Identification of large genomic rearrangement of BRCA1/2 in high risk patients in Korea.

BACKGROUND: While the majority of germline inactivating mutations in BRCA1/2 are small-scale mutations, large genomic rearrangements (LGRs) are also detected in a variable proportion of patients. However, routine genetic methods are incapable of detecting LGRs, and comprehensive genetic testing algorithm is necessary. METHODS: We performed multiplex ligation-dependent probe amplification assay for small-scale mutation negative patients at high-risk for LGR, based on previously published LGR risk criteria. The inclusion criteria for the high-risk subgroup were personal history of 1) early-onset breast cancer (diagnosed at ≤36 years); 2) two breast primaries; 3) breast cancer diagnosed at any age, with ≥1 close blood relatives (includes first-, second-, or third-degree) with breast and/or epithelial ovarian cancer; 4) both breast and epithelial ovarian cancer diagnosed at any age; and 5) epithelial ovarian cancer with ≥1 close blood relatives with breast and/or epithelial ovarian cancer. RESULTS: Two LGRs were identified. One was a heterozygous deletion of exon 19 and the other was a heterozygous duplication of exon 4-6. The prevalence of LGRs was 7% among Sanger-negative, high-risk patients, and accounted for 13% of all BRCA1 mutations and 2% of all patients. Moreover, LGRs reported in Korean patients, including our 2 newly identified cases, were found exclusively in families with at least one high-risk feature. CONCLUSIONS: Our result suggests that selective LGR screening for Sanger-negative, high-risk patients is necessary for Korean patients.

Pseudozyma aphidis fungaemia with invasive fungal pneumonia in a patient with acute myeloid leukaemia: case report and literature review.

Pseudozyma species rarely cause invasive diseases in humans, which are usually isolated from plants. There have been anecdotal reports regarding Pseudozyma species infections in patients with underlying diseases or in neonates. However, clinical data and the pathogenicity in humans are still insufficient. We experienced a case of Pseudozyma aphidis fungaemia with invasive fungal pneumonia that developed during reinduction chemotherapy in a 51-year-old male with acute myeloid leukaemia (AML). P. aphidis was suspected based on the morphology of the yeast isolated from the blood and was confirmed via rDNA gene sequencing analysis. The patient successfully underwent stem cell transplantation with continuing antifungal treatment and finally completely recovered from both the AML and infectious complications. Here, we report a case of P. aphidis infection that developed during neutropenia in an AML patient and review the global literature.

Using Non-Invasive Respiratory Monitoring for COVID-19 Pulmonary Embolism Diagnosis.

With the high incidence rate of pulmonary embolism (PE) and pneumonia reported in hospitalized patients with COVID-19, the ability to determine the dominant etiology for severe respiratory distress quickly and accurately is crucial to a patient's well-being. Traditionally, D-dimer blood tests and diagnostic imaging studies would be utilized to determine the presence of a PE or a venous thromboembolism. However, COVID-19 places patients in a prothrombotic state and performing diagnostic imaging studies on all patients with COVID-19 would be impractical, making the need for a simple and reliable method to determine the likelihood of PE or venous thromboembolism a priority for emergency departments. The authors believe the use of non-invasive respiratory monitoring technology to assess lung function in hospitalized patients with COVID-19 can aid in discerning the dominant hypoxia etiology and tailoring of their treatment. Here, the authors outline a case and method of using non-invasive respiratory monitoring of lung function in the successful diagnosis of a PE in a 62-year-old patient with COVID-19.

Development of a Novel Flow Cytometry-Based System for White Blood Cell Differential Counts: 10-color LeukoDiff.

BACKGROUND: Flow cytometry (FCM) is commonly used to identify many cell populations. We developed a white blood cell (WBC) differential counting system for detecting abnormal cells using FCM incorporating 10 colors and 11 antibodies in a single tube, called "10-color LeukoDiff," and evaluated its performance. METHODS: Ninety-one EDTA-anti-coagulated peripheral blood samples from 76 patients were analyzed using 10-color LeukoDiff. We compared 10 color LeukoDiff results with the results of manual differential count (manual diff). WBCs were classified into 17 cell populations: neutrophils, total lymphocytes, T lymphocytes, B lymphocytes, CD5 and CD19 co-expressing lymphocytes, natural killer cells, total monocytes, 16+ monocytes, eosinophils, immature granulocytes, basophils, myeloblasts, B-blasts, T-blasts, myeloid antigen-positive B-blasts, CD19- plasma cells, and 19+ plasma cells. RESULTS: The correlations between the 10-color LeukoDiff and manual diff results were strong (r>0.9) for mature neutrophils, lymphocytes, eosinophils, immature granulocytes, and blasts and moderate for monocytes and basophils (r=0.86 and 0.74, respectively). There was no discrepancy in blast detection between 10-color LeukoDiff and manual diff results. Furthermore, 10-color LeukoDiff could differentiate the lineage of the blasts and separately count chronic lymphocytic leukemic cells and multiple myeloma cells. CONCLUSIONS: The 10-color LeukoDiff provided an accurate and comprehensive WBC differential count. The most important ability of 10-color LeukoDiff is to detect blasts accurately. This system is clinically useful, especially for patients with hematologic diseases, such as acute leukemia, chronic lymphocytic leukemia, and multiple myeloma. Application of this system will improve the development of FCM gating strategy designs.

Considerations for monitoring minimal residual disease using immunoglobulin clonality in patients with precursor B-cell lymphoblastic leukemia.

BACKGROUND: Minimal residual disease (MRD) monitoring is a powerful tool to predict the risk of relapse. Herein, we present an MRD monitoring strategy for B-cell lymphoblastic leukemia (B-ALL) using high-throughput sequencing (HTS) of immunoglobulin (Ig) clonality before implementation into routine practice. METHODS: We selected 74 bone marrow (BM) specimens from 47 patients who were diagnosed with B-ALL. Ig clonality was analyzed using both fragment analysis and HTS. The performance of Ig clonality was evaluated through comparison of the results from real-time quantitative polymerase chain reaction (qPCR) of leukemia-specific fusion transcripts and flow cytometry. RESULTS: IGH clonality was observed in all patients, and the sum of clonal burden varied (9.47%-96.77%). IGK clonality was identified in 70% of patients and availed in cases with low IGH clonal burden. The total IGH clonal burden was significantly correlated with the proportion of leukemic blasts, leukemia-specific fusion transcripts, and flow cytometry. We recognized the different responses of each clone and emerging clones originating from the trace of Ig rearrangement presented in the initial specimen. IGH clonal burden after chemotherapy represented patient outcomes well. IGH assay also provided information of repertoire diversity of IGH rearrangement. CONCLUSION: The Ig clonality assay via HTS will be a promising tool for MRD monitoring of B-ALL through an adequate strategy to identify and monitor individual clones and determine repertoire diversity.

Genetic-pathologic characterization of myeloproliferative neoplasms.

Myeloproliferative neoplasms (MPNs) are clonal hematopoietic stem cell disorders characterized by the proliferation of one or more myeloid lineages. The current study demonstrates that three driver mutations were detected in 82.6% of 407 MPNs with a mutation distribution of JAK2 in 275 (67.6%), CALR in 55 (13.5%) and MPL in 6 (1.5%). The mutations were mutually exclusive in principle except in one patient with both CALR and MPL mutations. The driver mutation directed the pathologic features of MPNs, including lineage hyperplasia, laboratory findings and clinical presentation. JAK2-mutated MPN showed erythroid, granulocytic and/or megakaryocytic hyperplasia whereas CALR- and MPL-mutated MPNs displayed granulocytic and/or megakaryocytic hyperplasia. The lineage hyperplasia was closely associated with a higher mutant allele burden and peripheral cytosis. These findings corroborated that the lineage hyperplasia consisted of clonal proliferation of each hematopoietic lineage acquiring driver mutations. Our study has also demonstrated that bone marrow (BM) fibrosis was associated with disease progression. Patients with overt fibrosis (grade ⩾2) presented an increased mutant allele burden (P<0.001), an increase in chromosomal abnormalities (P<0.001) and a poor prognosis (P<0.001). Moreover, among patients with overt fibrosis, all patients with wild-type JAK2/CALR/MPL (triple-negative) showed genomic alterations by genome-wide microarray study and revealed the poorest overall survival, followed by JAK2-mutated MPNs. The genetic-pathologic characteristics provided the information for understanding disease pathogenesis and the progression of MPNs. The prognostic significance of the driver mutation and BM fibrosis suggests the necessity of a prospective therapeutic strategy to improve the clinical outcome.