Shashikant Chittal

Monoclonal antibodies in the diagnosis of Hodgkin's disease. The search for a rational panel.

A novel, comprehensive panel of monoclonal antibodies was tested in a large series of routinely processed lymph node biopsy specimens from patients with Hodgkins disease (69 cases), with the object of developing either definitive or adjunctive diagnostic criteria. B- and T-cell lymphomas and reactive states that could mimic Hodgkins disease were also assessed with the same monoclonal antibody panel. In addition to the popularly used anti-Leu-M1 (CD15), the panel included the recently produced Ber-H2 (CD30) antibody, which detects a formalin-resistant epitope of the Ki-1 antigen. The other monoclonal antibodies were directed against epithelial membrane antigen (Dako-EMA) and leukocyte common antigen (Dako-LC) (CD45), as well as B-cell (LN-1 and LN-2) and T-cell (MT1) associated antigens. The results showed clear phenotypic separation of nodular lymphocyte predominant subtype of Hodgkins disease from other subtypes. The lymphocytic and histocytic cells of nodular lymphocyte predominant Hodgkins disease were reactive for LN-1 (all cases) and anti-EMA (most cases) but negative for anti-Leu-M1 and Ber-H2. Within the other subtypes—i.e. nodular sclerosis and mixed cellularity—nearly all Reed-Steinberg cells and Hodgkins cells were positive for both anti-Leu-M1 and Ber-H2. Ber-H2 monoclonal antibody was observed to react more frequently with Reed-Sternberg cells and Hodgkins cells in Bouins- or formalin-fixed tissues. Pleomorphic T-cell lymphomas, which could mimic Hodgkins disease on morphology, created the same problem on phenotypic analysis. However, MT1 identified a significant proportion of T-cell lymphomas with Reed-Sternberg-like cells, having proven negative for Reed-Sternberg cells and Hodgkins cells in Hodgkins disease. Thus, a combination of anti-Leu-M1, Ber-H2, anti-EMA, LN-1, and MT1 monoclonal antibodies appears at present to be the most useful panel for the diagnosis and the differential diagnosis of Hodgkins disease.

Further phenotypic evidence that nodular, lymphocyte-predominant Hodgkin's disease is a large B-cell lymphoma in evolution.

Five cases of nodular, lymphocyte predominant Hodgkins disease (nLP HD), in which an association with (n = 3) and transformation to (n = 2) large cell lymphoma (LCL) were found, were studied with monoclonal antibodies against B-, T-, and Reed-Sternberg (R-S) cell-associated antigens and epithelial membrane antigen (EMA) on paraffin sections. Both lymphocytic (L) and histiocytic (H) cells of nLP HD and lymphoma cells of LCL expressed multiple B-cell-associated antigens (detected by LN-l/CDw75, L26, MB2, DBB.42, DBA.44, DND.53, DNA.7 antibodies) but did not react with antibodies against T-cell-associated (MTl, UCHLl/ CD45RO) (one exception for CD45RO in LCL) and R-S cell-associated (Leu-Ml/CD15, Ber-H2/CD30) antigens. EMA was expressed by L and H cells in all cases and conserved in LCL cells, emphasizing the frequent expression of EMA by the diagnostic cells of nLP HD. An antibody (BNH9) against blood group-related antigens (H and Y oligosaccharide antigens) that does not normally react with lymphoid cells was found to be reactive with few L and H cells in two of five and most LCL cells in four of five cases. The finding might be indicative of abnormal activation of lymphoid cells. The data reinforce current implications that nLP HD is a B-cell malignancy in evolution and that it is not truly representative of Hodgkins disease in terms of biological and clinical behavior.

Primary anterior mediastinal b-cell lymphoma. A clinicopathologic and immunohistochemical study of 16 cases

Sixteen cases of primary anterior mediastinal B‐cell lymphoma were characterized by morphologic, immunophenotypic, and clinical profiles. Twelve were men and four were women. The median age was 42 years. Virtually all tumors were of large cell type. Three main morphologic categories were identified, with one rare exception. In some tumors, the cells were compatible with centrocytes and centroblasts (four). Others had cells readily identifiable as centroblasts (six). Both these groups had a variable proportion of cells with multilobed nuclei. A third group was composed mainly of unclassifiable cells with multilobed nuclei (five). All had discernible sclerosis of varying intensity. A wider range of morphologic features and different sex distribution was noticed in comparison with previously reported clear cell features and younger women. The dominant phenotype of these B‐cell lymphomas was CD19+, CD22+, CD37+, CD21–, CD30–, CD10–, CD5–, and Ig‐negative. The finding of CD21–, Ig‐negative phenotype, as observed by the authors and others, overlaps with some high‐grade lymphomas of follicular center cell origin but is thought to bear similarity to a noncirculating population of thymic medullary B‐cells. The tumors attained large size without peripheral dissemination and responded to chemotherapy as well as radiotherapy.

Diagnostic features of primary malignant lymphomas of the thyroid with monoclonal antibodies

Primary malignant lymphomas (ML) of the thyroid are rare and their conclusive morphologic diagnosis is not always possible. The authors report diagnostic features of 11 cases of ML and one case of plasmacytoma of thyroid compared with chronic lymphocytic thyroiditis and undifferentiated carcinomas of thyroid in an immunohistochemical study using monoclonal antibodies (MoAb). The lymphoid nature of tumors could be identified in all cases with three MoAb on paraffin sections. In ML, tumor cells expressed leucocyte common antigen (Dako‐LC+) with negativity for epithelial membrane antigen (Dako‐EMA—) and cytokeratin (KL1—). Newer MoAb identifying B‐cell (LN‐1, LN‐2, MB2) and T‐cell‐associated antigens (MT1, UCHL1) not denatured by fixation, revealed B‐cell nature of tumor cells in all cases of ML. Among anti‐B MoAb, LN‐1 and MB2 were most consistent in their reactivity. In cryostat sections of three ML cases, the tumor cells expressed one or more B‐cell‐associated antigens. Plasmacytoma was negative for Dako‐LC and KL1 but positive for Dako‐EMA and monotypic cytoplasmic Ig.

Immunohistochemical profile of cutaneous B-cell lymphoma on cryostat and paraffin sections

Thirty cases of primary (23 cases) and secondary (seven cases) cutaneous B-cell lymphoma (CBCL) were studied by immunohistochemistry using a selected monoclonal antibody (MoAb) panel on both cryostat and paraffin sections. On cryostat sections all CBCL so tested were positive for surface membrane immunoglobulins (IgMk most often) and B-cell antigens (CD22+, CD37+ ) with a variable T-cell-reactive component identified by MoAbs against T-cell antigens (CD2, CD3, CD4, CD5, CD8). CD4-positive stromal T-cells were usually more numerous than CDS-positive cells. A strong (50–75% of total cells) stromal T-cell (CD2+, CD3+ ) reaction was found in centroblastic-centrocytic lymphoma. Small numbers of CD1+ Langerhans cells were found in most cases, but they were present in large numbers in follicular lymphoma. On paraffin sections, a combination of MoAbs against B-associated antigens (LN-1, MB2) identified B-cell lineage in virtually all cases of CBCL. CBCL was negative for MoAbs against T-associated antigens (MT1, UCHL1) with rare exceptions (two cases). However, MT1 and UCHL1 combined identified the T-cell nature of all cases of nonepidermotropic, nonmycosis T-cell lymphoma, which were initially predictive of B-lineage by histologic pattern.

Follicular lymphoma with abundant PAS-positive extracellular material: immunohistochemical and ultrastructural observations

A finding of large amounts of extracellular amorphous material is rare in malignant lymphomas. We report four such cases of follicular lymphoma that were investigated by electron microscopy and immunohistochemistry using monoclonal antibodies. This extracellular material was clearly different from the sclero-hyalinosis commonly found in malignant lymphomas. Our observations revealed that the material is ultrastructurally composed of membranous structures, membrane-bound vesicles, and electron-dense bodies. Immunohistochemistry confirmed its neoplastic cell origin and its identity with cell membrane antigens (Dako-LC+, anti-B antibodies positive). In one case investigated using frozen sections, it showed the same immunophenotype as the neoplastic cells: SB4+ (CD19), SB2+ (CD21), TO15+ (CD22), SB3+ (CD37), IgML+, and Calla+. The mechanism for accumulation of this extracellular amorphous material is probably a function of a deregulation of cell wall synthesis and an exocytosis of cell membranes.

Immunohistochemical detection of post-therapy residual testicular lymphoblasts in childhood acute lymphoblastic leukemia (ALL)

The aim of this study was to evaluate the diagnostic value of immunohistochemistry with monoclonal antibodies (MoAbs) in detecting residual blast cells in testicular biopsies from children with acute lymphoblastic leukemia (ALL). In a prospective study of 26 patients, testicular biopsies were performed after completion of therapy, and the average follow-up after biopsies was 29 months. After immunostaining, seven patients with negative biopsies on routine histology showed scattered, strongly calla-positive cells as well as cells reacting with anti-B (CD22) MoAb. Among these seven patients with residual blast cells, four had relapsed either in testes (n = 1), bone marrow and testes (n = 1), or in the bone marrow (n = 2). In contrast, among the 15 patients without residual blast cells, all but 1 remained in complete remission. In four other cases no definite conclusion was possible after immunohistochemical study. Four testicular biopsies from patients with occult infiltration were used as positive controls. Negative controls consisted of testicular biopsies from children with testicular ectopia and postmortem testicular tissue specimens. Results suggest that the risk of relapse is significantly higher in patients with positive immunohistochemical findings indicating persistent residual blast cells. However, the predictive value of these findings requires confirmation on a larger number of cases to have therapeutic implications.