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Objectives To increase awareness of potential diagnostic test interference associated with alemtuzumab, which is a therapeutic immunoglobulin G1 κ monoclonal antibody used in hematologic malignancies, autoimmune diseases, and transplant-related disorders. Methods Bone marrow and blood from patients with T-cell prolymphocytic leukemia treated with alemtuzumab were evaluated by flow cytometry. Healthy donor blood was analyzed with or without in vitro treatment with alemtuzumab for comparison. Results Immunophenotypic analysis of bone marrow collected 4 weeks after alemtuzumab treatment demonstrated artifactual surface κ light chain restriction in CD19+ B cells and CD3+ T cells. Similar findings were observed in blood from another patient in a specimen collected 3 days after alemtuzumab treatment. These findings were recapitulated in healthy donor blood incubated with alemtuzumab. Conclusions Alemtuzumab can produce direct interference during flow cytometry analysis, resulting in false-positive evidence of light chain clonality. Clinicians and laboratorians should be cognizant of this risk to avoid misdiagnosis of B-cell neoplasms. [ABSTRACT FROM AUTHOR]

Objectives: To increase awareness of potential diagnostic test interference associated with alemtuzumab, which is a therapeutic immunoglobulin G1 κ monoclonal antibody used in hematologic malignancies, autoimmune diseases, and transplant-related disorders.Methods: Bone marrow and blood from patients with T-cell prolymphocytic leukemia treated with alemtuzumab were evaluated by flow cytometry. Healthy donor blood was analyzed with or without in vitro treatment with alemtuzumab for comparison.Results: Immunophenotypic analysis of bone marrow collected 4 weeks after alemtuzumab treatment demonstrated artifactual surface κ light chain restriction in CD19+ B cells and CD3+ T cells. Similar findings were observed in blood from another patient in a specimen collected 3 days after alemtuzumab treatment. These findings were recapitulated in healthy donor blood incubated with alemtuzumab.Conclusions: Alemtuzumab can produce direct interference during flow cytometry analysis, resulting in false-positive evidence of light chain clonality. Clinicians and laboratorians should be cognizant of this risk to avoid misdiagnosis of B-cell neoplasms. [ABSTRACT FROM AUTHOR]

Keywords: leukapheresis; leukemia EN leukapheresis leukemia 277 278 2 02/07/23 20230201 NES 230201 A 70-year-old man presented to an outside hospital with a three-day history of fever and abdominal pain. CD2 expression, especially when coexpressed with CD34, is associated with the microgranular variant of APL.[2] In addition to astute morphologic assessment, recognizing this unique immunophenotype may be useful when considering emergent leukapheresis before t(15;17) studies result. [Extracted from the article]

Background: Acute myeloid leukemia (AML) with RAM immunophenotype is a newly recognized high-risk AML immunophenotypic subcategory characterized by blasts with bright expression of CD56 and weak to absent expression of CD45, HLA-DR, and CD38, as first described by the Children's Oncology Group (COG). The relationship between AML-RAM and other CD56-positive acute leukemias is unclear. The goal of this study is to characterize the clinicopathological characteristics of AML with RAM phenotype and compare them with other CD56 co-expressing acute leukemias., Methods: From a multi-institutional search, we identified a total of 160 CD56+ acute leukemia cases, including AML-RAM ( n  = 28), CD56+ acute undifferentiated leukemia (AUL) ( n  = 11), CD56+ T-lymphoblastic leukemia ( n  = 39), and CD56+ AML ( n  = 81). We compared the clinical and pathologic findings of these groups., Results: AML-RAM patients were significantly younger and presented with significantly higher platelet and white blood cell counts and bone marrow (BM) blast percentages when compared to AUL ( p  > 0.05) and had higher median BM blast percentages than T-ALL and CD56+ AML groups (both p  < 0.05). Flow cytometry showed significantly brighter expression of CD56 on blasts as compared to other CD56+ AML cases, partial CD34 expression compared to AUL, and AML, weak-to-absent CD38 expression compared to all groups, and absent HLA-DR and terminal deoxynucleotidyl transferase as compared to AUL and T-ALL (all p  < 0.05). The frequency of abnormal karyotypes was significantly higher among RAM when compared to all groups ( p  < 0.05). Next-generation sequencing profiles differed among the leukemia groups, with significant enrichment of CBFA2T3::GLIS2 fusions ( p  < 0.05) and TP53 mutations ( p  < 0.05) in RAM cases compared to other AML control groups, and U2AF1 ( p  < 0.05), serine and arginine-rich splicing factor 2 ( p  < 0.05), and BCL6 co-repressor ( p  < 0.05) mutations compared to AUL. Clinical outcome analysis demonstrated significantly lower 3-year overall survival of the RAM subgroup (36 months) compared to control groups ( p  = 0.002)., Conclusion: We find that AML with RAM phenotype occurs primarily in younger ages, with distinct clinicopathological, immunophenotypic, and mutational presentations, and worse prognosis. This diagnosis should be considered in the clinical differential diagnosis of CD56-positive acute leukemias., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). eJHaem published by British Society for Haematology and John Wiley & Sons Ltd.)

Objective: The aim of this study was to determine the efficacy of plerixafor for hematopoietic stem cell (HSC) mobilization prior to autologous stem cell transplantation (aSCT) for patients with multiple myeloma (MM) and various lymphomas, using an oncologist-guided HSC collection goal and markers of cell viability., Methods: A retrospective chart review of all aSCT patients at Yale New Haven Hospital between 2017 and 2021 who met diagnostic criteria for MM, non-Hodgkin, or Hodgkin lymphoma (n = 382) was undertaken. Logistic regression evaluated plerixafor's effect on meeting the individual's HSC goal. The use of t-tests determined plerixafor's relationship to HSC yield and analysis of variance testing assessed its effect on cell viability., Results: Mobilization with granulocyte colony-stimulating factor (G-CSF) and plerixafor (odds ratio [OR] = 0.08; P < .05) relative to G-CSF alone was negatively associated with meeting the individual's HSC goal. Diffuse large B-cell lymphoma in patients mobilized with plerixafor yielded fewer HSCs than those without plerixafor (t = -2.78; P = .03). Mobilization regimen (P = .13) had no association with HSC viability. Mobilization failure with plerixafor was rare but occurred in patients with multiple risk factors, including exposure to several rounds of HSC-affecting chemotherapy., Conclusion: Plerixafor is effective across multiple diagnoses using an oncologist-driven HSC collection endpoint. Its association with mobilization failure is likely attributable to its use in patients predicted to be poor mobilizers., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society for Clinical Pathology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

During the COVID-19 pandemic, hematopoietic stem cell transplant (HSCT) recipients faced an elevated mortality rate from SARS-CoV-2 infection, ranging between 10-40%. The SARS-CoV-2 mRNA vaccines are important tools in preventing severe disease, yet their efficacy in the post-transplant setting remains unclear, especially in patients subjected to myeloablative chemotherapy and immunosuppression. We evaluated the humoral and adaptive immune responses to the SARS-CoV-2 mRNA vaccination series in 42 HSCT recipients and 5 healthy controls. Peripheral blood mononuclear nuclear cells and serum were prospectively collected before and after each dose of the SARS-CoV-2 vaccine. Post-vaccination responses were assessed by measuring anti-spike IgG and nucleocapsid titers, and antigen specific T cell activity, before and after vaccination. In order to examine mechanisms behind a lack of response, pre-and post-vaccine samples were selected based on humoral and cellular responses for single-cell RNA sequencing with TCR and BCR sequencing. Our observations revealed that while all participants eventually mounted a humoral response, transplant recipients had defects in memory T cell populations that were associated with an absence of T cell response, some of which could be detected pre-vaccination.

A subset of myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) show complex karyotype (CK), and these cases include a relatively high proportion of cases of therapy-related myeloid neoplasms and TP53 mutations. We aimed to evaluate the clinicopathologic features of outcome of 299 AML and MDS patients with CK collected from multiple academic institutions. Mutations were present in 287 patients (96%), and the most common mutation detected was in TP53 gene (247, 83%). A higher frequency of TP53 mutations was present in therapy-related cases (P = .008), with a trend for worse overall survival (OS) in therapy-related patients as compared with de novo disease (P = .08) and within the therapy-related group; the presence of TP53 mutation strongly predicted for worse outcome (P = .0017). However, there was no difference in survival between CK patients based on categorization of AML vs MDS (P = .96) or presence of absence of circulating blasts ≥1% (P = .52). TP53-mutated patients presented with older age (P = .06) and lower hemoglobin levels (P = .004) and marrow blast counts (P = .02) compared with those with CK lacking TP53 mutation. Multivariable analysis identified presence of multihit TP53 mutation as strongest predictor of worse outcome, whereas neither a diagnosis of AML vs MDS nor therapy-relatedness independently influenced OS. Our findings suggest that among patients with MDS and AML, the presence of TP53 mutation (in particular multihit TP53 mutation) in the context of CK identifies a homogeneously aggressive disease, irrespective of the blast count at presentation or therapy-relatedness. The current classification of these cases into different disease categories artificially separates a single biologic disease entity., (© 2022 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Purpose of Review: In this review, we provide a comprehensive and contemporary understanding of malignant monocytosis and provide a framework by which the appropriate diagnosis with malignant monocytosis can be rendered., Recent Findings: Increasing data support the use of molecular data to refine the diagnostic approach to persistent monocytosis. The absence of a TET2, SRSF2, or ASXL1 mutation has ≥ 90% negative predictive value for a diagnosis of CMML. These data may also reliably differentiate chronic myelomonocytic leukemia, the malignancy that is most associated with mature monocytosis, from several other diseases that can be associated with typically a lesser degree of monocytosis. These include acute myelomonocytic leukemia, acute myeloid leukemia with monocytic differentiation, myelodysplastic syndromes, and myeloproliferative neoplasms driven by BCR-ABL1, PDGFRA, PDGFRB, or FGFR1 rearrangements or PCM1-JAK2 fusions among other rarer aberrations. The combination of monocyte partitioning with molecular data in patients with persistent monocytosis may increase the predictive power for the ultimate development of CMM but has not been prospectively validated. Many conditions, both benign and malignant, can be associated with an increase in mature circulating monocytes. After reasonably excluding a secondary or reactive monocytosis, there should be a concern for and investigation of malignant monocytosis, which includes hematopathologic review of blood and marrow tissues, flow cytometric analysis, and cytogenetic and molecular studies to arrive at an appropriate diagnosis.

A hallmark of lymphoid malignancies is the presence of a monoclonal lymphocyte population. Monoclonality of B- and T-cell populations can be established through immunoglobulin (IG) or T-cell receptor (TCR) gene rearrangement analysis, respectively. The biological rationale of IG and TCR gene rearrangement analysis is that due to the extensive combinatorial repertoire made possible by V(D)J recombination in lymphocytes, it is unlikely that any substantive lymphocyte population would share the same IG or TCR gene rearrangement pattern unless there is an underlying neoplastic or reactive origin. Modern IG and TCR gene rearrangement analysis is typically performed by polymerase chain reaction (PCR) using commercially available primer sets followed by gel capillary electrophoresis. This process is highly sensitive in the detection of nearly all lymphoid malignancies. Several pitfalls and limitations, both biological and technical, apply to IG/TCR gene rearrangement analysis, but these can be minimised with high quality controls, performance of assays in duplicate, and adherence to strict criteria for interpreting and reporting results. Next generation sequencing (NGS) will likely replace PCR based methods of IG/TCR gene rearrangement analysis but is not yet widespread due to the absence of standardised protocols and multicentre validation., (Copyright © 2020 Royal College of Pathologists of Australasia. Published by Elsevier B.V. All rights reserved.)

Context.—: The significance of positive immunoglobulin (IG) or T-cell receptor (TCR) gene rearrangement studies in the context of otherwise normal ancillary findings is unknown., Objective.—: To examine long-term hematologic outcomes of individuals with positive gene rearrangement studies with otherwise unremarkable blood or bone marrow studies in parallel., Design.—: Data from patients who underwent IG or TCR gene rearrangement testing at the authors' affiliated Veterans Affairs Hospital January 1, 2013 to July 6, 2018 were extracted from medical records. Date of testing, specimen source, and morphologic, flow cytometric, immunohistochemical, and cytogenetic characterization of the tissue source were recorded. Gene rearrangement results were categorized as test positive/phenotype positive (T+/P+), test positive/phenotype negative (T+/P-), test negative/phenotype negative (T-/P-), or test negative/phenotype positive (T-/P+) based on comparison to other studies and/or final diagnosis. Patient records were reviewed for subsequent diagnosis of hematologic malignancy for patients with positive gene rearrangements but no other evidence for a disease process., Results.—: A total of 136 patients with 203 gene rearrangement studies were analyzed. For TCR studies, there were 2 T+/P- and 1 T-/P+ results in 47 peripheral blood assays, as well as 7 T+/P- and 1 T-/P+ results in 54 bone marrow assays. Regarding IG studies, 3 T+/P- and 12 T-/P+ results in 99 bone marrow studies were identified. None of the 12 patients with T+/P- TCR or IG gene rearrangement studies later developed a lymphoproliferative disorder., Conclusions.—: Positive IG/TCR gene rearrangement studies in the context of otherwise negative bone marrow or peripheral blood findings are not predictive of lymphoproliferative disorders., Competing Interests: The authors have no relevant financial interest in the products or companies described in this article.

Context.—: A novel electronic consult (e-consult) system for a pathology and laboratory medicine service (PLMS) was implemented in 2015 at a high-complexity Veterans Administration health care facility. Consults were previously made through direct provider communication without documentation in the medical record., Objective.—: To evaluate the utilization trends of the laboratory e-consult system at the Department of Veterans Affairs Connecticut facility during the first 2 years since inception., Design.—: E-consultation involves pathology and laboratory medicine resident review followed by attending pathologist review and cosignature. E-consults to the pathology and laboratory medicine service from 2015 to 2017 were reviewed to record type of consult, requesting department, patient location, and turnaround time., Results.—: The pathology and laboratory medicine service received 351 e-consults from 2015 to 2017. The volume varied by subsection: hematology and coagulation (215 of 351; 61%), chemistry (109 of 351; 31%), blood bank (19 of 351; 6%), and microbiology/virology (8 of 351; 2%). Hematology and coagulation consults were entirely for peripheral blood smear review (215 of 215; 100%). Chemistry consults were placed for toxicology/drugs of abuse (81 of 109; 74%), test utilization (17 of 109; 16%), or nontoxicology (11 of 109; 10%). Three services placed the majority of consults: primary care (279 of 351; 80%), hematology/oncology (39 of 351; 11%), and psychiatry (27 of 351; 8%). The median turnaround time for completion of e-consults was 1.2 days. Since e-consult implementation, the mean number of consults increased from 8.6/mo in 2015 to 18.1/mo in 2017, peaking in the last quarter of analysis in 2017 with a mean of 25.3 consults/mo., Conclusions.—: This novel e-consult system improved accessibility to and documentation of answers to laboratory questions and increased the visibility of the pathology and laboratory medicine service. Future goals include development of outcomes-based measures to better assess the clinical impact of e-consults., (© 2021 College of American Pathologists.)

Background: People living with sickle cell disease (SCD) are prone to red blood cell (RBC) alloimmunization. We hypothesized that subjects with alloantibodies (responders) would have differences in circulating T-follicular helper (Tfh)-like cells compared to subjects without alloantibodies (non-responders)., Materials and Methods: Peripheral blood mononuclear cells were collected from 28 subjects, including those with SCD and controls. Circulating CD4 T-cell subsets were first evaluated at baseline. CD4 T-cell subsets were also evaluated after naïve CD4 T-cells were differentiated into Tfh-like cells following in vitro culture with CD3/CD28 beads, IL-7, IL-12, and Activin A. Transfusion and alloantibody histories were extracted from the electronic medical record., Results: Non-responders had a lower percentage of CD45RA negative Tmemory cells than responders or controls (p<0.05). Notably, there were no differences in circulating Tfh-like cells between any group. However, naïve CD4 T-cells from subjects with SCD were more likely to express CXCR5 after in vitro culture than cells from controls. After culture, CXCR5 expressing cells from responders were more likely to express PD1 and ICOS (16.43 %, sd. 20.23) compared to non-responders (3.69 %, s.d. 3.09) or controls (2.78 %, s.d. 2.04)., Discussion: The tendency for naïve CD4 T-cells from responders to differentiate into Tfh-like cells after in vitro culture may suggest these cells are prepared to assist B-cells with antibody production regardless of antigen specificity. Further studies are needed, but it is possible that these results may explain why some responders form RBC alloantibodies with multiple specificities, in addition to RBC autoantibodies and HLA alloantibodies., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest relevant to this manuscript., (Copyright © 2020 Elsevier Ltd. All rights reserved.)