II.	Recognition and Classification of Malignant Lymphomas Marsha Kinney, James Cook and Steven Swerdlow

Classification of malignant lymphomas and the avoidance of errors: The first step in avoiding the pitfalls of diagnostic hematopathology is to know your options. This means knowing the specific benign entities that we diagnose including those that mimic lymphomas and being aware of the long list of lymphomas that we are now required to recognize and diagnose. Even if one doesn't master the entire list (a fate left to a very few), one can manage by being extra careful when a lymphoid proliferation doesn't fit well into one of the entities that you do know about. In terms of the long list of lymphomas to master, at least in 2009, there is an internationally agreed upon classification with a just recently published well-illustrated monograph to use – the 2008 revision of the 2001 WHO classification of the haematopoietic and lymphoid tumours. [1] This monograph essentially includes much of what was included in the update of cutaneous lymphomas that were published in the 2005 WHO/EORTC consensus classification for cutaneous lymphomas. [2, 3, 4] The 2008 World Health Organization classification recognizes a list of distinct clinicopathologic entities agreed upon not only by hematopathologists but also by a clinical advisory group of hematologists/oncologists. The WHO classification, divides lymphomas into those of non-Hodgkin and Hodgkin type and then divides the precursor and mature non-Hodgkin lymphomas into those of B-cell or T-cell (or natural killer cell) origin, similar to the original "functional" Lukes/Collins and Kiel classifications from more than three decades ago. 2008 WHO Classification of lymphoid neoplasms [1] PRECURSOR LYMPHOID NEOPLASMS B lymphoblastic leukaemia/lymphoma B lymphoblastic leukaemia/lymphoma, NOS B lymphoblastic leukaemia/lymphoma with recurrent genetic abnormalities B lymphoblastic leukaemia/lymphoma with t(9;22)(q34;q11.2); BCR-ABL1 B lymphoblastic leukaemia/lymphoma with t(v;11q23); MLL rearranged B lymphoblastic leukaemia/lymphoma with t(12;21)(p13;q22); TEL-AML1(ETV6-RUNX1) B lymphoblastic leukaemia/lymphoma with hyperdiploidy B lymphoblastic leukaemia/lymphoma with hypodiploidy (hypodiploid ALL) B lymphoblastic leukaemia/lymphoma with t(5;14)(q31;q32); IL3-IGH B lymphoblastic leukaemia/lymphoma with t(1;19)(q23;p13.3); E2A-PBX1; (TCF3-PBX1) T lymphoblastic leukaemia/lymphoma MATURE B-CELL NEOPLASMS Chronic lymphocytic leukaemia/small lymphocytic lymphoma B-cell prolymphocytic leukaemia Splenic marginal zone lymphoma Hairy cell leukaemia Splenic lymphoma/leukaemia, unclassifiable Splenic diffuse red pulp small B-cell lymphoma Hairy cell leukaemia-variant Lymphoplasmacytic lymphoma Waldenström macroglobulinemia Heavy chain diseases Alpha heavy chain disease Gamma heavy chain disease Mu heavy chain disease Plasma cell myeloma Solitary plasmacytoma of bone Extraosseous plasmacytoma Extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) Nodal marginal zone lymphoma Paediatric nodal marginal zone lymphoma Follicular lymphoma Paediatric follicular lymphoma Primary cutaneous follicle centre lymphoma Mantle cell lymphoma Diffuse large B-cell lymphoma (DLBCL), NOS T-cell/histiocyte rich large B-cell lymphoma Primary DLBCL of the CNS Primary cutaneous DLBCL, leg type EBV positive DLBCL of the elderly DLBCL associated with chronic inflammation Lymphomatoid granulomatosis Primary mediastinal (thymic) large B-cell lymphoma Intravascular large B-cell lymphoma ALK positive DLBCL Plasmablastic lymphoma Large B-cell lymphoma arising in HHV8-associated multicentric Castleman disease Primary effusion lymphoma Burkitt lymphoma B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and Burkitt lymphoma B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and classical Hodgkin lymphoma MATURE T-CELL AND NK-CELL NEOPLASMS T-cell prolymphocytic leukaemia T-cell large granular lymphocytic leukaemia Chronic lymphoproliferative disorder of NK-cells Aggressive NK cell leukaemia Systemic EBV positive T-cell lymphoproliferative disease of childhood Hydroa vacciniforme-like lymphoma Adult T-cell leukaemia/lymphoma Extranodal NK/T cell lymphoma, nasal type Enteropathy-associated T-cell lymphoma Hepatosplenic T-cell lymphoma Subcutaneous panniculitis-like T-cell lymphoma Mycosis fungoides Sézary syndrome Primary cutaneous CD30 positive T-cell lymphoproliferative disorders Lymphomatoid papulosis Primary cutaneous anaplastic large cell lymphoma Primary cutaneous gamma-delta T-cell lymphoma Primary cutaneous CD8 positive aggressive epidermotropic cytotoxic T-cell lymphoma Primary cutaneous CD4 positive small/medium T-cell lymphoma Peripheral T-cell lymphoma, NOS Angioimmunoblastic T-cell lymphoma Anaplastic large cell lymphoma, ALK positive Anaplastic large cell lymphoma, ALK negative HODGKIN LYMPHOMA Nodular lymphocyte predominant Hodgkin lymphoma Classical Hodgkin lymphoma Nodular sclerosis classical Hodgkin lymphoma Lymphocyte-rich classical Hodgkin lymphoma Mixed cellularity classical Hodgkin lymphoma Lymphocyte depleted classical Hodgkin lymphoma POST-TRANSPLANT LYMPHOPROLIFERATIVE DISORDERS (PTLD) Early lesions Plasmacytic hyperplasia Infectious mononucleosis-like PTLD Polymorphic PTLD Monomorphic PTLD (B- and T/NK-cell types)* Classical Hodgkin lymphoma type PTLD* NOS, not otherwise specified. The italicized histologic types are provisional entities, for which the WHO Working Group felt there was insufficient evidence to recognize as distinct diseases at this time. *These lesions are classified according to the disease entity to which they correspond. The second step in avoiding errors is know to how to use histopathology together with a variety of ancillary tools to make the diagnoses that you now know exist . Learning how to judiciously use and wisely interpret the host of special studies that are a part of diagnostic hematopathology are just as critical as knowing the basic criteria for the hematopoietic/lymphoid disorders. Histopathology remains the most important element in evaluating diagnostic lymph node biopsies and extranodal hematopoietic/lymphoid proliferations even in 2009. It is far beyond the scope of this course to discuss the role of cytopathology in these circumstances. The cases that will be presented were hard enough even with entire tissue biopsies available. Immunophenotypic studies, whether performed using flow cytometric and/or immunohistochemical methods are only a photo-finish behind morphologic studies in terms of being essential in most cases that we deal with today. It is an acknowledged challenge knowing what methods to use in which circumstances and dealing with the growing list of antibodies that we have at our disposal. Once again, it is beyond the scope of this course to discuss this topic in general; however, you will see how important immunohistochemical studies were in the cases we will be reviewing today. Cytogenetic studies, essentially required in the diagnosis of acute leukemias, have been utilized much less in dealing with lymphoid disorders. However, they are gaining in both popularity and importance for several reasons. Classical cytogenetic studies (G-banded karyotypes) are used in some institutions for the work-up of lymphomas, even though they are labor intensive and therefore not inexpensive to perform. While not infrequently contributing supportive information to well-worked up cases or providing information that may have prognostic implications, the number of cases where classical cytogenetic studies provide information directly leading to a diagnosis that might otherwise have been missed is very few. [5] Furthermore, there are rare occasions where these studies can be misleading with evidence of a cytogenetic clone in the absence of a recognizable lymphoma. Cytogenetic molecular fluorescence in situ hybridization (ô ) studies, on the other hand, do not require fresh cells and are increasingly popular since they can be performed using cells in paraffin sections or extracted from paraffin blocks if metaphase or interphase cells in cytogenetic preparations or interphase cells on touch imprints are not available. In addition, there are increasing numbers of commercially available probes to look for many of the chromosomal abnormalities that characterize specific types of lymphomas or have prognostic implications. [6, 7, 8] Finally, there are molecular genetic studies, using Southern blot or polymerase chain reaction (PCR) analysis. These studies, also fraught with many hazards, can be critical; however, they are unnecessary in most cases at the current time. They may provide false negative and false positive results. The availability of the "Biomed-2" primers for PCR studies looking for evidence of clonal B-cell or T-cell proliferations may help improve this situation and have been reported to be superior to Southern blot analysis. [9, 10, 11] Remember, however, that just because a test is more sophisticated, it does not necessarily provide a more correct answer. Gene expression profiling is another type of molecular testing that is being used to study lymphomas (and other types of proliferations); however, it is not ready for routine use in diagnostic hematopathology. [12, 13] The third step in giving yourself the best chance at arriving at the correct diagnosis is to use all the information you can lay your hands on including the clinical and other laboratory data. Having an up-to-date list of diagnostic options and knowing or having access to their diagnostic criteria, personally selecting and then integrating the data from all of the above studies, and then educating yourself as much as you can about the patient's clinical and laboratory findings will give you the best chance of staying out of trouble and turn you into a star! References: Swerdlow SH, Campo E, Harris NL, et al. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. IARC: Lyon 2008. Burg G, Kempf W, Cozzio A, et al. WHO/EORTC classification of cutaneous lymphomas 2005: histological and molecular aspects. J Cutan Pathol 32: 647-674, 2005. LeBoit PE, Burg G, Weedon D, et al. Pathology and Genetics of Skin Tumours. IARC Press: Lyon 2006. Willemze R, Jaffe ES, Burg G, et al. WHO-EORTC classification for cutaneous lymphomas. Blood 105: 3768-3785, 2005. Cook JR, Shekhter-Levin S, Swerdlow SH. Utility of routine classical cytogenetic studies in the evaluation of suspected lymphomas: results of 279 consecutive lymph node/extranodal tissue biopsies. Am J Clin Pathol 121: 826-835, 2004. Martin-Subero JI, Gesk S, Harder L, et al. Interphase cytogenetics of hematological neoplasms under the perspective of the novel WHO classification. Anticancer Res 23: 1139-1148, 2003. Paternoster SF, Brockman SR, McClure RF, et al. A new method to extract nuclei from paraffin-embedded tissue to study lymphomas using interphase fluorescence in situ hybridization. Am J Pathol 160: 1967-1972., 2002. Cook JR. Paraffin section interphase fluorescence in situ hybridization in the diagnosis and classification of non-hodgkin lymphomas. Diagn Mol Pathol 13: 197-206, 2004. van Dongen JJ, Langerak AW, Bruggemann M, et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia 17: 2257-2317, 2003. Sandberg Y, van Gastel-Mol EJ, Verhaaf B, et al. BIOMED-2 multiplex immunoglobulin/T-cell receptor polymerase chain reaction protocols can reliably replace Southern blot analysis in routine clonality diagnostics. J Mol Diagn 7: 495-503, 2005. Evans PA, Pott C, Groenen PJ, et al. Significantly improved PCR-based clonality testing in B-cell malignancies by use of multiple immunoglobulin gene targets. Report of the BIOMED-2 Concerted Action BHM4-CT98-3936. Leukemia, 2006. Wiestner A, Staudt LM. Towards molecular diagnosis and targeted therapy of lymphoid malignancies. Semin Hematol 40: 296-307, 2003. Staudt LM. Molecular Diagnosis of the Hematologic Cancers. N Engl J Med 348: 1777-1785, 2003.