphocyte and B cell counts (500-5000/μl) Lymphocytosis High risk cytogenetic alterations (5-9%) Annual progression requiring treatment 1-2% Clinical monitoring Dagklis et al Blood 2009 Karube et al Sem Cancer Biol 2014

Non-CLL MBL (CD5- MBL / Clonal B-cell lymphocytosis) Bone marrow involvement (sinusoidal) / 80% lymphocytosis > 4 x 10 9 /L Phenotype/Genotype suggestive of splenic lymphoma CD49d+/CD38-, CD10-; 20% CD5+ weak 72% aberrant karyotypes, del(7q) 15%, transloc 7q 13%, i(17q) 17% V4-34 (23%) V1-2 (6%) (More similar to SDRP than SMZL) 17% Progression (most) with splenomegaly and other lymphomas (MZL) Xochelli A et al Blood 2014 MBL nodal counterpart Atypical cells with CLL phenotype in otherwise reactive LN Absence of proliferation centers; < 1.5 cm nodes Gibson SE et al Haematologica 2011

Histological Progression in CLL Clinical significance of proliferation centers Heterogeneous terminology: Accelerated CLL Proliferation center-rich-cll Histologically Aggressive CLL CLL with expanded proliferation centers Expanded / confluent proliferation centers - High proliferation (Ki67 >30%) - Del(11q) 25%, del(17p) 16%, t(14q) 31% Histological criteria not standardized Survival intermediate between CLL and Richter transformation P < 0.01 Ki67 Standard CLL Gine E et al Haematologica 2010; 95:1526-33 Ciccone M et al Leukemia 2012 26:499-508 Falchi L et al Blood 2014; 123; 2783-90 RS Expanded PC

New Markers in CLL LEF1 1 Transcitpion factor, wnt pathway Useful in the diagnosis of CLL in tissues: 100% expression in CLL but not in other small B-cell lymphomas Caveat: Expression in 40% DLBCL LEF1 CD200 2 - Immunoglobulin superfamily - Expressed in CLL, HCL, and FL - Negative in MCL but positive in 24% indolent SOX11-negative MCL 3 CD49d 4 Integrin family Prognostic value independent of CD38/ ZAP70 Predictive value for BCR-targeted therapies? 1 Tandomn B et al Mod Pathol. 2011, 24:1433 43 2 Challangundla L et al Am J Clin Pathol 2014; 142.837 44 3 Espinet B et al Clin Cancer Res 2013; 20:1007 19 4 Bullan P et al J Clin Oncol 2014; 32:897 904

Lymphomas with plasmacytic differentiation Role of MYD88 mutation >90% Waldenström macrogl/lpl >50% IgM MGUS TLR MYD88 IRAKs NF-kB pathway MAPK pathways Inflammatory cytokines and chemokines 70% LBCL CNS & Testis 29% DLBCL-ABC 7-9% MALT 0-10% SMZL* 0-24% nmzl* 3% CLL L265P, most common mutation, detectable in FFPE tissues Useful information in the differential diagnosis * Interpret with caution results in the literature Ngo VN et al Nature 2011; 470:115 9; Treon SP et al N Engl J Med 2012; 367:826 33; Landgren O et al Leukemia 2014; 28:1799 803; Hamadeh F et al Modern Pathol 2014 e pub

Lymphoplasmacytic Lymphoma MYD88 and CXCR4 Somatic mutations MYD88 90-100% WM/LPL CXCR4 25% WM; Resistance to Ibrutinib Broader spectrum of cytological and architectural patterns 1 Diffuse architecture Follicular colonization Monotonous lymphoid population, minor plasmacytic morphology Polymorphic composition most probably other entities 1 Recognition of anomalous phenotypes 2,3 CD10 + (3-16%), CD5+ (17% ) IgD+, CD21 IgM MGUS carries MYD88 mutations in 47-87% 4 Precursor of LPL/WM. Gamma heavy chain disease lacks MYD88 L265P and is not considered LPL 5 CD10 IgD 1 Hamadeh F et al Modern Pathol 2014 e pub; 2 Konoplev et al Am J Clin Pathol 2005; 124: 414 20; 3 Lin P et al Am J Clin Pathol 2011; 136: 195 210; 4 Landgren O et al Leukemia 2014; 28:1799 803; 5 Haematologica 2014;99(9):e154 5

Early steps in Follicular Lymphoma In Situ and early involvement lesions t(14;18) in peripheral blood Detection require sensitive methods Most do not progress but some evidences of FL precursor in situ follicular neoplasia Incidental finding Low incidence of progression (<5%) Need to exclude systemic lymphoma Jegalian AG et al Blood 2011 Partial involvement by FL 50% progress to overt FL Adam P et al AJSP 2005 Mamessier E et al Haematologica 2014; 99: 802 810

Follicular lymphoma grading Maintain current grading system 1 2, 3a and 3b Some cases do not fit well in classical definitions Low morphological grade with high proliferation 1 FL «pediatric type» 2 FL with blastoid morphology 3 FL grade 3B Relation to GC type DLBCL Low frequency of t(14;18) High IRF4 expression in some cases (3A/B, elderly, nodal, CD10 negative, high Ki67) 4 IRF4 1 Wang SA et al Am J Surg Pathol. 2005;29:1490 6; 2 LiuQ et al Am J Surg Pathol. 2013;37:333 43; 3 Natkunam Y et al Am J Surg Pathol. 2000;24:525 34; 4 Karube K et al Blood. 2007;109:3076 9

Different subtypes of t(14;18) negative FL FL with conventional morphology 1 30% BCL2 protein positive Lower GC expression signature (CD10 negative) Higher proliferation No clinical impact Diffuse variant of FL 2 Large nodal tumors in inguinal region Localized disease CD10, BCL2, BCL6, CD23 positive Del 1p36 Primary extranodal FL (e.g. cutaneous) 1 Leich et al. Blood 2009;114:826-34 2 Katzeberger T et al Blood 2009;113:1053-61

Pediatric lymphomas (come of age) Follicular Lymphoma Pediatric Type Children and young adults Striking male predominance Nodal presentation, head and neck Expansile GC with grade 3, blastic No diffuse areas High proliferation rate CD21 Lack of t(14;18) Excellent prognosis Watch & wait therapy recommended BCL2 Liu Q et al Am J Surg Pathol. 2013;37:333 43 Louissaint A Jr et al Blood. 2012, 120:2395 404

Large B-cell lymphoma with IRF4 rearrangement New provisional entity segregated from other pediatric FL Waldeyer s ring, head and neck nodal, bowel presentation Most commonly in children/young adults Follicular and diffuse areas with grade 3 Germinal center phenotype (CD10/BCL6) BCL2 expression but no t(14;18) IRF4 Strong IRF4 expression and IRF4 translocation Cases without the genetic alteration may not be detected Treatment is often required IRF4 BAP Caution: name subject to change, precise criteria yet to be determined since IRF4 + not specific Salaverria I et al Blood 2011;118:139 47; Liu Q et al Am J Surg Pathol. 2013;37:333 43

Mantle cell lymphoma CCND1-negative variant CCND2 trans 55% Classic MCL Cyclin D1 CCND1 neg MCL Cyclin D1 Sox11 Mozos et al Haematologica 2009 Salaverria et al Blood 2013

Mantle cell lymphoma Indolent Variants Clinical concept with different pathological conditions In situ MC neoplasia, Mantle zone pattern, low proliferation index (SOX11+ or SO11-) In situ MC neoplasia Mantle Zone MCL Low proliferation Non-nodal leukemic subtype of MCL Non nodal leukemic (splenomegaly) disease SOX11 negative Hypermutated IGHV Simple karyotypes May transform into blastoid MCL (TP53 mut) Richard et al J Clin Pathol 2006; Jares P et al J Clin Invest 2013

NGS in small B cell lymphomas Complex landscape of somatic mutations in different tumors with two major patterns One mutated gene in most cases of the disease Great heterogeneity with few frequently mutated genes and higher number of genes with low recurrence rate Mutated genes cluster in functional pathways Common mutated genes and pathways across entities

Diagnostic value of somatic mutations in mature samll B-cell lymphoid neoplasms 90% WM Waldenstrom M MYD88 L265P 29% DLBCL-ABC 6% MZL 3% CLL Hairy Cell Leukemia BRAF V600E 79-100% HCL 4% Plasma cell myeloma 3% NHL (Other BRAF mut) HCL-v HCLc IGHV4-34 MAP2K1 50% HCLv 50% HCLc IGHV4-34 0% HCL BRAFmut Tiacci et al NEJM 2011 Ngo Nature 2011; Puente Nature 2011; Xi L et al Blood 2012; Xu et al ASH 2011; Hunter Z et al ASH 2011;Waterfall Nat Genet 2014

Recurrently mutated pathways in small B-cell lymphoid neoplasms Pathway U-CLL M-CLL MCL FL GCB NFkB + +/- Chromatin Remodeling + + TLR/MYD88 +/- DNA-damage + + NOTCH1/2 + +/- + + SF3B1 (RNA metabolism) + +/- SMZL HCL BRAF +

Clinical Relevance of Mutational Profiles in Lymphoid Neoplasms Diagnostic criteria to refine entities Identification of subsets of patients Prognostic and predictive significance Monitoring disease evolution: Dynamic evolution of mutational landscape Targets for therapy: Actionable mutations

Updating WHO classification What s new in small B-cell lymphomas? Refinement of definitions, diagnostic criteria and terminology MBL variants, LPL, in situ Follicular and Mantle cell neoplasias Inclusion of new defined entities and variants FL t(14;18) negative, MCL variants Provisional entities: Updates and open questions FL Pediatric type, IRF4 + LBCL, Relevance and clinical impact of NGS