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OBJECTIVES: The diagnosis of hematologic malignancies integrates multiple diagnostic and clinical disciplines. Historically, targeted (single-analyte) genetic testing has been used as reflex to initial prescreening by other diagnostic modalities including flow cytometry, anatomic pathology, and clinical cytogenetics. Given the wide range of mutations associated with hematologic malignancies a DNA/RNA-based NGS panel can provide a more effective and economical approach to comprehensive testing of patients as an initial, tier-1 screen. METHODS: Using a cohort of 380 patients, we performed clinical validation of a gene panel designed to assess 40 genes (DNA), and 29 fusion driver genes with over 600 gene fusion partners (RNA), including sample exchange data across three clinical laboratories, and correlation with cytogenetic testing results. RESULTS: The clinical validation of this technology demonstrated that its accuracy, sensitivity, and specificity are comparable to the majority of targeted single-gene approaches, while assessment of the initial patient cohort data demonstrated a high diagnostic yield of 50.5%. CONCLUSIONS: Implementation of a tier-1 NGS-based protocol for gene panel screening provides a comprehensive alternative to targeted molecular testing in patients with suspected hematologic malignancies, with increased diagnostic yield, scalability, reproducibility, and cost effectiveness, making it ideally suited for implementation in clinical laboratories.
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Biomarcadores de Tumor , Neoplasias Hematológicas/diagnóstico , Neoplasias Hematológicas/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Proteínas de Fusión Oncogénica/genética , Biología Computacional/métodos , Predisposición Genética a la Enfermedad , Pruebas Genéticas , Variación Genética , Genómica/métodos , Neoplasias Hematológicas/epidemiología , Humanos , Mutación , Estudios RetrospectivosRESUMEN
Bone marrow aspirate showed diffuse infiltration by a population of monomorphic cells with scant cytoplasm, markedly increased nuclear-to-cytoplasmic ratio, and numerous indistinct nucleoli. Bone marrow biopsy confirmed extensive marrow infiltration by a malignant neoplasm with strong and diffuse expression of synaptophysin by immunohistochemistry, consistent with metastases from Merkel Cell carcinoma.
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This retrospective cross-sectional study investigates the impact of laboratory-specific hemoglobin reference intervals on electronic consultation (eConsult) referral patterns for suspected anemia and elevated hemoglobin at a tertiary care center in London, Ontario that serves Southwestern Ontario. The study analyzed referrals through the Ontario Telemedicine Network's eConsult platform for hemoglobin abnormalities, excluding patients under 18 years old, between July 1, 2019, and June 30, 2023.The main outcome measures were influence of hemoglobin reference intervals on the referral patterns for suspected anemia and elevated hemoglobin, as well as the extent of pre-referral laboratory testing. Of the 619 eConsults reviewed, 251 referrals for suspected anemia and 93 for elevated hemoglobin were analyzed. Referral patterns showed significant variance in hemoglobin levels based on different laboratory thresholds. Referrals for suspected anemia in females from laboratories whose lower limit was 120 g/L or greater had a hemoglobin concentration 7.5 g/L greater than referrals that used laboratories with a threshold lower than 120 g/L. The study also identified potential areas for improvement in pre-referral investigations; 44% of eConsults did not provide a ferritin level, 53% were missing a B12 level, and 81% were missing a reticulocyte count. In conclusion, laboratory reference intervals for hemoglobin significantly influence referral patterns for suspected hemoglobin abnormalities in Ontario's eConsult system. There is a need for standardized reference intervals and comprehensive pre-referral testing to avoid unnecessary medicalization and referrals. We propose an anemia management algorithm to guide primary care providers in the pre-referral investigation process.
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Flow cytometry (FC) has proven to be an extremely versatile and useful tool in the diagnosis and monitoring of hematological diseases in addition to numerous other applications. Major advances in electronics, software, and reagents over the past years have simplified some aspects of FC, while at the same time the ability to combine 8-10 antibodies in a single tube can create both technical and interpretation issues that are more difficult to detect when using only 3-4 color combinations. Use of multiparameter panels can facilitate identification of abnormal populations; however, characteristics of the neoplastic population may create potential diagnostic pitfalls. An understanding of normal immunophenotypic patterns in states of rest, recovery, and activation is a critical first step in order to appropriately identify the abnormal populations that characterize hematopoietic neoplasms. Additionally, incorporation of newer therapeutic strategies, in particular targeted therapies, can confound standard methods for flow cytometric data analysis and knowledge of the impact of therapy on flow cytometric data is critical for accurate data interpretation. This manuscript will review preanalytical, instrument, and interpretation issues that may lead to incorrect interpretation of results.
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Citometría de Flujo , Enfermedades Hematológicas/diagnóstico , HumanosRESUMEN
Paroxysmal nocturnal hemoglobinuria (PNH) is an uncommon disorder characterized by hemolysis, thrombosis and marrow failure. Whereas venous and arterial thrombosis is a very common symptom of the disease, the frequency of PNH clones in patients with unexplained venous thromboembolism, including deep vein thrombosis and pulmonary embolism, has not been studied. We conducted a cross sectional study evaluating the presence of PNH clones in patients with prevalent venous thromboembolism using a high sensitivity flow cytometry assay for erythrocytes and neutrophils. Among the 388 patients enrolled in the study one patient had a detectable PNH clone of 0.02% in the neutrophil population (0.26%; 95% CI 0.05 to 1.45) and no detectable erythrocyte clone. We conclude that the presence of PNH clones in patients with idiopathic venous thrombosis is rare. Screening for PNH clones among VTE patients might be better reserved for patients with signs of hemolysis.