Monoclonal Immunoglobulin Gene Rearrangement Research Articles

Molecular pathology assists the diagnosis of lymphoepithelial sialadenitis, Sjögren's syndrome and extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue

Background/purposeLymphoepithelial sialadenitis (LESA), Sjögren's syndrome (SS), and salivary MALT lymphoma are diseases characterized by lymphoepithelial lesions, and the differential diagnosis between them in the salivary glands is challenging. This study aimed to explore clinicopathological and genetic characteristics of the three diseases. Materials and methodsWe retrospectively analyzed the clinical features, the histomorphology, immunohistochemistry, and genetic profiling by polymerase chain reaction (PCR) and next-generation sequencing (NGS). ResultsThere included 68 LESAs, 25 SSs, and 62 MALT lymphomas. Ten cases relapsed in total, and 3 of MALT lymphomas died due to high-level transformation. Immunohistochemical double staining showed FCRL4 cells co-expressed Pax-5 and Ki-67, suggesting FCRL4 cells were proliferative B-cells. The expression level of the FCRL4 was significantly higher in MALT lymphoma than LESA and SS. The detection rates of clonal IGH, IGK, and IGL gene rearrangements in MALT lymphoma with a sensitivity of 83.33%. Monoclonal immunoglobulin gene rearrangements were confirmed in five suspected patients by NGS (100%). ConclusionFCRL4 B cells might play an important role in the formation of lymphoepithelial lesions and might be as a diagnostic positive marker of salivary MALT lymphoma. The application of multiple detection methods could significantly improve the diagnostic accuracy for MALT lymphomas from LESA and SS.

Flow Cytometry Analysis Reveals a Wide Cytologic and Immunophenotypic Spectrum of Peripheral B Lymphocytes in Angioimmunoblastic T-Cell Lymphoma

Introduction: Angioimmunoblastic T-cell lymphoma (AITL) is an aggressive T-cell lymphoma commonly associated with B-cell dysregulation. Correlations involving B-cell dysregulation and clinicopathological features remain unclear. Methods: We prospectively collected blood samples from 11 AITL patients and 17 healthy controls. The percentages of B-cell subpopulations and lymphocytes with IL-21 production were assessed using flow cytometry. Peripheral blood lymphocyte morphology was evaluated microscopically. Results: Six of 11 (54.5%) patients presented with polyclonal hypergammaglobulinemia. Three of 11 (27.3%) tumor biopsies showed monoclonal immunoglobulin gene rearrangement. The patients exhibited significantly lower levels of naive (p < 0.001) and class-switched (p < 0.001) B cells than controls. The percentages of IgD−CD27− B cells (p = 0.007) and antibody-secreting cells (ASCs) (p = 0.001) were increased. Blood smears revealed atypical lymphocytes and immature plasma cells with morphological diversity. In comparison to normal controls, IL-21 production significantly increased in CD4+ (p < 0.001) and CD8+ (p = 0.020) T cells. B-cell clonality, RHOA G17V mutation, and the presence of sheets of clear cells and immature/mature plasma cells in lymph nodes were significantly associated with percentages of class-switched B cells and ASCs. The patients with circulating EBV DNA had a lower percentage of naive B cells (p = 0.009). Conclusions: Our results demonstrated a wide spectrum of peripheral B-cell morphologies and immunophenotypes of peripheral B cells in AITL. These findings correspond to dysregulated B-cell immunity and heterogeneous clinicopathological features.

The prognostic significance of monoclonal immunoglobulin gene rearrangement in conjunction with histologic B‐cell aggregates in the bone marrow of patients with diffuse large B‐cell lymphoma

Bone marrow involvement (BMI) is a well‐known poor prognostic factor in patients with diffuse large B‐cell lymphoma (DLBCL). This study robustly investigated the significance of monoclonal immunoglobulin gene rearrangement combined with histologic B‐cell aggregates in bone marrow (BM) in the detection of a poor prognostic group. Pretreatment BM samples of 394 DLBCL patients were analyzed via the immunoglobulin gene rearrangement study and the microscopic examination. Monoclonal immunoglobulin gene rearrangement was detected in 25.4% of cases. Histologic B‐cell aggregates with the features of large B‐cell lymphoma aggregates, small cell B‐cell lymphoma aggregates, or B‐cell aggregates of unknown biological potential were observed in 12% of cases (6.9%, 1.3%, and 3.8%, respectively). Histologic B‐cell aggregates were more associated with monoclonality than polyclonality. Cases with both monoclonality and histologic B‐cell aggregates demonstrated close association with poor prognostic factors such as a higher International Prognostic Index score and showed an inferior overall survival rate when compared to cases with only monoclonality or only histologic B‐cell aggregates. From the findings, a combination of monoclonality and histologic B‐cell aggregates within the bone marrow was highly associated with poor prognosis and could be used to determine high‐risk DLBLC patients with greater sensitivity and specificity than conventional microscopic examination or immunoglobulin gene rearrangement study alone.

Long-term Tumor-free Survival With Untreated Primary Diffuse Large B-cell Lymphoma of the Tonsil: A Report of 9 Cases.

In some cases, long-term tumor-free survival might be possible for untreated primary diffuse large B-cell lymphoma (DLBCL) of the tonsil. Here, we report of 9 untreated patients who had primary tonsil DLBCL with long-term tumor-free survival. All these patients were children or young adults (4 male and 5 female individuals; ages: 6 to 38 y, median age: 25 y) with clinically evident swollen tonsils or papillary neoplasms. Tonsillectomy and biopsies indicated partial structural destruction of the tonsils with diffuse infiltration of large lymphoid cells. The large cells expressed CD20 with a Ki-67 proliferative index >50%. All samples were negative for CD5, Epstein-Barr virus-encoded RNA, and t(14;18) translocation. Except for cases 2 and 4, all samples showed monoclonal immunoglobulin gene rearrangements. Although chemotherapy and radiotherapy had not been administered after tonsillectomy (for various reasons), periodic imaging and clinical evaluation showed that none of the 9 patients developed or died of lymphoma (median follow-up: 40 mo). In conclusion, primary tonsil DLBCL does not always behave in a malignant manner, and some patients can achieve long-term tumor-free survival without chemotherapy or radiotherapy. On the basis of this case series, we concluded that close follow-up and observation might be possible for pediatric and young-adult patients who have undergone complete tonsillectomy for primary tonsil DLBCL with the following features: short disease course, morphologically early lesions, negativity for Epstein-Barr virus-encoded RNA and t(14;18) translocation, and no involvement of any other sites, after careful clinical evaluation.

Cutaneous plasmacytosis: a clinicopathologic study of 12 cases in Taiwan revealing heterogeneous underlying causes.

Cutaneous plasmacytosis is clinically characterized by multiple pigmented papules and plaques that occur mainly on the trunk and many plasma cells in the lesional skin. Most of the cases reported have occurred in Japan. Whether cutaneous plasmacytosis is a unique reactive disease or a neoplastic condition has not been established. We collected 12 cases fulfilling the criteria for a diagnosis of cutaneous plasmacytosis at our institution from 2001 to 2013. We analyzed clinicopathologic features and performed immunohistochemical and immunoglobulin gene rearrangement studies. All 12 patients presented with characteristic cutaneous lesions mainly on the trunk and proximal extremities. Many plasma cells were observed in the lesional skin, and the basal layer was hyperpigmented. Four patients had numerous plasma cells in lymph nodes, and one patient also had increased plasma cells in bone marrow. One patient fulfilled the criteria for immunoglobulin G4 (IgG4)-related lymphadenopathy. Elevated serum IgG and IgG4 concentrations were found in seven patients. Monoclonal immunoglobulin gene rearrangements were detected in four patients. The median duration of follow-up was four years. One patient was lost from follow-up. Partial remission of the cutaneous lesions was observed in five patients, and progressive disease was noted in five patients. One patient died of respiratory failure. Our study suggests that the so-called "cutaneous plasmacytosis" has heterogeneous underlying causes with or without systemic involvement and may be associated with clonal immunoglobulin gene rearrangements and IgG4-related lymphadenopathy. No effective treatment is available for this condition.

Splenic lymphoma with villous lymphocytes, associated with type II cryoglobulinemia and HCV infection: a new entity?

AbstractHepatitis C virus (HCV) has been associated with the development of B-cell non-Hodgkin lymphomas. We recently reported the regression of splenic lymphoma with villous lymphocytes (SLVL) in patients with HCV after antiviral treatment, demonstrating a direct role of HCV in lymphomagenesis. This study expands our previous results in 18 patients with chronic HCV and SLVL. Mixed cryoglobulinemia (MC) was present in all cases and was symptomatic in 13 (72%). All patients were treated with interferon alone or in association with ribavirin. Hematologic and virologic responses were correlated. Fourteen (78%) patients achieved a sustained complete hematologic response after clearance of HCV RNA. Two patients had a virologic partial response and achieved a complete hematologic response. Two virologic nonresponders achieved partial hematologic response. Regardless of the response, monoclonal immunoglobulin gene rearrangement persisted after treatment. This study underscores the role of HCV in the lymphomagenesis and the benefit of antiviral treatment for patients presenting with HCV-driven lymphoproliferations.

Demonstration of Light Chain Restricted Clonal B-Lymphoid Infiltrates in Archival Bone Marrow Trephines by Quantitative Real-Time Polymerase Chain Reaction

Assessment of clonality either by demonstrating light chain restriction or showing monoclonal immunoglobulin gene rearrangement is a valuable and indispensable adjunct to diagnosis in hematopathology. The study of light chain restriction by immunohistochemistry on archival material is hampered by a very low sensitivity especially regarding low grade lymphomas of B cell origin. DNA rearrangement studies of the immunoglobulin locus do improve sensitivity markedly but for lymphomas of follicle center origin they are prone to false negative results due to hypermutations. Therefore we developed a new clonality assay based on the quantification of immunoglobulin light chain transcripts using real-time polymerase chain reaction technology, which is also suitable for the analysis of archival bone marrow trephines. We tested the reproducibility and sensitivity of this approach by comparatively analyzing a series of bone marrow trephines with multiple myeloma (n = 26), reactive lymphoid hyperplasia (n = 37), and focal infiltration by low grade B cell lymphoma (n = 29). We could raise the detection rate of clonality from an average of 17% by immunohistochemistry and 66% as assessed by polymerase chain reaction rearrangement studies to 83% by this new technique. Despite false negative results due to light chain hypermutation in some cases, the detection rate of clonality could be improved even for B cell lymphomas of follicle center origin (follicular lymphoma or marginal zone lymphoma) thus making this novel approach a valuable additional tool for the hematopathologist.

Study on the role of G1 cyclins in Epstein-Barr virus-associated human lymphomas maintained in severe combined immune deficiency (SCID) mice

Five Epstein-Barr virus (EBV)-positive human lymphoma cell lines maintained in severe combined immune deficiency (SCID) mice were used to investigate the role of G1 cyclins in EBV-induced lymphomagenesis. All the primary tumors had been negative for EBV but became positive after establishment in SCID mice, with monoclonal immunoglobulin gene rearrangement and EBV monoclonality. To compare the expression status of G1 cyclins, these EBV-associated lymphoma lines (6 EBV[-] human SCID mouse lymphoma lines, 13 human B cell lymphomas and 8 samples of human tonsil tissue) were examined by reverse transcription-polymerase chain reaction-Southern blotting, Western blotting and immunohistochemistry. mRNA expression of cyclin D1 (CCND1), cyclin D2 (CCND2), cyclin E (CCNE), cyclin-dependent kinase 2 (CDK2) and 4 (CDK4) was found in all 3 types of lymphomas. Western blotting demonstrated identical results. Immunohistochemistry revealed CCND1 to be negative in all lymphomas. CCND2 was positive and restricted to the nuclei in all EBV(+) SCID mouse lymphoma lines, whereas it was limited to the cytoplasm in half of the EBV(-) counterparts. CCNE was positive in the nuclei in all EBV(+) but negative in all EBV(-) SCID mouse lymphoma lines. Immunoprecipitation of EBV(+) and (-) SCID mouse lymphomas for CCND1, CCND2 and CCNE vs. p21, PCNA and CDK2 or CDK4 demonstrated that, in EBV(+) SCID lines, CCND2/CDK4 complexes were present without binding to p21, suggesting independence from p21 regulation. In EBV(-) SCID mouse lymphomas, half of the cases showed complex formation of CCND2/CDK4 without binding of p21. In contrast, CCND1/CDK4 and CCNE/CDK2 were under regulation of p21 in both EBV(+) and (-) lymphomas. These results suggest that differential expression of CCNDs, CCNE and CDKs, as well as variation in their subcellular localization and association with CDK-inhibitor protein, could explain differences in cell proliferation between EBV(+) and EBV(-) lymphomas.

Molecular and virologic characteristics of lymphoid malignancies in children with AIDS.

To characterize AIDS-associated lymphoid malignancies in children. We studied lymphomas and B-cell leukemias from 25 children with AIDS for immunoglobulin heavy chain gene clonality, c-myc oncogene abnormalities, and presence of HIV and Epstein-Barr virus. Monoclonal immunoglobulin gene rearrangements were identified in 22 of 23 cases tested, the single exception being one of mucosa-associated lymphoid tissue. Immunoglobulin gene/c-myc translocations were found in 3 of 4 cases of B (surface immunoglobulin-positive)-acute lymphoblastic leukemia, 8 of 11 small noncleaved cell lymphomas, and 1 of 5 large cell lymphomas. Mutations of c-myc were found in 2 of 13 small noncleaved cell lymphomas, 1 of 2 Epstein-Barr virus-positive mucosa-associated lymphoid tissue neoplasms, and 1 of 4 Epstein-Barr virus-negative B-acute lymphoblastic leukemia. Six small noncleaved cell lymphomas, both mucosa-associated lymphoid tissue neoplasms and one of large cell lymphoma had high levels of Epstein-Barr virus in tumor tissue. Hodgkin's disease tissue and B-acute lymphoblastic leukemia tumors were negative for EBV. Proviral HIV-1 was not detected in any tumor. AIDS-associated lymphoid malignancies in children appear to have a different distribution of histologic subtypes than adult HIV-infected individuals, fewer large cell lymphomas occur in children. The small noncleaved cell lymphomas exhibit a lower frequency as well as different locations of c-myc mutations than AIDS-associated small noncleaved cell lymphomas in adults.

Detection of clonal excess in lymphoproliferative disease by kappa/lambda analysis: correlation with immunoglobulin gene DNA rearrangement

Previous studies have suggested that analysis of the distribution of surface immunoglobulin light chain isotypes by flow cytometry provides evidence for monoclonality of B cell tumors and may detect populations of circulating tumor cells in patients with lymphoproliferative disease. We have used simultaneous flow cytometry and DNA restriction enzyme analysis on 58 samples of tissue and blood to determine whether lymphocyte populations detected by "kappa/lambda" analysis are indeed monoclonal. In greater than 90% of cases, abnormalities detected by flow cytometry correlated with monoclonal rearrangements of immunoglobulin genes as detected by Southern blot analysis. By analyzing tissue and blood from the same patients, we have also demonstrated that monoclonal circulating cells detected by flow cytometry reflect peripheral circulating tumor cells, since DNA from these cells shows the same immunoglobulin rearrangement as DNA from the original tumors in these patients. Although mixing studies suggested that DNA rearrangement studies were more sensitive than was flow cytometry in detecting minor populations of monoclonal lymphocytes, we found only one case in which this affected the diagnostic accuracy of the kappa/lambda analysis, with one notable exception, that of detection of a monoclonal proliferation of B cells that did not express surface immunoglobulin. The kappa/lambda test thus offers a powerful diagnostic tool in the evaluation of lymphoproliferative disease.

Detection of Clonal Excess in Lymphoproliferative Disease by K/λ Analysis: Correlation With Immunoglobulin Gene DNA Rearrangement

Abstract Previous studies have suggested that analysis of the distribution of surface immunoglobulin light chain isotypes by flow cytometry provides evidence for monoclonality of B cell tumors and may detect populations of circulating tumor cells in patients with lymphoproliferative disease. We have used simultaneous flow cytometry and DNA restriction enzyme analysis on 58 samples of tissue and blood to determine whether lymphocyte populations detected by “kappa/lambda” analysis are indeed monoclonal. In greater than 90% of cases, abnormalities detected by flow cytometry correlated with monoclonal rearrangements of immunoglobulin genes as detected by Southern blot analysis. By analyzing tissue and blood from the same patients, we have also demonstrated that monoclonal circulating cells detected by flow cytometry reflect peripheral circulating tumor cells, since DNA from these cells shows the same immunoglobulin rearrangement as DNA from the original tumors in these patients. Although mixing studies suggested that DNA rearrangement studies were more sensitive than was flow cytometry in detecting minor populations of monoclonal lymphocytes, we found only one case in which this affected the diagnostic accuracy of the kappa/lambda analysis, with one notable exception, that of detection of a monoclonal proliferation of B cells that did not express surface immunoglobulin. The kappa/lambda test thus offers a powerful diagnostic tool in the evaluation of lymphoproliferative disease.