PCNEO, a New Proficiency Testing Program for Flow Cytometric Analysis of Plasma Cell Neoplasms From the College of American Pathologists Diagnostic Immunology and Flow Cytometry Committee.

CONTEXT.­: In 2018 the College of American Pathologists Diagnostic Immunology and Flow Cytometry Committee designed and implemented a new plasma cell neoplasia flow cytometry proficiency testing program-PCNEO-to allow clinical flow cytometry laboratories to monitor and assess their performance compared with a peer group. OBJECTIVE.­: To report the results from the first 4 years of the PCNEO program. DESIGN.­: Program participants were sent 2 sets of challenges per year, each including 1 wet challenge and 2 dry challenges, with associated clinical and laboratory findings. The wet challenges were composed of myeloma cell line specimens (with or without dilution in preserved whole blood) for flow cytometric analysis. The dry (paper) challenges were composed of clinical case summaries and images of flow cytometric test results from various flow cytometry laboratories of committee members. RESULTS.­: A total of 116 to 145 laboratories from 17 countries enrolled in the proficiency testing program. For the wet challenges, almost all participants (97%-100%; cumulative, 98.2%) correctly identified the presence of neoplastic plasma cell populations based on flow cytometric analysis of undiluted myeloma cell lines. Slightly fewer participants (89.0%-97.4%; cumulative, 95.2%) correctly identified the presence of neoplastic plasma cell populations based on flow cytometric analysis of diluted myeloma cell lines (10% or 50% dilutions into peripheral blood) intended to better represent a typical clinical sample. There was generally agreement among 80% or more of participants for positive or negative staining for CD38, CD138, CD19, CD20, and surface and cytoplasmic κ and λ light chains. Similarly, 84% to 100% of participants were able to correctly identify the presence of neoplastic plasma cell populations in paper challenges, including the presence of small, neoplastic plasma cell populations (0.01%-5.0% clonal plasma cells), or the presence of nonneoplastic plasma cell populations (correctly identified by 91%-96% of participants). CONCLUSIONS.­: Participant performance in the new proficiency testing program was excellent overall, with the vast majority of participants able to perform flow cytometric analysis and identify neoplastic plasma cell populations, and to identify small plasma cell clones or expanded populations of reactive plasma cells in dry challenge flow cytometry results. This program will allow laboratories to verify the accuracy of their testing program and test interpretations for the assessment of patients suspected of having a plasma cell neoplasm.

Evaluation and Comparison of Performance Parameters and Impact of Telepathology and Operator Experience on Endobronchial and Endoscopic Ultrasound-Guided Fine-Needle Aspiration.

OBJECTIVES: Endobronchial ultrasound- and endoscopic ultrasound-guided fine-needle aspiration (EBUS-/EUS-FNA) are minimally invasive techniques of diagnosing and staging malignancies. The procedures are difficult to master, requiring specific feedback for optimizing yield. METHODS: Over 2 years, EBUS-/EUS-FNA cases were gathered using the institutional pathology database. Patient and specimen characteristics were collected from the pathology database and electronic medical record. RESULTS: In 2 years, 789 unique FNA specimens were collected (356 EBUS and 433 EUS specimens). The cohort and each subgroup had excellent performance, which was enhanced by telepathology. The discrepancy rate was satisfactorily low. Hematolymphoid neoplasms are overrepresented in discrepant EBUS cases. The malignancy rates of cytology diagnostic categories were comparable to the literature. CONCLUSIONS: Using diagnostic yield and concordance results allow for comprehensive evaluation of the entire process of EBUS-/EUS-FNAs. This study's findings can influence patient management, training methods, and interpretation of results, while also acting as a model for others to investigate their own sources of inadequacy, discrepancy, and training gaps.

Participation in the College of American Pathologists Laboratory Accreditation Program Decreases Variability in B-Lymphoblastic Leukemia and Plasma Cell Myeloma Flow Cytometric Minimal Residual Disease Testing: A Follow-up Survey.

CONTEXT.­: Minimal residual disease (MRD) testing by flow cytometry is ubiquitous in hematolymphoid neoplasm monitoring, especially B-lymphoblastic leukemia (B-ALL), for which it provides predictive information and guides management. Major heterogeneity was identified in 2014. Subsequently, new Flow Cytometry Checklist items required documentation of the sensitivity determination method and required lower level of detection (LLOD) inclusion in final reports. This study assesses Laboratory Accreditation Program (LAP) participation and new checklist items' impact on flow cytometry MRD testing. OBJECTIVES.­: To survey flow cytometry laboratories about MRD testing for B-ALL and plasma cell myeloma. In particular, enumerate the laboratories performing MRD testing, the proportion performing assays with very low LLODs, and implementation of new checklist items. DESIGN.­: Supplemental questions were distributed in the 2017-A mailing to 548 flow cytometry laboratories subscribed to the College of American Pathologists FL3 Proficiency Testing Survey (Flow Cytometry-Immunophenotypic Characterization of Leukemia/Lymphoma). RESULTS.­: The percentage of laboratories performing MRD studies has significantly decreased since 2014. Wide ranges of LLOD and collection event numbers were reported for B-ALL and plasma cell myeloma. Most laboratories determine LLOD by using dilutional studies and include it in final reports; a higher proportion of LAP participants used these practices than nonparticipants. CONCLUSIONS.­: Several MRD testing aspects vary among laboratories receiving FL3 Proficiency Testing materials. After the survey in 2014, new checklist items were implemented. As compared to 2014, fewer laboratories are performing MRD studies. While LLOD remains heterogeneous, a high proportion of LAP subscribers follow the new checklist requirements and, overall, target LLOD recommendations from disease-specific working groups are met.

Concurrent Subcutaneous Panniculitis-like T-Cell Lymphoma and B-Cell Acute Lymphoblastic Leukemia in 2 Pediatric Patients.

Subcutaneous panniculitis-like T-cell lymphoma is a cutaneous lymphoma characterized by CD8+ T-cell infiltrate in the subcutis that is rare in children. Acute lymphoblastic lymphoma is the most common pediatric malignancy and often presents with fevers and pancytopenia. Herein, we report 2 pediatric patients presenting with subcutaneous panniculitis-like T-cell lymphoma and B-cell acute lymphoblastic lymphoma, distinct hematologic malignancies arising from different lymphoid lineages, with no identifiable germline cancer predisposition.

Metabolic reprogramming augments potency of human pSTAT3-inhibited iTregs to suppress alloreactivity.

Immunosuppressive donor Tregs can prevent graft-versus-host disease (GVHD) or solid-organ allograft rejection. We previously demonstrated that inhibiting STAT3 phosphorylation (pSTAT3) augments FOXP3 expression, stabilizing induced Tregs (iTregs). Here we report that human pSTAT3-inhibited iTregs prevent human skin graft rejection and xenogeneic GVHD yet spare donor antileukemia immunity. pSTAT3-inhibited iTregs express increased levels of skin-homing cutaneous lymphocyte-associated antigen, immunosuppressive GARP and PD-1, and IL-9 that supports tolerizing mast cells. Further, pSTAT3-inhibited iTregs significantly reduced alloreactive conventional T cells, Th1, and Th17 cells implicated in GVHD and tissue rejection and impaired infiltration by pathogenic Th2 cells. Mechanistically, pSTAT3 inhibition of iTregs provoked a shift in metabolism from oxidative phosphorylation (OxPhos) to glycolysis and reduced electron transport chain activity. Strikingly, cotreatment with coenzyme Q10 restored OxPhos in pSTAT3-inhibited iTregs and augmented their suppressive potency. These findings support the rationale for clinically testing the safety and efficacy of metabolically tuned, human pSTAT3-inhibited iTregs to control alloreactive T cells.

Cytomorphologic features of malignant solitary fibrous tumor with mediastinal involvement sampled by endoscopic and endobronchial ultrasound-guided fine-needle aspiration: A comparison of two cases.

Solitary fibrous tumor (SFT) is an uncommon fibroblastic neoplasm with considerable risk of local recurrence. SFT is histologically characterized by bland spindled-to-epithelioid cells in alternating hyper- and hypocellular zones, a "patternless pattern," ectatic "staghorn" vessels with variable edematous perivascular stroma, and thick ropey collagen. Cytologically, smears are variably cellular with spindled-to-epithelioid cells with oval nuclei, wispy cytoplasm, multiple inconspicuous nucleoli, and occasional nuclear pseudoinclusions. Small vessels and bare/stripped nuclei are generally present while mild atypia is not uncommon. STAT6 nuclear expression is the most useful immunohistochemical stain and is the product of a NAB2-STAT6 gene fusion. SFTs with mediastinal involvement may be diagnostically challenging due to proximity to vital structures and anticipated patient risks. Endobronchial and endoscopic ultrasound-guided fine-needle aspiration (EBUS/EUS-FNA) are minimally-invasive tissue sampling methods that provide diagnostic material while minimizing patient risk, and the mediastinum is accessible by both procedures. Small aspirate samples and SFT nonspecific features can compound the diagnostic difficulty, although familiarity with the cytologic, morphologic, immunophenotypic, and genetic features of SFTs assist the pathologist in confirming the diagnosis. Pathologists must also be aware of high-risk SFT features to ensure appropriate therapy and management. Case #1 describes a recurrent mediastinal SFT with high-risk features sampled by EUS-FNA. Case #2 describes a primary diagnosis of mediastinal SFT with malignant behavior made on an EBUS-FNA specimen.

Comparison of evaluation techniques, including digital image analysis, for MYC protein expression by immunohistochemical stain in aggressive B-cell lymphomas.

Incorporation of an MYC immunohistochemical stain in the workup of large B-cell lymphomas has become common in hematopathology practice. Evaluation of this stain can be difficult because of staining heterogeneity and can have interobserver variability, particularly when performed on the entire tumor sections. We identified 87 cases of aggressive B-cell lymphoma (34 core needle and 53 excisional biopsies) and compared the following methods of MYC immunohistochemical staining evaluation: the original pathologist's interpretation, a systematic retrospective method of evaluation by manual analysis, and a retrospective method of evaluation by digital image analysis (using scanned slides analyzed via the Aperio Nuclear algorithm). Overall, concordance among these methods was around 80% with κ statistics showing good agreement. However, nearly one-third of our cases had a percent MYC positivity in the 30% to 50% range, and for these cases, concordance among the various methods was marginal/poor. This suggests limited utility as a prognostic or predictive marker using 40% as a cutoff value. In our series, core biopsy specimens were poor predictors of MYC gene rearrangement, and there was no association between MYC immunohistochemical stain and MYC gene gain/amplification. Our retrospective digital image analysis showed strong correlation in MYC percent positivity with our retrospective manual review (correlation coefficient of 0.90) and similar concordance to pathologist interpretation as among pathologists, suggesting that digital image analysis is a viable alternative to manual determination of MYC percent positivity. Digital image analysis provides further opportunities for more sophisticated and standardized scoring systems, which may be helpful in future prognostic/predictive studies.