in tumor suppressor genes such as p16 or p53, and often have a complex karyotype including tetraploidy (92 chromosomes). Thus, although all such cases can now be recognized within the spectrum of MCL because of characteristic staining for cyclin D1, the diverse clinical behavior is beginning to be explained by specific genetic alterations evident within the neoplastic cells.

CURRENT APPROACH

The current approach to the classification of non-Hodgkin’s lymphomas takes advantage of all of the diagnostic tools available to reach an accurate diagnosis. As detailed above, some lymphoma entities can be reliably diagnosed using morphology alone. Examples would include most follicular lymphomas and small lymphocytic lymphoma (SLL). A diffuse infiltrate of large lymphoid cells now requires that one first establish lineage (B-cell vs. T-cell) followed by more targeted immunophenotypic studies that help with specific classification. For example, mediastinal large B-cell lymphoma requires clinical input to know that the patient’s main site of tumor is the mediastinum. Primary effusion lymphoma typically occurs in the setting of HIV infection, characterized by an effusion without mass-like lesions. The immunophenotype is often ambiguous (lack of B-cell-associated markers, CD19 and CD20), but molecular genetic evidence of commitment to a B-cell lineage with clonal immunoglobulin gene rearrangements is virtually always present. The neoplastic cells usually show coinfection with both Epstein–Barr virus (EBV) and human herpes virus-8 (HHV-8). Nasal T-=natural killer (T=NK) cell lymphoma most frequently occurs in the nasopharynx and is virtually always associated with latent EBV infection in the neoplastic cells. The tumor cells express CD56, but often lack T-cell gene rearrangements reflecting their true natural killer (NK) cell lineage. Anaplastic large cell lymphomas can occur as systemic lymphomas or as isolated cutaneous lesions. Biopsies of either disorder reveal a pleomorphic lymphoid infiltrate with large, CD30-positive malignant cells and evidence of T-cell clonality. However, the primary cutaneous lesions do not express anaplastic lymphoma kinase (ALK)

Table 5 Approach to Diagnosis of Lymphoma

1.Clinical correlation

2.Morphologic assessment

3.Immunophenotypic studies

4.Molecular genetic data

5.Cytogenetics

6.Gene expression analysis (not routine)

protein, whereas virtually all of the systemic cases do, providing a unique immunophenotypic signature to distinguish these tumors. Both SLL and MCL are examples of CD5-positive B-cell lymphomas. However, they are clinically quite distinct. MCL is an aggressive lymphoma whereas SLL tends to be quite indolent. A combination of immunophenotypic data and molecular cytogenetic results can be used to clearly distinguish these two neoplasms. SLL is usually dim CD20-positive, expresses CD23, and may express CD11c. MCL on the other hand, brightly expresses CD20, and does not express CD23 or CD11c. MCL is characterized by a recurrent cytogenetic abnormality, the t(11;14), which is not seen in SLL. Importantly, as a result of the t(11;14), MCL expresses cyclin D1, which is not expressed by the neoplastic cells of SLL. Moreover, MCL cases with pleomorphic morphology were not considered part of the spectrum of MCL; however, the routine use of immunostains to cyclin D1 is now used to make the diagnosis in such cases, establishing them as extreme examples of the morphologic diversity of MCL. In summary, the current approach to lymphoma classification may take advantage of all of these approaches in any given case to more precisely render an accurate diagnosis (Table 5). The ultimate goal of these efforts is to provide our clinical colleagues with precise diagnoses that may have both prognostic and therapeutic consequences.

LYMPHOMAS OF THE OCULAR ADNEXA

AND EYE

While lymphomas of the eye itself are very rare, extranodal lymphomas of the ocular adnexal, including the conjunctiva,

eyelids, lacrimal gland, and orbit, constitute as many as 5–8% of all extranodal non-Hodgkin’s lymphomas. The vast majority of the ocular adnexa lymphomas are extranodal marginal zone B-cell lymphomas of MALT type, while intraocular lymphomas are virtually all diffuse large B-cell lymphomas (DLBCL). Most of the progress in distinguishing low-grade non-Hodgkin’s lymphomas involving the orbit from atypical lymphoid hyperplasia (so-called pseudolymphoma) has been the result of improvements in diagnostic techniques for the demonstration of B-cell monoclonality using either immunohistochemical techniques for light chain restriction or gene rearrangement studies showing monoclonal immunoglobulin (IgH) rearrangements. Although clonality is not synonymous with malignancy, it certainly favors a lymphoma. Importantly, the presence of clonality does not necessarily translate into clinically aggressive disease.

MALT lymphomas at this site are thought to arise following acquired reactive MALT tissue that follows chronic antigenic stimulation. Hence, a proliferation of polyclonal B-cells occurs followed in some cases by the emergence of a dominant B-cell clone. The histopathology is characteristic of MALT lymphomas at other sites, with reactive lymphoid follicles, plasma cells, a proliferation of centrocytes-like cells, and lympho-epithelial lesions. Scattered, large transformed cells may be present. The diagnosis can typically be established by immunostains demonstrating an excess of B-lymphocytes with light chain restriction evident in some cases. B-cells are seen lying within the epithelium (lympho-epithelial lesions) and can be highlighted with stains for cytokeratin. Gene rearrangement studies typically reveal a clonal IgH rearrangement. If cytogenetic studies are performed, presence of a t(11;18)(q21;q21) may be found. This novel translocation has been documented in MALT lymphomas from a variety of different anatomical sites. Others lymphomas at this site are uncommon, but include primary follicular lymphoma, lymphoplasmacytic lymphoma, and mantle cell lymphoma. Immunostains together with molecular genetic studies can usually provide the necessary data to allow a distinction between these lymphoma subtypes.

FUTURE DIRECTIONS

Current research efforts in non-Hodgkin’s lymphoma are taking several directions. Most are designed to improve our understanding of the biology, but may also have the added advantage of providing insights into determining prognosis and providing rationale targets for future therapies. One of the new technologies that is critical in this effort is gene expression profiling using microarrays. These techniques utilize tumor tissue mRNA to survey genome-wide expression profiles from individual cases. Consequently, in contrast to the past where typically a single marker was studied in a cohort of patients, this technique allows 15,000–30,000 data points to be generated for each patient. The overwhelming amount of data now requires bio-informatic assistance for proper interpretation, but has already led to significant insights into the heterogeneous biology of some lymphoid tumors, the most recent example being DLBCL. The WHO currently groups the majority of such cases under the rubric ‘‘unspecified DLBCL.’’ Gene expression profiling has now revealed at least two or three major subtypes of disease based on cell of origin. Not only have these data provided an improved understanding of the biology, but also have been shown to be powerful predictors of clinical behavior. In the near future, similar techniques using customized ‘‘mini-chips’’ may become part of the routine diagnostic armamentarium.

REFERENCES

1.Anonymous. National Cancer Institute sponsored study of classifications of non-Hodgkin’s lymphomas: summary and description of a working formulation for clinical usage. The Non-Hodgkin’s Lymphoma Pathologic Classification Project. Cancer 1982; 49(10):2112–2135.

2.Anonymous. A clinical evaluation of the international lymphoma study group classification of non-Hodgkin’s lymphoma. Blood 1997; 89(11):3903–3918.

3.Harris NL, Jaffe ES, Diebold J, Flandrin G, Muller-Hermelink HK, Vardiman J, Lister TA, Bloomfield CD. World Health Organization classification of neoplastic diseases of the hematopoietic and lymphoid tissues: report of the Clinical Advisory Committee Meeting, House, Virginia, November 1997. J Clin Oncol 1999; 17(12):3835–3849.

4.Harris NL, Jaffe ES, Stein H, Banks PM, Chan JK, Cleary ML, Delsol G, De Wolf-Peeters C, Falini B, Gatter KC. A revised European–American classification of lymphoid neoplasms: a proposal from the International Lymphoma Study Group. Blood 1994; 84(5):1361–1392.

5.Jaffe ES, Harris NL, Stein H, Vardiman JW. World Health Organization Classification of Tumours: Tumours of Haematopoietic and Lymphoid Tissues. Lyon: IARC Press, 2001.

6.Rosenwald A, Wright G, Chan WC, Connors JM, Campo E, Fisher RI, Gascoyne RD, Muller-Hermelink HK, Smeland EB, Giltnane JM, Hurt EM, Zhao H, Averett L, Yang L, Wilson WH, Jaffe ES, Simon R, Klausner RD, Powell J, Duffey PL, Longo DL, Greiner TC, Weisenburger DD, Sanger WG, Dave BJ, Lynch JC, Vose J, Armitage JO, Montserrat E, LopezGuillermo A, Grogan TM, Miller TP, LeBlanc M, Ott G, Kvaloy S, Delabie J, Holte H, Krajci P, Stokke T, Staudt LM. The use of molecular profiling to predict survival after chemotherapy for diffuse large-B-cell lymphoma. N Engl J Med 2002; 346(25): 1937–1947.