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Howard Ratech

Publications and source records attributed to Howard Ratech.

9 recordsLinked to original sources
PubMed2006

AIDS and non-AIDS diffuse large B-cell lymphomas express different antigen profiles.

Based on gene expression profiling, diffuse large B-cell lymphomas arising in immunocompetent patients can be divided into germinal center and activated B-cell types. Since little is known about acquired immunodeficiency syndrome associated diffuse large B-cell lymphomas, we tested whether the protein expression of germinal center and activated B-cell markers differed between acquired immunodeficiency syndrome (AIDS) vs non-AIDS diffuse large B-cell lymphomas. We immunohistochemically stained tissue microarrays of 39 de novo diffuse large B-cell lymphomas: 12 AIDS associated and 27 non-AIDS, with germinal center (BCL6, CD10, CyclinH) and activated B-cell markers (MUM1, CD138, PAK1, CD44, BCL2). We scored each case for percent positive cells (0-19%=0; 20-49%=1; 50-100%=2). The activated B-cell and germinal center summation scores of each case were used as (x, y) coordinate data points to construct two-dimensional contour-frequency plots. The contour plot of non-AIDS diffuse large B-cell lymphomas showed two distinct clusters: a cluster with a high germinal center phenotype (cluster 1) and a cluster with a high activated B-cell phenotype (cluster 3). In contrast, the AIDS-related diffuse large B-cell lymphomas formed a single aggregate (cluster 2) (P=0.02, Fisher exact test). When the contour plots of the AIDS-related and the non-AIDS cases were superimposed, cluster 2 of the AIDS cases expressed an intermediate germinal center/activated B-cell phenotype compared to clusters 1 and 3 of the non-AIDS diffuse large B-cell lymphomas. Our results confirm that non-AIDS diffuse large B-cell lymphomas segregate into two groups with either germinal center or activated B-cell phenotype. We report the new finding that the AIDS status of the patient predicts the immunophenotype of the diffuse large B-cell lymphomas.

Adult↗
PubMed2005

Rapid detection of clonal T-cell receptor-beta gene rearrangements in T-Cell lymphomas using the LightCycler-polymerase chain reaction with DNA melting curve analysis.

Various molecular methods have been developed to diagnose clonal T-cell receptor (TCR) gene rearrangements in clinical samples. Most polymerase chain reaction strategies for detecting clonal TCR gene rearrangements rely on either gel or capillary electrophoresis. However, a cumbersome manual transfer step separates amplification from analysis. Recently, we developed a novel polymerase chain reaction assay using the LightCycler system to detect clonal immunoglobulin heavy chain gene rearrangement. In the current study, we extend this work to include the TCR. We report that clonal TCR-beta (TCR-beta) gene rearrangements can be detected in less than 1 hour after preparing the DNA by measuring DNA melting immediately after amplification in a single closed capillary tube. We retrospectively studied 52 fresh-frozen tissue samples from patients clinically suspected of T-cell malignancy. A clonal TCR-beta gene rearrangement was detected in 14 samples by DNA melting curve analysis. When DNA melting was compared to the gold standard methods of Southern blot or denaturing gradient gel electrophoresis, it achieved a sensitivity equal to 71% and a specificity equal to 94%. We also compared melting curve analysis and polyacrylamide gel electrophoresis: melting curve analysis reached a sensitivity equal to 100% and a specificity equal to 97%. We conclude that DNA melting curve analysis in the LightCycler system has potential for clinical use as a new, ultra-fast method for the initial diagnosis of clonal TCR-beta gene rearrangements.

Adult↗
PubMed2005

Germinal center and activated b-cell profiles separate Burkitt lymphoma and diffuse large B-cell lymphoma in AIDS and non-AIDS cases.

Morphologic features of Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL) overlap. No single phenotypic marker or molecular abnormality is pathognomonic. We tested a panel of 8 germinal center (GC) and activated B-cell (ABC) markers for their ability to separate BL and DLBCL. We diagnosed 16 BL and 39 DLBCL cases from 21 patients with AIDS and 34 without AIDS based on traditional morphologic criteria, Ki-67 proliferative index, and c-myc rearrangement (fluorescence in situ hybridization). After immunohistochemically staining tissue microarrays of BL and DLBCL for markers of GC (bcl-6, CD10, cyclin H) and ABC (MUM1, CD138, PAK1, CD44, bcl-2), we scored each case for the percentage of positive cells. Hierarchical clustering yielded 2 major clusters significantly associated with morphologic diagnosis (P < .001). For comparison, we plotted the sum of the GC scores and ABC scores for each case as x and y data points. This revealed a high-GC/low-ABC group and a low-GC/high-ABC group that were associated significantly with morphologic diagnosis (P < .001). Protein expression of multiple GC and ABC markers can separate BL and DLBCL.

Adolescent↗
PubMed2004

Rapid diagnosis of clonal immunoglobulin heavy chain gene rearrangements in cutaneous B-cell lymphomas using the LightCycler-Polymerase Chain Reaction with DNA melting curve analysis.

We have recently developed a novel Immunoglobulin heavy chain gene rearrangement (IgH-R) assay that combines polymerase chain reaction (PCR) amplification and analysis in the same closed capillary tube using the LightCycler System. IgH-R can be identified by DNA melting curve analysis within 40 minutes after DNA preparation and amplification. To test the clinical utility of this new IgH-R assay for rapidly diagnosing cutaneous B-cell lymphomas, we prospectively analyzed 44 formalin-fixed, paraffin-embedded tissues suspected of B-cell malignant lymphoma: skin (n = 31), lymph node (n = 7), stomach (n = 3), spleen (n = 1), colon (n = 1), and soft tissue (n = 1). We detected IgH-R in 12 DNA samples, including 8 skin biopsies, with the following diagnoses: B-cell chronic lymphocytic leukemia (n = 4), extranodal marginal zone B-cell lymphoma (n = 4), diffuse large B-cell lymphoma (n = 2), Burkitt lymphoma (n = 1), and precursor B-lymphoblastic lymphoma (n = 1). DNA melting curve analysis, compared with polyacrylamide gel electrophoresis, achieved a sensitivity equal to 92.3% and a specificity equal to 100%. There was a single false negative result because DNA melting curve analysis could not detect less than 10.0% clonal B-cells. We conclude that this new, rapid PCR assay for detecting IgH-R based on DNA melting curve analysis can be clinically useful for confirming the initial diagnosis of B-cell malignant lymphoma.

Adult↗
PubMed2003

Evidence for expression of early myeloid antigens in mature, non-blast myeloid cells in myelodysplasia.

Myelodysplastic syndromes (MDS) are clonal hematopoietic stem cell disorders with frequent cytogenetic abnormalities. They can arise de novo or be related to therapy. Although blasts in MDS have been studied extensively, there is little information available on the mature, non-blast myeloid cells (NBMCs). We used a retrospective case-control study design. NBMC populations in MDS (48 cases) and in tumor-free control (12 cases) bone marrow samples were analyzed using multiparameter flow cytometry for mean side scatter (SSC) channel number and for expression of aberrant cell surface antigens. MDS cases were stratified on the basis of cytogenetic abnormalities. We report that NBMCs in MDS with normal karyotype expressed significantly higher HLA-DR than controls (P = 0.034). NBMCs in MDS cases with cytogenetic abnormalities and with > or =5% marrow blasts, compared with controls, had significantly higher CD34 and higher HLA-DR but lower CD10 and lower SSC mean channel number. CD34 expression in NBMCs was significantly greater in therapy-related MDS compared with de novo MDS ( P = 0.01), although the presence of cytogenetic abnormalities was not different ( P > 0.05). These data suggest that bone marrow, mature, NBMCs have phenotypic changes in MDS that are not seen in normal controls.

Adult↗
PubMed2003

Phase II trial of infusional cyclophosphamide, idarubicin, and etoposide in poor prognosis non-Hodgkin's lymphoma.

The purpose of this study was to determine the complete response (CR) rate, failure-free survival (FFS), and overall survival (OS) of patients with poor-prognosis intermediate-grade non-Hodgkin's lymphoma (NHL) after treatment with cyclophosphamide, idarubicin, and etoposide given as a continuous intravenous infusion (CIVI) over 96 hours (infusional CIE), including patients with relapsed/refractory disease and patients with no prior therapy but at least two poor-risk features by the age-adjusted International Prognostic Index. Forty-two patients with previously untreated NHL (N = 24) or relapsed/refractory (N = 18) NHL received cyclophosphamide (200 mg/m2/d), idarubicin (2.5-3.0 mg/m2/d) and etoposide (60 mg/m2/d) given by a 96-hour CIVI every 3 weeks for a maximum of 8 cycles. All patients also received granulocyte-colony-stimulating factor. CR occurred in 10 of 24 patients (42%; 95% confidence intervals [CI] 22%, 62%) treated with CIE as first-line therapy, and in 3 of 18 patients (17%; 95% CI 20%, 32%) treated with CIE as second-line or greater therapy. One-year FFS and OS were 42% and 64%, respectively, in patients with no prior therapy, and 17% and 56% in patients with prior therapy. Severe (grade III) or life-threatening (grade IV) toxicity included leukopenia (59%), anemia (61%), thrombocytopenia (31%), and infection (10%). Two patients (4%) died due to treatment related infectious complications. It is unlikely that infusional CIE produces a CR rate more than about 60% in poor-risk patients with intermediate-grade NHL when used as first-line therapy, or more than about 30% in patients receiving the regimen as second-line therapy. Substitution of idarubicin for doxorubicin in this setting, therefore, is not associated with an improved response rate.

Adult↗
PubMed2003

Human immunodeficiency virus-associated lymphoma.

Infection with the human immunodeficiency virus (HIV) is associated with an increased risk of systemic non-Hodgkin's lymphoma, Hodgkin's disease, and primary central nervous system lymphoma (PCNSL). Systemic lymphoma usually involves extranodal sites (80%-90%) and is usually of intermediate-grade (diffuse large-cell or immunoblastic( or high-grade (diffuse small noncleaved) histology. Approximately one third to one half of patients are cured with the cytotoxic treatment regimens that are used in immunocompetent patients with lymphoma. Careful attention must be paid to appropriate treatment of HIV infection and to primary and secondary infection prophylaxis. Colony-stimulating factors are commonly used in conjunction with cytotoxic therapy because of the high risk of febrile neutropenia. Patients with HIV-associated Hodgkin's disease also frequently have extranodal involvement and mixed cellularity histology, features associated with an adverse prognosis in immunocompetent patients. Treatment regimens used to treat Hodgkin's disease in immunocompetent patients have been used with some success, although the prognosis is not favorable in HIV-infected patients with PCNSL is generally poor because such patients typically present with advanced immunodeficiency (CD4 <50/microL), and the lymphoma often relapses after transient initial response to whole brain irradiation. There are anecdotal reports of responses to therapy directed against Epstein-Barr virus (ie, high-dose zidovudine, gancyclovir, and interleukin-2).

Anti-Retroviral Agents↗
PubMed2002

Rapid and accurate detection of monoclonal immunoglobulin heavy chain gene rearrangement by DNA melting curve analysis in the LightCycler System.

The detection of immunoglobulin heavy chain gene rearrangement (IgH-R) is a standard tool for distinguishing polyclonal from monoclonal B-cell populations. Current DNA-based polymerase chain reactions (PCR) strategies can diagnose monoclonal IgH-R either by measuring the length of the amplicon or by detecting gel mobility variations owing to sequence-dependent conformational changes. However, amplification and analysis remain sequential operations usually requiring manual transfer. We have developed a novel PCR strategy for detecting monoclonal IgH-R that monitors fluorescence of the specific double-stranded DNA binding dye SYBR Green I during melting curve analysis using the LightCycler System. We compared polyacrylamide gel electrophoresis (PAGE) versus melting curve analysis in 130 clinical DNA samples from formalin-fixed, paraffin-embedded (FFPE) tissues (mostly skin biopsies) of 128 patients. The identical FR3 primers were used to amplify the IgH variable region for both analytic techniques. We detected IgH-R in 24 DNA samples from FFPE tissue of 22 patients. Melting curve analysis, compared to PAGE, revealed no false negative and no false positive results, yielding both sensitivity and specificity equal to 100%. We also compared Southern blot analysis versus melting curve analysis in 23 clinical DNA samples from fresh-frozen lymph nodes of 23 patients. We detected IgH-R by melting curve analysis in 7 DNA samples from fresh-frozen lymph nodes. Melting curve analysis, compared to Southern blot analysis, revealed sensitivity equal to 58.3% (7 of 12) and specificity equal to 100% (11 of 11). We conclude that continuous fluorescence monitoring of PCR products with DNA melting curve analysis can rapidly and reproducibly distinguish polyclonal from monoclonal B-cell populations.

Adult↗