Comprehensive gene expression profiling and immunohistochemical studies support application of immunophenotypic algorithm for molecular subtype classification in diffuse large B-cell lymphoma: a report from the International DLBCL Rituximab-CHOP Consortium Program Study.

Gene expression profiling (GEP) has stratified diffuse large B-cell lymphoma (DLBCL) into molecular subgroups that correspond to different stages of lymphocyte development-namely germinal center B-cell like and activated B-cell like. This classification has prognostic significance, but GEP is expensive and not readily applicable into daily practice, which has lead to immunohistochemical algorithms proposed as a surrogate for GEP analysis. We assembled tissue microarrays from 475 de novo DLBCL patients who were treated with rituximab-CHOP chemotherapy. All cases were successfully profiled by GEP on formalin-fixed, paraffin-embedded tissue samples. Sections were stained with antibodies reactive with CD10, GCET1, FOXP1, MUM1 and BCL6 and cases were classified following a rationale of sequential steps of differentiation of B cells. Cutoffs for each marker were obtained using receiver-operating characteristic curves, obviating the need for any arbitrary method. An algorithm based on the expression of CD10, FOXP1 and BCL6 was developed that had a simpler structure than other recently proposed algorithms and 92.6% concordance with GEP. In multivariate analysis, both the International Prognostic Index and our proposed algorithm were significant independent predictors of progression-free and overall survival. In conclusion, this algorithm effectively predicts prognosis of DLBCL patients matching GEP subgroups in the era of rituximab therapy.

Acute promyelocytic leukemia, hypogranular variant, with uncharacteristic staining with chloroacetate esterase.

A diagnosis of the hypogranular variant of acute promyelocytic leukemia (APLv) may be difficult to establish based on cytomorphology alone. However, the great majority of cases have a classical immunophenotype by flow cytometric immunophenotyping (FCI) (CD13+, CD33+, dim CD64+, HLA-DR-, and CD34-) and a classical enzyme cytochemical (EC) staining pattern. [intensely staining with myeloperoxidase, Sudan Black B, and chloroacetate esterase (CAE) and negative with alpha'-naphthyl acetate and butyrate esterases]. Although the immunophenotype of APLv by FCI has varied in the literature (HLA-DR +/- and CD34 +/-), the EC staining pattern has remained constant. We report a case of APLv with characteristic cytomorphology, compatible FCI data (CD13+, CD33+, dim CD64+, HLA-DR +/-, and CD34-), chromosomal detection of t(15; 17), and molecular detection of the PML/RAR alpha fusion gene; however, staining of the leukemic cells with CAE was quite uncharacteristic. We describe our findings.

Composite lymphoma of Hodgkin lymphoma and mycosis fungoides: previously undescribed in the same extracutaneous site.

Mycosis fungoides (MF), diagnosed and limited to the skin, has been associated with the subsequent development of Hodgkin lymphoma (HL), most commonly of the nodular sclerosing (NS) subtype. In the previously described cases, there are none in which the extracutaneous tissue was simultaneously involved by HL and residual/relapsing MF. Here we report a case of HL, mixed cellularity (MC) subtype, arising in an inguinal lymph node in a patient with a previous diagnosis of MF. We describe the immunophenotypic, histologic and immunohistochemical findings of a composite lymphoma containing the HL, MC subtype and MT. The importance of detecting MF in addition to the HL in the extracutaneous site with available diagnostic modalities is highlighted.

CD15(+) acute lymphoblastic leukemia and subsequent monoblastic leukemia: persistence of t(4;11) abnormality and B-cell gene rearrangement.

The abnormality in the translocation of chromosomes 4 and 11 (t[4;11]) has been characteristically associated with calla-negative CD15(+) acute lymphoblastic leukemia (ALL) of early pre-B-cell origin. Transformation of a lymphoblastoid to a monoblastoid morphologic structure has rarely been described at relapse in these cases; however, these cases have lacked flow cytometric immunophenotyping (FCI) and genotypic studies (GS) to define the immunophenotype of and the presence of a B-cell gene rearrangement in the monoblastoid component. We report a case of CD15(+), CD10(-) ALL of early pre-B-cell origin defined by morphologic testing and FCI with the t(4;11) abnormality. At relapse, the morphologic testing, enzyme cytochemistry, and FCI data were characteristic of monoblastic leukemia. The t(4;11) abnormality persisted with associated additional chromosomal abnormalities, and the monoblasts contained a B-cell gene rearrangement by GS. These findings support the concept that both processes arose from a multipotential progenitor cell.

CD30 expression in follicular lymphoma.

CONTEXT: CD30(+) anaplastic large cell lymphomas were originally described as being of T-cell, null cell, and B-cell origin. CD30, however, is not a specific marker of anaplastic large cell lymphoma and has been found to be expressed in reactive as well as neoplastic populations as a probable activation marker. In addition, CD30(+) cells have also been described in both diffuse large B-cell and follicular lymphomas (FLs), resembling the pattern seen in reactive tonsils and lymph nodes. OBJECTIVE: We report an index case of FL with CD30 expression, which on initial touch preparations and flow cytometric immunophenotyping revealed a prominent population of CD30(+) cells with marked cellular pleomorphism (anaplasia) in a background of typical FL. Immunohistochemistry of the paraffin section for CD30 in our index case confirmed unequivocal CD30(+) pleomorphic cells in the malignant nodules in occasional clusters. This case prompted a study of additional cases of FL for pattern of immunoreactivity with CD30 on paraffin sections. DESIGN: Twenty-two additional cases of FL (grades 1-3) were retrieved for CD30 immunoperoxidase staining as in the index case. RESULTS: This study demonstrated 32% of the additional cases of FL had definitive CD30(+), large, pleomorphic malignant cells by paraffin immunohistochemistry. In 2 cases (9%), the pattern of immunoreactivity with CD30 showed clustering and variable staining of large cells, as our index case. CONCLUSION: This study underscores the morphologic and immunophenotypic spectrum of FL that includes CD30 staining and cellular pleomorphism.

Evaluation of bone marrow specimens with acute myelogenous leukemia for CD34, CD15, CD117, and myeloperoxidase.

CONTEXT: Immunophenotyping of bone marrow (BM) specimens with acute myelogenous leukemia (AML) may be performed by flow cytometric (FC) or immunohistochemical (IH) techniques. Some markers (CD34, CD15, and CD117) are available for both techniques. Myeloperoxidase (MPO) analysis may be performed by enzyme cytochemical (EC) or IH techniques. OBJECTIVE: To determine the reliability of these markers and MPO by these techniques, we designed a study to compare the results of analyses of these markers and MPO by FC (CD34, CD15, and CD117), EC (MPO), and IH (CD34, CD15, CD117, and MPO) techniques. MATERIALS AND METHODS: Twenty-nine AMLs formed the basis of the study. These AMLs all had been immunophenotyped previously by FC analysis; 27 also had had EC analysis performed. Of the AMLs, 29 had BM core biopsies and 26 had BM clots that could be evaluated. The paraffin blocks of the 29 BM core biopsies and 26 BM clots were stained for CD34, CD117, MPO, and CD15. These results were compared with results by FC analysis (CD34, CD15, and CD117) and EC analysis (MPO). RESULTS: Immunodetection of CD34 expression in AML had a similar sensitivity by FC and IH techniques. Immunodetection of CD15 and CD117 had a higher sensitivity by FC analysis than by IH analysis. Detection of MPO by IH analysis was more sensitive than by EC analysis. There was no correlation of French-American-British (FAB) subtype of AML with CD34 or CD117 expression. Expression of CD15 was associated with AMLs with a monocytic component. Myeloperoxidase reactivity by IH analysis was observed in AMLs originally FAB subtyped as M0. CONCLUSIONS: CD34 can be equally detected by FC and IH techniques. CD15 and CD117 are better detected by FC analysis and MPO is better detected by IH analysis.

Paraffin immunoreactivity of CD10, CDw75, and Bcl-6 in follicle center cell lymphoma.

Follicle center cell lymphoma(FCCL) has the following immunophenotype(IP): sIg+, Pan B+, CD10+/-, CD5-, CD23-/+, CD43-, CD11c-, CD25-. In addition, reactivities of a malignant lymphoma with CDw75(LN-1) and bcl-6 are considered indicators of FCCL. Bcl-6 expression is common in Grade 1 FCCL (100%) and rare in other indolent B-cell lymphomas(BCL). In contrast, bcl-2 expression is common in FCCL (80%) and in other BCL subtypes. Since no previous study has correlated paraffin immunoreactivity(PIR) of CD10, CDw75, and bcl-6 in FCCL (Grades 1-3), this is this study's purpose. Twenty-nine FCCL's were identified and reviewed (6, Grade 1; 10, Grade 2; 13, Grade 3) from the Division of Hematopathology, St. Louis University. The diagnoses were based on morphology and immunohistochemistry(IH)(21 cases) +/- the flow cytometric IP(14 cases). The paraffin blocks were stained for CD10 (Novacastra, Vector Laboratories, Burlingame, CA), CDw75 and bcl-6 (DAKO Corporation, Carpinteria, CA). Results showed that, CD10 by paraffin IH(PIH) was positive in 23 [18(strong); 3(moderate); 2(weak)] and negative in 6(3, Grade 2; 3, Grade 3). All CD10-cases were CDw75+; 4, bcl-6+. The two CD10-, bcl-6-cases were Grade 2. CDw75 was positive in 28 cases [16(strong); 11(moderate); 1(weak)] and negative in 1 (Grade 3; CD10+, bcl-2+, bcl-6+). Bcl-6 was positive in 26 [16(strong); 6(moderate); 4(weak)] and negative in 3(Grade 2's). Thus, the sensitivity of CD10, CDw75, and bcl-6 by PIH for FCCL was 79%, 97%, and 90%, respectively. Of the three stains evaluated by PIH in FCCL, CDw75 was the most sensitive, closely followed by bcl-6. CD10 was least sensitive-79%. By combining these 3 stains, the sensitivity was 100%; thus, a combined approach is recommended.

Mantle cell leukemia, prolymphocytoid type: a rarely described form.

Leukemic manifestations of mantle cell lymphoma are seen in a minority of cases, usually associated with extensive tumor. Usually the neoplastic cells in the peripheral blood resemble mantle cells with a mature chromatin pattern and irregular nuclear contours, or less frequently with a more "blastic" chromatin pattern. A prolymphocytic leukemia with t(11;14)(q13;q32) has previously been reported; however, a complete flow cytometric immunophenotypic profile was lacking. Mantle cell leukemia, prolymphocytoid type with complete flow cytometric data has not previously been described and is the purpose of this report. We report such a case in a 90 year-old female who presented with an elevated white blood cell count. The diagnosis was based on flow cytometric immunophenotyping and the cytomorphology of the peripheral blood combined with cyclin D1 immunohistochemical staining of the bone marrow. We describe our findings and her clinical course in order to heighten awareness of this previously rarely described entity.

Large cell variants of CD5+, CD23- B-cell lymphoma/leukemia.

CONTEXT: Mantle cell lymphoma (MCL), and its leukemic phase, constitute a well-studied hematologic malignancy with known overall survival, prognostic indicators, morphologic findings at diagnosis and in bone marrow, and known incidence of the bcl-1 immunoglobulin gene rearrangement. Large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype (CD5+, CD23-), including but not limited to blastic MCL, prolymphocytoid MCL, blastic mantle cell leukemia, and prolymphocytic mantle cell leukemia, are not as well characterized. Although blastic MCL is known to be associated with a shorter overall survival than conventional MCL, the large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype have not been described as fully as conventional MCL. OBJECTIVE: The purpose of the present study was to describe the large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype. DESIGN: Nineteen cases of large cell variants of CD5+, CD23- B-cell lymphoma/leukemia are reviewed and described in regard to morphology, bone marrow morphological findings, Cyclin D1 immunostaining, and bcl-1 analysis. Clinical data were not available owing to the varied clinical sources of the specimens. SETTING: Tertiary-care academic institution. RESULTS: Lymph node involvement in blastic CD5+, CD23- B-cell lymphoma was diffuse (100%) with a nodular component (33%) or focal mantle zone pattern (10%). Bone marrow involvement in blastic CD5+, CD23- B-cell lymphoma was seen in only 27% of cases and was composed predominantly of small, slightly irregular lymphocytes. Cyclin D1 was demonstrated in 60% of the 15 cases analyzed and more sensitive in B5-fixed tissue. Bcl-1 (performed in 5 cases) was not detected in the 4 cases of blastic CD5+, CD23- B-cell lymphoma analyzed and was detected in the case of the prolymphocytoid MCL. Cyclin D1 was demonstrated in all 4 bcl-1 negative cases and was negative in the bcl-1 positive prolymphocytoid MCL. CONCLUSION: Careful analysis of clinical data, morphology, immunophenotype, Cyclin D1 expression, and molecular analysis are required to differentiate the unusual large cell variants of MCL from other processes.

Contribution of flow cytometric immunophenotyping to the evaluation of tissues with suspected lymphoma?

BACKGROUND: A critical analysis of the contribution of flow cytometric immunophenotyping (FCI) to the evaluation of lymph nodes and extranodal tissues with suspected lymphoma by a large, retrospective approach has not been reported previously and represents the purpose of this study. METHODS: A total of 278 lymph nodes and 95 extranodal tissue specimens submitted over a 2-year period with complete histologic, FCI, and immunohistochemical (IH) data formed the basis of the study. RESULTS: The FCI data contributed significantly to or was consistent with the final tissue diagnosis in the majority (94%) of the tissue samples. There is no well-described utility of flow cytometry markers for Hodgkin's lymphoma (HL) due to the usual scarcity of tumor cells in the final cell suspensions obtained from these tumors. However, the FCI data excluded non-Hodgkin's lymphoma (NHL) and suggested the possible usefulness of CD15 and CD30 by FCI in HL. In addition, immunophenotypic data by FCI in combination with touch imprint cytomorphology was useful in excluding a diagnosis of NHL in cases of nonhematopoietic malignancies and was particularly useful in defining the following hematopoietic tumors and malignancies: thymoma, T-cell lymphoblastic lymphoma, leukemia cutis, and plasma cell dyscrasia. Thus, IH was not essential for the diagnosis in these latter cases and was performed in only two cases (one thymoma and one plasma cell dyscrasia). Of interest, FCI supported the diagnosis in 3 cases of Ewing's sarcoma/primitive neuroectodermal tumor by detection of CD56 on the surface of the malignant cell. Only 11% of NHL were "negative" by FCI (i.e., an aberrant T-cell or monoclonal B-cell population was not identified). Reasons for these discrepancies included partial tissue involvement by the NHL with sampling differences, T-cell rich or lymphohistiocytic-rich variants with a small population of monoclonal B cells, marked tumoral sclerosis, poor tumor preservation, and T-cell NHL without an aberrant immunophenotype. Only 60% of CD30+ anaplastic large cell lymphomas (ALCL) were CD30+ by FCI. CONCLUSIONS: FCI data should always be correlated with light microscopy if no FCI abnormalities are detected; IH may need to be performed in selected cases. It is less necessary to perform microscopic examination of tissues when the FCI data are positive and indisputable. However, in selected cases in which FCI data is diagnostic, microscopic observations may provide additional information due to sampling.

Simultaneous presence of t(2;8)(p12;q24) and t(14;18)(q32;q21) in a B-cell lymphoproliferative disorder with features suggestive of an aggressive variant of splenic marginal-zone lymphoma.

We report a case of an aggressive variant of splenic marginal-zone lymphona (SMZL) with circulating villous lymphocytes. The karyotype of all examined cells had multiple structural and numerical abnormalities, including two lymphoma characteristic translocations, t(2;8)(p12;q24) and t(14;18)(q32;q21). Based on a literature review of cytogenetic aberrations of splenic lymphoma with villous lymphocytes (SLVL) and SMZL, this is apparently the first documentation of these two translocations in a case of SMZL, and could reflect the heterogeneity of the disorder.

Malignant mesothelioma with CD30-positivity. A case report and review of the literature.

The activation marker CD3O is useful in the diagnosis of Hodgkin and non-Hodgkin lymphomas. It has also been described in nonhematopoietic tumors, including pancreatic carcinomas, salivary gland tumors, and embryonal carcinomas. We report a case of malignant mesothelioma with intense CD30 positivity. This finding has not previously been described and is important in broadening the differential diagnosis of a CD30(+) cohesive large cell malignancy.

Low-grade B-cell lymphoma and concomitant extensive sarcoidlike granulomas: a case report and review of the literature.

Sarcoidlike granulomas may occur in association with Hodgkin lymphoma and non-Hodgkin lymphoma. The granulomas may be concomitant and so extensive that they obscure the malignant process. In addition, a sarcoidosis-lymphoma syndrome has been described in which there appears to be a relationship between sarcoidosis and the development of a lymphoproliferative disorder. We report a case of a low-grade B-cell lymphoma with concomitant extensive sarcoidlike granulomas. The patient had no diagnostic clinical evidence of sarcoidosis, although she had an elevated serum calcium level and increased serum angiotensin converting enzyme activity. Increased serum calcium and serum angiotensin-converting enzyme activity have been associated with clinical sarcoidosis but have also occasionally been described in association with Hodgkin lymphoma and non-Hodgkin lymphoma without evidence of sarcoidosis. We describe our findings and illustrate the usefulness of immunoperoxidase immunophenotyping techniques in such a case.

Mucosa-associated lymphoid tissue-type lymphomas occurring in post-transplantation patients.

Post-transplantation lymphoproliferative disorders (PTLDs) are usually Epstein-Barr virus (EBV)-associated B-cell lymphoproliferative disorders that vary in their morphologic spectrum. Extranodal marginal zone lymphomas of the mucosa-associated lymphoid tissue-type (MALT-type) have not been considered to be part of this spectrum. The authors encountered five such cases recently. The clinical, histopathologic, and immunophenotypic features are reported. There were three men and two women with a mean age of 51.2 years (range, 48-63 years). Two patients were cardiac transplant recipients, two patients were liver transplant recipients, and the remaining patient was a renal transplant patient. Sites of lymphoma were the stomach in three patients and the parotid gland in two patients. Mean time to the lymphoma was 84 months after transplantation. All patients had morphologic features of low-grade extranodal marginal zone lymphomas of the MALT-type, and Helicobacter pylori was present in all three gastric cases. All patients exhibited the B-cell immunophenotype and were negative for EBV by in situ hybridization. These lymphomas were treated with a variety of modalities, including reduction of immunosuppression, antibiotics, surgical resection, radiation therapy, and chemotherapy. At last follow-up, one patient had developed signet ring adenocarcinoma at 27 months but had no evidence of PTLD, one patient relapsed at 17 months but is alive with stable disease at 24 months, and the remaining patients were alive without disease at 11, 12, and 14 months. Extranodal low-grade MALT-type lymphomas can occur in the post-transplantation setting and generally develop years after transplant. As seen in immunocompetent patients, EBV appears to play no role in the pathogenesis of these lymphomas. These lymphomas appear to have more in common with MALT-type lymphomas in nonimmunocompromised patients than conventional PTLDs, although they occur in "at-risk" patients due to their immunosuppressive therapy. These lymphomas do not appear to be clinically aggressive. Recognition of MALT-type lymphomas in the post-transplantation setting as an indolent disease avoids unnecessary treatment.

CD30+ anaplastic large-cell lymphoma with aberrant expression of CD13: case report and review of the literature.

CD13 is commonly expressed in hematopoietic malignancies of myelomonocytic origin and has less commonly been described in lymphoid neoplasms, including acute lymphoblastic leukemia, B-cell lymphoproliferative disorders, and plasma cell malignancies. Aberrant CD13 expression has rarely been described in KP-1 (CD68)-positive large-cell lymphomas. However, CD13 positivity has not previously been described in a case of CD30+ (ALK-1+) anaplastic large-cell lymphoma of presumed null-cell origin without histiocytic differentiation. The purpose of this case report is to describe a CD30+ anaplastic large-cell lymphoma of presumed null-cell origin with aberrant expression of CD13. The case illustrates the unique usefulness of immunophenotypic and molecular techniques in establishing the correct diagnosis. The case was referred with a diagnosis of "rule out granulocytic sarcoma versus megakaryocytic malignancy" due to the morphology and a limited flow cytometric immunophenotypic (FCI) panel that had been performed and revealed expression of CD45, HLA-DR, and CD13. Subsequent morphologic review at our institution combined with an expanded FCI panel established the diagnosis. The differential diagnosis of a CD13+ hematopoietic malignancy should include this entity. The prognostic significance of this finding has yet to be determined.

CD5+ true SLL/CLL with plasmacytic differentiation and an unusual 1p36 translocation: case report and review of the literature.

Lymphoplasmacytic lymphoma (LPL) and small lymphocytic lymphoma/chronic lymphocytic leukemia (SLL/CLL)are distinct clinicopathologic entities. Although some cases of SLL/CLL may show plasmacytic differentiation and be associated with monoclonal immunoglobulin in serum, such cases appear to be very rare, and if plasma cell differentiation were marked, differentiation of SLL/CLL from LPL could be difficult. We report a rare case of true CD5-positive small lymphocytic lymphoma/chronic lymphocytic leukemia with unequivocal plasmacytic differentiation. This case also showed an abnormality of chromosome 1p36 not previously described in small lymphocytic lymphoma/chronic lymphocytic leukemia.

CD7 and CD56-positive primary effusion lymphoma in a human immunodeficiency virus-negative host.

Primary effusion lymphoma is an entity with distinctive features. The majority of cases are diagnosed in patients infected with human immunodeficiency virus. We report a case of pleural-based primary effusion lymphoma in an elderly patient negative for human immunodeficiency virus. By flow cytometry, lymphoma cells expressed CD7, CD38, CD45, CD56, HLA-DR, and kappa surface light chains. A monoclonal rearrangement of the immunoglobulin heavy chain and the presence of human herpesvirus 8 genome were detected. Our case lacked CD30 or CD138 with expression of surface light chains. There was strong expression of CD7 and CD56. These findings are unusual or unique in primary effusion lymphoma. Our report suggests that aberrant expression of T cell and natural killer cell markers can be seen in primary effusion lymphoma.

Primary cutaneous T-cell-rich B-cell lymphomas with flow cytometric immunophenotypic findings. Report of 3 cases and review of the literature.

BACKGROUND: Primary cutaneous T-cell-rich B-cell lymphoma is a relatively rare entity that has been diagnosed most commonly using immunohistochemical and molecular techniques. Flow cytometric immunophenotyping (FCI) has not been described in this entity. We report the demonstration of B-cell monoclonality by FCI in 3 cases of primary cutaneous T-cell-rich B-cell lymphoma. METHODS: Clinical and pathologic data were recorded for 3 cases of primary cutaneous T-cell-rich B-cell lymphoma. Immunohistochemical and FCI data were available in all cases; DNA analysis was performed in 1 case. RESULTS: Flow cytometric immunophenotyping revealed a monoclonal B-cell population exclusively in the monocyte (large cell) region in all 3 cases. Immunohistochemistry confirmed the T-cell richness of the infiltrates within the cutaneous lymphomas; T cells accounted for 65% to greater than 90% of the cells within the infiltrates. DNA analysis by polymerase chain reaction in 1 case did not demonstrate a monoclonal rearrangement of the immunoglobulin heavy-chain gene. CONCLUSIONS: Flow cytometric immunophenotyping in primary cutaneous T-cell-rich B-cell lymphoma may be useful in demonstrating monoclonality in these cases, especially if there is selective gating on the relatively small population of cells in the large cell region. The FCI data should be correlated with histology and immunohistochemistry.