Clonal Immunoglobulin Heavy Chain Gene Rearrangement Research Articles

Immunoglobulin and T Cell Receptor β Chain Gene Rearrangement Analysis of Ocular Adnexal Lymphoid Neoplasms: Clinical and Biologic Implications

We investigated the organization of the immunoglobulin heavy chain (IgH) and the T cell receptor beta chain (T beta) gene loci in 20 ocular adnexal and four extraocular lymphoid neoplasms obtained from 18 patients presenting with an ocular adnexal lymphoid neoplasm. Fifteen ocular adnexal and four extraocular lymphoid neoplasms occurring in 13 patients were classified by morphological examination and immunophenotypic analysis as monoclonal B cell lymphomas. Each one of these 19 lymphoid neoplasms exhibited clonal IgH gene rearrangements upon hybridization of EcoRI- or HindIII-digested DNA to a heavy-chain joining region (JH)-specific DNA probe. The bilateral ocular adnexal monoclonal B cell neoplasms occurring simultaneously in two individuals exhibited identical clonal IgH gene rearrangements, which indicated their derivation from an identical B cell clone. The ocular adnexal and the extraocular monoclonal B cell neoplasms occurring in two of three patients also exhibited identical clonal IgH gene rearrangements, which suggested that they too were derived from an identical B cell clone. Five ocular adnexal lymphoid neoplasms were classified by morphological examination and immunophenotypic analysis as benign, polyclonal pseudolymphomas. Three of these five ocular adnexal lymphoid neoplasms exhibited clonal IgH gene rearrangements, which suggested the presence of monoclonal B cell populations that escaped detection by morphological and immunophenotypic examination. None of the 24 pathological samples exhibited clonal T beta gene rearrangements upon hybridization of EcoRI- or BamHI-digested DNA to a T beta gene DNA probe. The results of these studies demonstrate the value of Southern blot hybridization analysis for clonal IgH and T beta gene rearrangements in the diagnosis, classification, and investigation of extranodal lymphoid neoplasms originating and/or presenting in the ocular adnexa.

Prognostic markers for immunodeficiency-associated primary central nervous system lymphoma

Immunodeficiency is a major risk factor for primary central nervous system lymphoma (PCNSL), but data on the disease in immunocompromised hosts are scarce. We aimed to define clinical and imaging features and determine prognostic factors for immunodeficiency-associated PCNSL. All PCNSL cases seen at Yale-New Haven Hospital between 2002 and 2017 were retrospectively screened for immunodeficiency. For patients with immunosuppression, biopsies were evaluated and clinical data were collected. Predictors of survival were identified using Kaplan-Meier survival analysis and log-rank test. p values < 0.05 were considered significant. 23 patients with immunodeficiencies were identified: eleven on immunosuppressants after solid organ transplantation, seven with human immunodeficiency virus infection, and five on immunosuppressive treatment due to various autoimmune disorders. PCNSL cases were largely Epstein-Barr-Virus positive (78%), histologically classified as diffuse large B cell lymphomas (87%), and showed peripheral contrast enhancement (81%) and corresponding heterogeneous diffusion-weighted imaging patterns (DWI) on magnetic resonance imaging (MRI) (71%). Median overall survival was 31months. Age > 60years at diagnosis (p < 0.01), peripheral enhancement of the mass on MRI (p = 0.04), heterogeneous DWI patterns (p = 0.04), and clonal immunoglobulin heavy chain gene rearrangement (IgHR) (p = 0.03) were found to be negative prognostic markers. Immunodeficiency-associated PCNSL presents with similar clinical, pathological and imaging features. Age > 60years, clonal IgHR, heterogeneous DWI pattern and peripheral enhancement on MRI may serve as predictors of less favorable outcome.

Rapid detection of immunoglobulin heavy chain gene rearrangement by PCR and melting curve analysis using combined FR2 and FR3 primers

BackgroundImmunoglobulin heavy chain (IgH) gene rearrangement test is a standard tool in diagnosing B-cell lymphoma. The BIOMED-2 multiplex PCR protocol has become the most commonly used laboratory method for detecting clonal IgH gene rearrangement. However, post-PCR procedure requires manual transfer of PCR product for analysis and is time-consuming. A novel strategy using LightCycler to continuously monitor fluorescence during melting curve analysis (MCA) can overcome these shortcomings. The previous studies published on this method were all restricted to FR3 primers of BIOMED-2.MethodsReal-time PCR and subsequent MCA were performed on 71 clinical DNA samples from formalin-fixed, paraffin-embedded tissues, including 40 with B-cell non-Hodgkin lymphomas and 31 with reactive lymphoid hyperplasia. We optimized the current method using FR3 primers and applied FR2 primers for the first time into MCA to detect IgH gene rearrangement. Polyacrylamide gel electrophoresis and capillary gel electrophoresis were also performed on all lymphoma samples with the identical FR2 primers.ResultsMCA of combined FR2 and FR3 primer sets yielded the sensitivity and the specificity equal to 70 % (28/40) and 100 % (31/31), respectively. Addition of FR2 primers increased the sensitivity by 12.5 % (5/40) comparing to FR3 primers alone. MCA was slightly more sensitive than polyacrylamide gel electrophoresis and comparable to capillary gel electrophoresis to detect clonal IgH gene rearrangement.ConclusionsCombined PCR and DNA melting curve analysis in a closed system can reduce cross-contamination risk. This method can test 96 samples simultaneously within 90 min and therefore, it is high-throughput and faster. PCR-MCA in the LightCycler system has potential for evaluating monoclonal IgH gene rearrangement in a clinical environment.

Florid Reactive Lymphoid Hyperplasia of the Lower Female Genital Tract (Lymphoma-like Lesion): A Benign Condition That Frequently Harbors Clonal Immunoglobulin Heavy Chain Gene Rearrangements

Lymphoma-like lesions (LLL) of the lower female genital tract are florid reactive inflammatory processes that mainly occur in women in their reproductive years. Histologically, they are characterized by a dense lymphoid infiltrate with admixed large cells that is often suspicious for lymphoma. In contrast to lymphoma, however, they are superficial lesions that typically show surface erosion and a mixed lymphoid infiltrate and do not have evidence of a mass, deep invasion, or prominent sclerosis. With the advent of widespread molecular genetic testing, it would seem that LLLs should be polyclonal helping make the correct diagnosis. However, we have found cases with morphologic and immunophenotypic features of LLLs and evidence of clonal rearrangement of the immunoglobulin heavy chain (IGH) gene, potentially leading to misdiagnosis. We examined the clinicopathologic features and outcome of 12 patients with LLL (9 in the cervix and 3 in the endometrium). The patients ranged in age from 18 to 54 (median 37) years and came to medical attention because of squamous dysplasia (8 patients), vaginal bleeding (3), or adnexal mass (1). One patient had an endocervical polyp, but otherwise none had a discrete mass. The specimens contained a dense, polymorphous inflammatory infiltrate, commonly associated with mucosal erosion. Immunohistochemical studies showed a mixture of B and T cells without immunoglobulin light chain restriction. Four cases (all cervical) had a clonal IGH gene rearrangement by PCR. There was no evidence of lymphoma on staging or on follow-up in any patient, including the 4 patients with clonal IGH rearrangement, after a mean follow-up of 3.5 years (range: 4 mo to 13 y). In summary, we describe 12 patients with LLL of the lower female genital tract. Four of the 9 cases (44%) analyzed by PCR had a clonal IGH gene rearrangement. The clinical and pathologic features of these cases suggest that a clonal IGH rearrangement in this setting does not warrant a diagnosis of lymphoma. Careful correlation of clinical, histologic, immunophenotypic, and genetic features is required to avoid misdiagnosis and inappropriate treatment. Routine microscopic findings and detailed clinical information remain paramount in establishing the correct diagnosis.

Clinical significance in detection of immunoglobulin heavy chain clonal rearrangement in bone marrow of patients with B cell lymphoma

To explore the feasibility of semi-nested PCR technique for detection of immunoglobulin heavy chain (IgH) clonal rearrangement in bone marrow of B-cell lymphoma patient and to further evaluate its clinicopathological value. Gene clonal rearrangement of IgH was detected by semi-nested PCR using primers of FR2 & FR3A in 105 bone marrow samples of patients with B-cell lymphoma. The PCR detection results were compared with the cytomorphology of bone marrow aspiration biopsy. The correlation between PCR detection results and clinicopathological factors were evaluated. Among 105 cases of B-cell lymphoma, bone marrow involvement was detected by PCR technique in 48 cases (45.7%), while only 22 cases (21.0%) were detected by bone marrow cytological analysis. There was a significant difference between two methods (P < 0.05), and the concordance rate was 71.4%. The incidence of bone marrow involvement at the time of initial diagnosis detected by PCR technique was 30.8% for diffuse large B cell lymphoma (DLBCL), 25.0% for follicular lymphoma (FL), and 100.0% for small lymphocytic lymphoma (SLL), respectively. Bone marrow involvement detected by PCR detection correlated with Ann Arbor stage. Rate of clonal IgH gene rearrangement by PCR in early B-cell lymphoma was lower than that in advanced stage B-cell lymphoma patients (P = 0.02). There was no statistically significant difference in efficacy between patients with positive and negative results detected by PCR (P > 0.05). But difference in complete response (CR) rate (23.3% and 46.3%) had significant difference (P = 0.019). Semi-nested PCR analysis may be an effective method for detection of abnormalities in bone marrow in patients with B-cell lymphoma and is superior to cytomorphology. The positive rate in patients with advanced Ann Arbor stage is higher than that in patients with early Ann Arbor stage, and patients with PCR negative result have more chances to achieved CR after treatment.

Primary Hepatic Lymphoma of Mucosa-Associated Lymphoid Tissue Type: A Case Report With Cytogenetic Study

Primary hepatic lymphoma of mucosa-associated lymphoid tissue type is extremely rare. Only 38 cases have been reported to date. A case of a 59-year-old man with Helicobacter pylori-resistant gastric ulcers and Buerger disease who was followed up since 1999 is reported. A 2-cm hepatic nodule was incidentally found during partial gastrectomy and corresponded to mucosa-associated lymphoid tissue-type lymphoma without underlying liver disease. Molecular studies showed a clonal immunoglobulin heavy-chain gene rearrangement. Investigations for the mucosa-associated lymphoid tissue lymphoma-associated translocations t(11;18) and t(14;18), as well as the t(3;14)(q27;q32), were negative, whereas trisomy 3 and trisomy 18 were detected.

Bone marrow trephines containing lymphoid aggregates from patients with rheumatoid and other autoimmune disorders frequently show clonal B-cell infiltrates

In bone marrow trephines, morphological and immunohistochemical criteria may not be sufficient to discriminate reactive from malignant lymphoid infiltrates. The aim of this study was to determine whether the detection of clonal immunoglobulin heavy chain (IGH) gene rearrangements is a reliable and specific marker for malignant B-cell clones in bone marrow biopsies. Bone marrow trephines with infiltration by different types of low-grade B-cell non-Hodgkin lymphoma (n = 32), reactive lymphoid hyperplasia (n = 18), and reactive lymphoid aggregates (n = 15), including 5 patients with rheumatoid or other autoimmune disorders, were analyzed by morphology, immunohistochemistry, IGH gene rearrangement (polymerase chain reaction), and DNA sequence analysis in selected cases. In 22 (68.8%) of 32 patients with B-cell non-Hodgkin lymphoma, a clonal IGH gene rearrangement was detected. Of the reactive cases, 1 of 18 patients with lymphoid hyperplasia demonstrated clonality, and 9 (60%) of 15 patients with reactive lymphoid aggregates gave a clonal result (GeneScan analysis). DNA sequence analysis was performed in 7 of the latter patients confirming clonality in 6. Four of the patients with B-cell clonality had an autoimmune disorder. None of these patients developed a malignant lymphoma during follow-up. Thus, the molecular detection of a clonal rearrangement of the IGH gene may support the diagnosis of a malignant lymphoma infiltrating the bone marrow. However, morphologically and immunohistochemically benign lymphoid aggregates might also harbor B-cell clones especially in patients with autoimmune disorders. Therefore, the detection of clonality has to be interpreted with utmost care and does not qualify for the unequivocal diagnosis of a malignant B-cell lymphoma.

Distinction between lymphoma-like lesions and lymphoma of uterine cervix: a clinicopathologic study of 26 cases

To study the clinicopathologic features and differential diagnosis of lymphoma-like lesions and lymphoma of uterine cervix. Clinical data and hematoxylin and eosin-stained slides of 10 cases of lymphoma-like lesion and 16 cases of lymphoma of uterine cervix were reviewed. Immunohistochemical study for B- and T-cell markers and light chains (kappa, lambda) were performed on paraffin sections. The rearrangement status of immunoglobulin heavy chain (IgH) gene was analyzed with semi-nested polymerase chain reaction in 4 cases lymphoma-like lesion and 4 cases of lymphoma of uterine cervix. The age of patients with lymphoma-like lesion ranged from 24 to 54 years (medium = 43 years). The lesion generally presented with cervical erosion or polyp. Microscopically, it is characterized by focal or diffuse superficial infiltration of immunoblast-like large B cells intermingled with a polymorphic population of inflammatory cells, including plasma cells, eosinophils and neutrophils. Maturation of the transformed large B cells was also noticed. On the other hand, the age of the patients with lymphoma of uterine cervix varied from 28 to 78 years (medium = 58 years). Cervical mass or diffuse enlargement of cervix were the commonest clinical findings. The cases included 12 examples of diffuse large B-cell lymphoma and 4 examples of follicular lymphoma. The former was characterized by a diffuse monomorphic population of large atypical lymphoid cells, while neoplastic follicles were identified in the latter. Neither polymorphic inflammatory infiltrates nor maturation phenomenon was found. The immunostaining for kappa and lambda light chains was inconclusive. Molecular study showed clonal rearrangement of IgH gene in all cases of cervical lymphoma, as well as 2 cases of lymphoma-like lesion. The distinction between lymphoma-like lesion and lymphoma of uterine cervix depends primarily on the clinical and histopathologic features. Assay for rearrangement of IgH gene may be helpful in differential diagnosis, though monoclonality can be detected in some benign lesions as well.

Eradication of Minimal Residual Disease during Treatment of Mantle Cell Lymphoma: CALGB 59909.

Recent studies suggest that detection of subclinical, or minimal residual disease (MRD) in apheresis products used for autografting correlates with poor disease-free survival following ASCT for mantle cell lymphoma (MCL). To validate this observation and to gain insights into the kinetics of MRD during treatment, we are performing a prospective analysis of MRD using quantitative real-time PCR (Q-PCR) in patients (pts) undergoing treatment for MCL on a CALGB study (59909). Q-PCR of sequential paired bone marrow (BM) and blood (B) samples and of apheresis products was performed using either a patient-specific immunoglobulin heavy chain (IgH) or BCL-1 gene rearrangement. All samples were analyzed in triplicate using LightCycler technology and reported as a normalized ratio of IgH or BCL-1 copy number to GAPDH copy number. The sensitivity of the assay ranged from 1 X 104–1 X 105. To date, a clonal IgH or BCL-1 gene rearrangement was detected in 36 of 41 (88%) pts entered on study. Patient-specific primers and consensus probes were used for Q-PCR monitoring of MRD following two courses of intensive induction therapy, during stem cell mobilization, and 3 and 12 months after ASCT and post-transplant immunotherapy with Rituximab (R). 27 pts have completed all protocol treatment with a median follow-up of 7 months (range: 0–28). 26 pts were evaluated for MRD following two courses of induction therapy with cyclophosphamide, methotrexate, doxorubicin, vincristine, prednisone, and R. 10 of 26 became MRD negative (−) following induction while 16 remained MRD positive (+). Following mobilization with high-dose cytarabine, etoposide and R, apheresis products from 9 of 10 MRD- pts were evaluated and all products were MRD- (1 pt not evaluable). Of the 16 pts who were MRD+ prior to mobilization therapy, MRD- apheresis products were collected in 8; 5 had MRD+ stem cell collections, and 3 were not evaluable. In total, apheresis products were evaluable in 22 of 26 pts; 17 (77%) had MRD- stem cells collected prior to ASCT. None of these MRD- pts has relapsed to date although 2 pts with MRD- products became weakly MRD+ 12 months following ASCT and R. Of the 5 pts with MRD+ apheresis collections, 4 have remained persistently MRD+ following ASCT and R; 1 has relapsed 19 months after completion of all treatment. Rising levels of MRD in BM and B samples were noted in this patient 3 and 12 months post-ASCT. Statistical comparison of MRD values in paired BM and B samples prior to, and post-ASCT demonstrated good agreement with an intraclass correlation coefficient of.814 and.777, respectively. In conclusion, our results demonstrate that prospective MRD monitoring using Q-PCR provides important insights into the kinetics of response during treatment of MCL. MRD- apheresis products were collected in the majority of pts following intensive induction and mobilization chemo-immunotherapy with R on CALGB 59909. To date, no pts with MRD- apheresis products have relapsed following ASCT. In contrast, it appears that pts with MRD+ apheresis products remain MRD+ following ASCT and R and may be more likely to relapse. Longer clinical follow-up is needed to clarify the significance of the persistence of MRD in apheresis products and following ASCT for MCL.

Immunoglobulin Gene Rearrangement of Lymphoid Malignancies: Application of the PCR Technique to Formalin-Fixed, Paraffin-Embedded Tissues

Every B-lymphocyte carries a unique sequence of the immunoglobulin heavy chain (IgH) gene, which serves as a marker for tumor cells derived from clonal B-lymphocyte expansion. Southern blot analysis for IgH gene rearrangement is not suitable for small biopsies and formalin-fixed, paraffin-embedded tissues. We aimed to assess the application of PCR techniques to demonstrate clonal IgH gene rearrangement in B-cell non-Hodgkin's lymphoma (B-NHL). A series including 119 B-NHL, 10 T-cell NHL, 6 reactive lymphoid hyperplasia, and 15 non-lymphoid malignancies were examined. DNA was extracted from paraffin-embedded tissues by proteinase K digestion. A semi-nested PCR amplification targeting two framework regions (Fr2A and Fr3A) and the joining region (JH) of the variable gene segment of the IgH gene was performed. Clonal IgH gene rearrangement was detected in 30% and 63% of B-NHL with Fr2A and Fr3A assays, respectively, and 74.8% when both assays were combined. Clonal IgH gene rearrangement was not demonstrated in any of the reactive lymphoid hyperplasia and non-lymphoid malignancies. However, 20.0% of the T-NHL showed clonal IgH gene rearrangement. Differences were noted among different subtypes of B-NHL. Clonal IgH gene rearrangement was detected in all of the low-grade B-NHL, except follicular lymphoma. High-grade B-NHL generated heterogeneous results.

Primary plasmacytoma arising in an endocervical polyp with detection of neoplastic cells on papanicolaou test. A case report and review of the literature.

Primary plasmacytomas of the female genital tract are extremely rare and present a substantial diagnostic challenge. Five cases morphologically representing plasmacytomas and localized to the uterine cervix have been reported previously; however, only 1 was shown to be monotypic for immunoglobulin light-chain expression. We report the case of a 37-year-old woman who had highly atypical plasma cells on her Papanicolaou test. A clinically detected endocervical polyp was removed and revealed a plasmacytoma, the diagnosis of which was substantiated by demonstrating monotypic lambda-light-chain restriction and a clonal immunoglobulin heavy-chain gene rearrangement. The cytologic and histopathologic findings of plasmacytomas of the uterine cervix are discussed, including the utility of immunophenotypic and molecular techniques to confirm the neoplastic diagnosis.

The application of a pCR technique for the detection of immunoglobulin heavy chain gene rearrangements in fresh or paraffin-embedded skin tissue

Although detection of a clonal sequence of the heavy chain gene of immunoglobulin by the polymerase chain reaction (PCR) is frequently used to assess lymphoid infiltrates in skin biopsy specimens, there are no data on the sensitivity and specificity of this test in detecting clonal B cell populations. Having refined a PCR technique for the detection of immunoglobulin heavy chain (IgH) gene rearrangement in both fresh and formalin-fixed, paraffin-embedded skin samples, we undertook to define the role of this assay in the diagnostic setting. Thirty-one cases of cutaneous B cell lymphoma (CBCL), 19 cases of B cell pseudolymphoma (lymphocytoma cutis), 34 cases of benign lymphocytic infiltrates of the skin and one case of cutaneous T cell lymphoma (CTCL) were studied using the polymerase chain reaction assay. All biopsies were formalin-fixed, paraffin-embedded skin sections apart from 13 of the 31 CBCL specimens which were fresh skin specimens. DNA from the framework region 3 (FR3) sequence of the IgH genes was amplified to ascertain the presence of a clonal IgH gene rearrangement. The findings were correlated with histological and immunophenotyping results on all samples. The assay performed with 73% sensitivity and 100% specificity, comparable to results obtained examining fresh lymphoid tissue specimens from patients with B cell tumours. The results indicate that this technique is a useful tool in the work up of suspected CBCL and in differentiating between CBCL and mixed lymphocytic infiltrates, a clearly important distinction with regards to prognosis and treatment.

PCR Analysis of Immunoglobulin Heavy Chain (IgH) and TcR-γ Chain Gene Rearrangements in the Diagnosis of Lymphoproliferative Disorders: Results of a Study of 525 Cases

This report summarizes a cumulative 4-year experience in polymerase chain reaction (PCR) analysis of immunoglobin heavy chain (IgH) and TcR-γ chain gene rearrangements in 525 cases of lymphoproliferative disorders. Because the sensitivity of the PCR methodology was found to be tissue dependent, in the study of the presence of clonal cell population in tissues containing a small number of polyclonal lymphocytes, such as skin and gastrointestinal biopsy specimens, we used the multiple–PCR run approach. In this latter methodology, we repeat the PCR reaction from the same sample at least three times to confirm the reproducibility of the results. In the study of 273 cases of B- or T-cell lymphomas with characteristic immunomorphological and clinical features, a clonal IgH or TcR-γ chain gene rearrangement was detected in approximately 80% of cases. A clonal rearrangement involving both IgH and TcR-γ chain genes was found in 10% of cases of both B-cell and T-cell lymphomas. The study of 167 cases of nonneoplastic lymphoid tissue samples showed the presence of clonally rearranged cell populations for IgH or TcR-γ genes in 3 and 9% of cases, respectively. We also applied PCR for the study of 85 cases of lymphoproliferations with no definite diagnosis (i.e., benign versus malignant) after immunomorphological analysis. In 65 cases (76%), the correlation of immunomorphological features with the presence (48 cases) or the absence (17 cases) of clonal lymphoid cell populations led to a definite diagnosis. In almost all these cases, the final diagnosis was found to be in agreement with the clinical course. In the 20 remaining cases (24%), no definite diagnosis could be made. We also assessed the value of PCR in detecting bcl-2/JH gene rearrangement as an additional clonal marker in the diagnosis of follicular lymphoma. Bcl-2/JH rearrangement and/or IgH gene rearrangement was found in approximately 85% (71/85) of follicular lymphoma cases studied.

Frequent T and B Cell Oligoclones in Histologically and Immunophenotypically Characterized Angioimmunoblastic Lymphadenopathy

The identification of clonal rearrangements of T cell receptor (TCR) genes is central to the diagnosis of T cell lymphomas. However, in angioimmunoblastic lymphadenopathy (AILD), first described as a nonneoplastic proliferation associated with immunodeficiency, the heterogeneity of TCR and IgH gene rearrangements suggest that some cases may harbor multiple lymphoid clones. In this study we have isolated DNA from archival paraffin biopsy material from 22 cases of AILD identified on the basis of classical histological and immunohistochemical features with the aim of establishing the occurrence of clones and oligoclones, the frequency of TCR and immunoglobulin heavy chain (IgH) variable (v) gene use, and the relationship of these findings to the presence of Epstein-Barr virus. DNA extracted from the biopsies was amplified using the polymerase chain reaction (PCR) and sequenced to detect functional and nonfunctional gene rearrangements. Epstein-Barr virus-encoded short RNA species (EBERs) were detected using in situ hybridization combined with immunochemistry to identify the phenotype of the Epstein-Barr virus-infected cells. Fifty-seven clonal products were found in 20/22 patients: TCRgamma clonal products were identified in 16/22, TCRbeta clonal products in 16/22 and IgH clonal products in 6/22 cases. Oligoclonal PCR products were seen for TCR in 3/22 and for IgH in 3/22 cases. In one biopsy PCR products from all reactions were polyclonal. Sequence analysis revealed functional TCRgamma, TCRbeta, and IgH sequences in 6/12, 9/11, and 8/8 cases, respectively. Functional TCR and/or IgH oligoclones were detected in 6/20 (30%) cases. In addition, nonfunctional TCR and IgH sequences were found in 11 cases. EBERs were identified in 18/20 cases varying from occasional to 25 to 30% nuclei staining and were associated with both T and B cells, although the majority were of indeterminate phenotype. The presence of EBERs was not associated with all clonal IgH gene rearrangements but was associated with B cell oligoclones. Patterns of gene recombinations indicated that the majority of TCRgamma recombinations used GV1 and GJ1S3/2S3 genes. Six out of eleven cases used TCR BV4S1 or BV2S1 genes associated with various BJ and BD1/2 genes. No common IgH gene usage was identified, but 8 clones had varying degrees of replacement and silent mutations (0.6-10.1%), consistent with B cell clones having undergone somatic mutation in the germinal center, and 3 clones harbored unmutated V genes, consistent with naive B cells. Our data do not support the concept of AILD as a clearly defined peripheral T cell lymphoma (PTCL). Rather, they suggest that AILD as defined by histology and immunohistochemistry is either a heterogeneous entity or represents a lymphoproliferation associated with immunodeficiency in which clonal T cell or B cell proliferation may occur.

Prognostic Significance of Immunoglobulin and T Cell Receptor Gene Rearrangements in Patients with Acute Myeloid Leukemia: Taiwan Experience

We investigated the prognostic significance of lymphoid antigen receptor gene rearrangement in patients with newly diagnosed acute myeloid leukemia (AML). Thirty-nine patients were included in the study. Clonal gene rearrangement of immunoglobulin heavy chain (IgH) and T cell receptor β chain (TCRβ) was found in leukemic cells in 11 (28.2%) and 10 (25.6%) patients, respectively. Five (12.8%) had both IgH and TCRβ gene rearrangements. Three of the seven (42.9%) B-lymphoid marker-positive and eight of the 32 (25%) B-lymphoid marker-negative patients had clonal IgH gene rearrangements. Five of the 11 (45.5%) T-lym-phoid marker-positive and 5 of the 28 (17.9%) T-lymphoid marker-negative patients had clonal TCRβ gene rearrangements. All patients were treated with similar regimens. The complete remission rate (62.5% vs 65.296, p=1.000) and median survival (13 vs 14 months, p=4.366) were similar in patients with and without clonal IgH or TCRβ gene rearrangements. In conclusion, while clonal rearrangements of IgH or TCRβ genes were found in AML patients, they did not appear to effect the prognosis.

Pseudolymphoma (Reactive Lymphoid Hyperplasia) of the Liver Containing Epithelioid Cell Granulomas and Schaumann's Bodies in Giant Cells

Nodular lymphoid hyperplasia, also known as pseudolymphoma (PL), of the liver is a benign reactive process of lymphoid tissue and is extremely rare. To our knowledge, only four cases of hepatic PL have been reported to date. We present a 59-year-old woman patient with hepatic PL that was preoperatively diagnosed as hepatocellular carcinoma (HCC), based on the findings of a variety of imaging modalities. However, resection of the tumor specimen at hepatic lobectomy revealed that the tumor was not a true tumor but a PL composed of proliferated nonneoplastic, polyclonal lymphocytes and plasma cells with abundant lymphoid follicles, together with mononuclear and multinuclear histiocytes with granuloma formation. Sporadic multinucleated giant cells had concentrically or eccentrically laminate ‘conchoidal’ bodies, known as Schaumann's bodies. Immunohistochemical staining of lymphocytes and plasma cells for T and B cells, or IgGK and IgGk revealed the polyclonal nature of these cells. Furthermore, clonal immunoglobulin heavy-chain gene rearrangement was not detected in the lymphoid cells in the lesion with polymerase chain reaction (PCR) analysis. The background hepatic tissue showed no cirrhosis, but some lymphocytes had infiltrated into the portal tracts around the tumor. Since hepatic PL is extremely rare and no association of PL with granuloma with Schaumann's bodies has been described in previously reported cases with hepatic PL, the clinical and histologic features in this case are described with a literature review.

Prognostic significance of immunoglobulin heavy chain gene rearrangement in patients with acute myelogenous leukemia.

Recently the immunoglobulin heavy chain (IgH) gene rearrangement in B cell malignancies has been analyzed. Clonality can be determined using the polymerase chain reaction (PCR). Little attention, however, has been given to the relationship between prognosis and IgH gene rearrangement in patients with acute myelogenous leukemia (AML). In this study, we examined IgH gene rearrangement in 35 untreated AML patients by PCR. PCR was performed using consensus heavy chain complimentarity-determining region (CDR)-3 primers. Clonal IgH gene rearrangement was detected in 14 patients (40%). Four of five patients (80%) who were positive for B cell markers had clonal IgH gene rearrangement. Ten of 30 B cell antigen-negative patients (33%) also showed IgH rearrangement. All patients were treated with a daunorubicin-based regimen, resulting in complete remission for 29 patients (83%). Sixty-four percent of those with IgH rearrangement and 95% of those without rearrangement had complete remission. Overall survival of IgH-PCR positive and negative patients at 25 months was 29 and 88%, respectively. IgH-PCR positivity may be a poor prognostic factor in AML.

Polymerase Chain Reaction Based Gene Rearrangement Studies in the Diagnosis of Follicular Lymphoma — Performance in Formaldehyde-Fixed Tissue and Application in Clinical Problem Cases

The diagnostic potential of PCR-based studies on the immunoglobulin heavy chain (IgH) gene rearrangements and the t(14;18) translocation was evaluated in a series of 30 benign and 46 malignant follicular lymphoid lesions including difficult problem cases using formaldehyde-fixed and paraffin-embedded (FFPE) tissue. Clonal IgH gene rearrangement was documented by PCR in 38/46 follicular lymphomas using two alternative (FR3A and FR2A-based) primer systems: all benign hyperplasias showed a polyclonal pattern. The t(14;18) translocation was documented in 20/46 cases, including cases that did not show a clonal IgH gene amplification; a weak signal was seen in 6 benign hyperplasias probably related to the recently described rare non-neoplastic cells carrying the t(14;18) translocation. Of the translocations in follicular lymphomas, 18 involved the major breakpoint region (MBR) and two the minor cluster region (MCR). The diagnosis of follicular lymphoma could be reached with at least one marker in over 90% of the cases. With these tests we diagnosed an incidental follicular lymphoma in the axillary dissection for breast carcinoma, and confirmed benign nature of an extreme follicular hyperplasia with a 2200 g spleen. One diffuse large cleaved cell lymphoma with subsequent follicular small cleaved cell lymphoma was diagnosed to have a similar t(14;18) translocation showing a link between the two seemingly different lymphomas.

THE SIGNIFICANCE OF B‐CELL CLONALITY IN GASTRIC LYMPHOID INFILTRATES

The significance of the demonstration of a clonal B-cell population in gastric lymphoid infiltrates was investigated by analysis of immunoglobulin heavy chain (IgH) gene rearrangements using sensitive polymerase chain reactions, employing fluorescently labelled primers to target the FR3 and FR1 regions. Tissue blocks were studied showing different histological features (high-grade lymphoma, low-grade lymphoma, and chronic gastritis) from 12 gastrectomies for primary gastric lymphoma, together with blocks showing chronic gastritis from 13 cases of gastric adenocarcinoma and biopsies from 33 patients with active Helicobacter-associated chronic gastritis. Clonal IgH gene rearrangements were detected in lymphoma samples from eight of the gastrectomies for lymphoma (67 per cent). In four of these eight specimens, clonal rearrangements were also detectable in the samples showing only chronic gastritis. Three of 28 (11 per cent) informative biopsies showing active Helicobacter-associated chronic gastritis had detectable clonal populations. Clonal rearrangements were also demonstrated in two of eight (25 per cent) informative blocks showing chronic gastritis from eight gastrectomies for adenocarcinoma. It is concluded that the detection of a clonal population in a suspicious lymphoid infiltrate does not confirm the diagnosis of lymphoma, nor does the absence of such a population imply benignity.

The significance of B-cell clonality in gastric lymphoid infiltrates.

The significance of the demonstration of a clonal B-cell population in gastric lymphoid infiltrates was investigated by analysis of immunoglobulin heavy chain (IgH) gene rearrangements using sensitive polymerase chain reactions, employing fluorescently labelled primers to target the FR3 and FR1 regions. Tissue blocks were studied showing different histological features (high-grade lymphoma, low-grade lymphoma, and chronic gastritis) from 12 gastrectomies for primary gastric lymphoma, together with blocks showing chronic gastritis from 13 cases of gastric adenocarcinoma and biopsies from 33 patients with active Helicobacter-associated chronic gastritis. Clonal IgH gene rearrangements were detected in lymphoma samples from eight of the gastrectomies for lymphoma (67 per cent). In four of these eight specimens, clonal rearrangements were also detectable in the samples showing only chronic gastritis. Three of 28 (11 per cent) informative biopsies showing active Helicobacter-associated chronic gastritis had detectable clonal populations. Clonal rearrangements were also demonstrated in two of eight (25 per cent) informative blocks showing chronic gastritis from eight gastrectomies for adenocarcinoma. It is concluded that the detection of a clonal population in a suspicious lymphoid infiltrate does not confirm the diagnosis of lymphoma, nor does the absence of such a population imply benignity.