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Castleman disease (CD) is a rare hematologic disorder characterized by pathologic lymph node changes and a range of symptoms due to excessive cytokine production. While uncontrolled infection with human herpesvirus-8 (HHV-8) is responsible for the cytokine storm in a portion of multicentric CD (HHV-8-associated MCD) cases, the etiology of unicentric CD (UCD) and HHV-8-negative/idiopathic MCD (iMCD) is unknown. Several hypotheses have been proposed regarding the pathogenesis of UCD and iMCD, including occult infection given the precedent established by HHV-8 infection. To investigate potential active infections in UCD and iMCD, we implemented Viral-Track, a computational method that identifies viral mRNA sequences from next-generation sequencing data. We applied Viral-Track to short sequencing reads from a cohort of UCD (n = 22), iMCD (n = 19), and controls (n = 86). While viral sequences for several unusual viruses were identified in individual CD patients, sequences for the same virus were not found across multiple CD patients or they were not specific to CD samples and were also found in non-CD samples. These results suggest that active viral infection is unlikely to be a pathological driver of UCD or iMCD. [ABSTRACT FROM AUTHOR]

Background: Despite recent advances in the treatment of aggressive lymphomas, a significant fraction of patients still succumbs to their disease. Thus, novel therapies are urgently needed. As the anti-CD20 antibody rituximab and the CD19-targeting antibody tafasitamab share distinct modes of actions, we investigated if dual-targeting of aggressive lymphoma B-cells by combining rituximab and tafasitamab might increase cytotoxic effects. Methods: Antibody single and combination efficacy was determined investigating different modes of action including direct cytotoxicity, antibody-dependent cellmediated cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP) in in vitro and in vivo models of aggressive B-cell lymphoma comprising diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma (BL). Results: Three different sensitivity profiles to antibody monotherapy or combination treatment were observed in in vitro models: while 1/11 cell lines was primarily sensitive to tafasitamab and 2/11 to rituximab, the combination resulted in enhanced cell death in 8/11 cell lines in at least one mode of action. Treatment with either antibody or the combination resulted in decreased expression of the oncogenic transcription factor MYC and inhibition of AKT signaling, which mirrored the cell line-specific sensitivities to direct cytotoxicity. At last, the combination resulted in a synergistic survival benefit in a PBMChumanized Ramos NOD/SCID mouse model. Conclusion: This study demonstrates that the combination of tafasitamab and rituximab improves efficacy compared to single-agent treatments in models of aggressive B-cell lymphoma in vitro and in vivo. [ABSTRACT FROM AUTHOR]

Objectives: Patients with clonal cytopenia of undetermined significance (CCUS) are at increased risk of developing myeloid neoplasia (MN). We evaluated whether a simple flow cytometry immunophenotyping (FCIP) assay could differentiate the risk of development of MN in patients with CCUS.Methods: Bone marrow aspirates were assessed by FCIP panel in a cohort of 80 patients identified as having CCUS based on next-generation sequencing or cytogenetics from March 2015 to May 2020, with available samples. Flow cytometric assay included CD13/HLA-DR expression pattern on CD34-positive myeloblasts; CD13/CD16 pattern on maturing granulocytic precursors; and aberrant expression of CD2, CD7, or CD56 on CD34-positive myeloblasts. Relevant demographic, comorbidity, and clinical and laboratory data, including the type and extent of genetic abnormalities, were extracted from the electronic health record.Results: In total, 17 (21%) patients with CCUS developed MN over the follow-up period (median survival follow-up, 28 months [95% confidence interval, 19-31]). Flow cytometry immunophenotyping abnormalities, including the aberrant pattern of CD13/HLA-DR expression, as detected at the time of the diagnosis of CCUS, were significantly associated with risk of developing MN (hazard ratio, 2.97; P = .006). Additional FCIP parameters associated with the development of MN included abnormal expression of CD7 on myeloblasts and the presence vs absence of any FCIP abnormality.Conclusions: A simple FCIP approach that includes assessment of CD13/HLA-DR pattern on CD34-positive myeloblasts can be useful in identifying patients with CCUS at higher risk of developing MN. [ABSTRACT FROM AUTHOR]

Objectives: Prognostically relevant chromosomal abnormalities in chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) are routinely identified by fluorescence in situ hybridization (FISH) on peripheral blood or bone marrow specimens. We studied the prevalence of chromosomal abnormalities on extramedullary tissues involved by CLL/SLL and evaluated their association with prominent proliferation centers (PPCs).Methods: FISH for recurrent abnormalities in CLL/SLL was performed on formalin-fixed, paraffin-embedded biopsy sections. PPCs were identified on H&E-stained sections. Available FISH results on peripheral blood or bone marrow specimens were also reviewed.Results: Recurrent FISH abnormalities were detected in 69% of 320 CLL/SLL biopsy specimens studied, including +12 (35%), 13q- (24%), 11q- (15%), 17p- (6%), 6q- (2%), and IGH/BCL2 (0.9%). Forty-three patients had abnormal blood or bone marrow FISH analyses, of whom 7 (16%) had discordant +12 and/or 13q-, and 3 (7%) had discordant 17p- or 11q-. Morphology was positive (17%), negative (78%), or equivocal (6%) for PPCs on 247 evaluable biopsy specimens, a finding not significantly associated with FISH results (P = .7).Conclusions: Trisomy 12 is overrepresented in tumoral CLL/SLL involvement, compared with the known predominance of 13q- in blood. Discrepancies between leukemic and tissue FISH findings are occasionally encountered. FISH results do not correlate with the presence of PPCs. [ABSTRACT FROM AUTHOR]

Objectives: Flow cytometric detection of T-cell clonality is challenging, particularly in differential diagnosis of immature T-cell proliferations. Studies have shown utility of TRBC1, in conjunction with other T-cell markers, as reliable means to identify T-cell clonality by flow cytometry. One limitation of surface TRBC1 (sTRBC1) evaluation is it cannot be detected in surface CD3 (sCD3)-negative T cells, such as normal or abnormal immature T-cell precursors. Here, we assess surface and cytoplasmic TRBC1 expression patterns in the differential diagnosis of T-lymphoblastic leukemia/lymphoma (T-ALL) vs normal thymocyte expansions.Methods: Forty-three samples containing T-ALL, thymoma, normal thymus, and/or indolent T-lymphoblastic proliferation (i-TLBP), were evaluated.Results: All 24 cases with normal thymocytes or i-TLBPs revealed a characteristic and reproducible sCD3/sTRBC1 expression pattern indicative of polytypic T-cell maturation. In contrast, all 19 T-ALLs lacked this polytypic maturation pattern and were either completely negative for sCD3/sTRBC1 or showed a minor sCD3-positive subset with a monotypic TRBC1 expression pattern. Cytoplasmic TRBC1 evaluation in 9 T-ALLs demonstrated a monotypic intracellular TRBC1-positive (n = 4) or TRBC1-negative (n = 5) expression, indicative of clonality.Conclusions: Our findings demonstrate flow cytometric evaluation of surface and cytoplasmic TRBC1 expression can aid detection of T-cell clonality and differential diagnosis of immature T-cell proliferations. [ABSTRACT FROM AUTHOR]

Objectives: The diagnosis of T-cell large granular lymphocytic leukemia (T-LGLL) is challenging because of overlapping immunophenotypic features with reactive T cells and limitations of T-cell clonality assays. We studied whether adding an antibody against T-cell receptor β constant region 1 (TRBC1) to a comprehensive flow cytometry panel could facilitate the diagnosis of T-LGLL.Methods: We added TRBC1 antibody to the standard T-cell and natural killer (NK) cell panel to assess T-cell clonality in 56 T-LGLLs and 34 reactive lymphocytoses. In addition, 20 chronic lymphoproliferative disorder of NK cells (CLPD-NKs) and 10 reactive NK-cell lymphocytoses were analyzed.Results: Clonal T cells were detected in all available T-LGLLs by monotypic TRBC1 expression and clonal/equivocal T-cell receptor gene rearrangement (TCGR) studies, compared with only 27% of T-LGLLs by killer-cell immunoglobulin-like receptor (KIR) restriction. Overall, 85% of T-LGLLs had a blood tumor burden greater than 500 cells/µL. Thirty-four reactive cases showed polytypic TRBC1 expression, except for 5 that revealed small T-cell clones of uncertain significance. All CLPD-NKs showed expected clonal KIR expression and negative TRBC1 expression.Conclusions: Addition of TRBC1 antibody to the routine flow cytometry assay could replace the TCGR molecular study and KIR flow cytometric analysis to assess clonality, simplifying the diagnosis of T-LGLL. [ABSTRACT FROM AUTHOR]

The aim is to prospectively evaluate the impact of a multidisciplinary lymphoma virtual tumor board. The utility of multi-site interactive lymphoma-specific tumor boards has not been reported. The Mayo Clinic Lymphoma Tumor Board is a component of the International Mayo Clinic Care Network (MCCN). The format includes the clinical case presentation, presentation of radiology and hematopathology findings by the appropriate subspecialist, proposed treatment options, review of the literature pertinent to the case, pharmacy contributions, and discussion followed by recommendations. Three hundred and nine consecutive highly selected real-time cases with a diagnosis of lymphoma were presented at the Mayo Clinic Lymphoma Tumor Board from January 2014 to June 2018 and decisions were prospectively tracked to assess its impact on the treatment decisions. A total of 309 cases were prospectively evaluated. One hundred and forty (45.3%) cases had some changes made or recommended. The total changes suggested were 179, as some cases had more than one recommendation. There were 93 (30%) clinical management recommendations, 45 (14.6%) additional testing recommendations, 29 (9.4%) pathology changes, and 6 (1.9%) radiology changes. In an electronic evaluation process, 93% of the responders reported an improvement in knowledge and competence, and 100% recommended no change in format of the board. A multidisciplinary lymphoma tumor board approach was found to have a meaningful impact on lymphoma patients while enhancing interdisciplinary interactions and education for multiple levels of the clinical care team. [ABSTRACT FROM AUTHOR]

Objectives: Distinguishing between T-cell and natural killer (NK)-cell neoplasms could be difficult given their overlapping immunophenotype. In this study, we investigated whether a flow cytometry assay with cytoplasmic staining for CD3 could be used for this purpose.Methods: Flow cytometry immunophenotyping was performed on 19 surface CD3 (sCD3)-negative mature T-cell neoplasms, 10 sCD3-positive mature T-cell neoplasms, 13 mature NK-cell neoplasms, and 19 normal controls. In addition to routine antibody panels (CD2, sCD3, CD4, CD5, CD7, CD8, CD16, CD45, CD56, CD57, CD94, CD158a, CD158b, CD158e, NKG2A TCRγ/δ), cytoplasmic staining for a monoclonal CD3 antibody (clone SK7/Leu-4) was assessed in all cases. A molecular study for T-cell receptor (TCR) gene rearrangement and an immunohistochemical study for TCRβ were performed.Results: Our data showed all T-cell neoplasms were uniformly positive for cytoplasmic CD3 (cCD3) regardless of sCD3 expression, whereas 85% of NK-cell neoplasms completely lacked cCD3 expression. The 2 cases with classic NK-cell immunophenotype but partial cCD3 expression showed no molecular genetic features of T-cell lineage by TCR gene rearrangement studies.Conclusions: Uniform cCD3 positivity and homogeneous cCD3 negativity highly suggest T-cell and NK lineage, respectively. When partial cCD3 expression is encountered, additional confirmatory studies should be pursued for the most accurate lineage assignment. [ABSTRACT FROM AUTHOR]