Flow cytometry detection of neoplastic plasma cells: employing a new statistical model in diagnosing and monitoring Multiple Myeloma

Multiple myeloma (MM) is a clonal B-cell disorder in which malignant plasma cells (PCs) accumulate in the bone marrow (BM), producing lytic lesions, excessive amounts of monoclonal protein in the serum or urine and evidence of end-organ damage (hypercalcemia, renal insufficiency, anemia or bone lesions). A conventional diagnosis in MM is based on a variety of laboratory results: morphology, analysis of M component, haematological features, biochemical parameters, immunophenotyping, cytogenetics, DNA ploidy and labelling index-proliferative activity of PC . Immunophenotypic studies on MM have now been performed for more than 15 years and flow cytometry (FC) represents an attractive approach, not only for research purposes but also in guiding clinical practice. In this sense, FC has many advantages: a) to distinguish among normal, reactive and malignant PC, b) to evaluate the risk of progression from monoclonal gammopathy of unknown significance (MGUS) to MM,, c) to detect prognostic markers,, d) to evaluate minimal residual disease (MRD) and e) to identify new targets for myeloma therapy. It may be difficult to define the clonality of a very small phenotypically abnormal plasma cell population in MRD, both by histology and FC. At this time histology remains the gold standard exam but we here define a new diagnostic model that is objective and reproducible at describing the correlation between phenotype and histology. Based upon our experience we found 8-color FC analysis to be a superior technique for identifying pathologic PC, especially in minimal residual disease (MRD). In a previous study we found that CD19 showed good correlation with the presence of disease using a cut-off of 61% to distinguish between normal and neoplastic PCs. However, this study evaluated the role of each antigen separately in detecting the presence of disease (this study was performed in Massachusetts General Hospital (MGH), Boston. We studied 15 control specimens and 55 patients). In another study (this study was performed in Pisa, at Hematology section, Santa Chiara Hospital, together with the collaboration of MGH. 15 control samples and 177 patients were studied. Patients were from both Santa Chiara Hospital and MGH) we demonstrated that the contribution of different antigens assessed simultaneously improved the correlation with histological results. We described a statistical model where, among all antigens tested, assessing CD19 and CD27 together resulted in the best concordance with histology. Its practical application is simple, rapid and does not require specialized technicians. We propose to use this formula as routine diagnostic tool. It could be used by a simple excel sheet or by a database, where, putting CD19 and CD27 expression values for each patient studied, a value of probability of disease can be obtained. A model plot could also be used for a quick test. A difference value of 0.2 could be used as cutoff of concordance between histology and model. Anyway, although if statistically acceptable, this model was found by analyzing samples collected in just two different laboratories and so, to improve the forecasting efficiency, our next goal will be to perform a multicenter study. Then we would like to evaluate the role that this flow cytometric model could have in evaluating response to therapy, in addition to the actual standardized criteria of evaluation. Thus, a multicenter national and international study, involving 21 Italian laboratories and laboratory of Patholgy of MGH, in Boston, is in progress. Until now, we have collected and analyzed MM samples from Hospital of Lucca. In the last year we have collected 64 MM samples from Lucca Hospital. We have tested the actual statistical model on these cases and we found that it works in 95% of cases. We then recalculated the statistical model, including samples from Lucca to those one from Pisa and Boston, and we found that CD19 (p-value=0.0000) and CD27 (p-value=0.0008) are still the two antigens best correlated with histological results, with an an R2 adjusted of 73,6%, so similar to that one previousely found. On all samples studied (120 from Pisa and Boston and 64 from Lucca), the percentage of concordant cases was 91.3%, with 4.7% discordant cases and 4% uncertain cases. Discordant cases were CD19 and CD27 positive myleoma cases and those cases where a monoclonal plasma cell population was detected together with a polyclonal plasma cell population by flow cytometry. Cases having uncertain results by the model have a difference between the value of the event observed by histology and the value of the event predicted by the model ranging between 0.20 and 0.46. These values are less than 0.50 and so could be considered acceptable, although, to make the model very restrictive, a value of difference between 0 and 0.20 was considered acceptable. This model makes predictions about the probability that the event “presence of disease” will occur for each patient, given the values of the variables CD19 and CD27

Idiopathic thrombocytopenic purpura and coronary artery disease: comparison between coronary artery bypass grafting and percutaneous coronary intervention.

Idiopathic thrombocytopenic purpura (ITP) is an autoimmune disorder characterized by a low platelet count and an increased risk of bleeding. At the same time, ITP patients present an increased risk of thrombosis and atherosclerosis related to the high presence of haemostatic factors and chronic steroid therapy. Although relatively rare, the association of ITP and coronary artery disease represents a complex therapeutic challenge. In particular, no recommendations exist regarding the best management approach. We reviewed the literature making a comparison between coronary artery bypass grafting and percutaneous coronary intervention. © 2011 Published by European Association for Cardio-Thoracic Surgery. All rights reserved

Prethymic cytoplasmic CD3 negative acute lymphoblastic leukemia or acute undifferentiated leukemia: a case report

Acute undiffentiated leukemia (AUL) is an acute leukemia with no more than one membrane marker of any given lineage. Blasts often express HLA-DR, CD34, and/or CD38 and may be positive for terminal deoxynucleotidyl transferase (TdT). The expression of CD34, HLA-DR, and CD38 has been shown in pro-T-ALL, although in this case, blasts should also express CD7 and cyCD3. However, some cases of T-ALL without CD3 in the cytoplasm and all TCR chain genes in germ line configuration are reported, features that fit well with a very early hematopoietic cell. We report a case of acute leukemia CD34+/-HLADR+CD7+CD38+cyCD3- in which a diagnosis of AUL was considered. However the blasts were also positive for CD99 and TCR delta gene rearrangement which was found on molecular studies. Therefore a differential diagnosis between AUL and an early cyCD3 negative T-ALL was debated

Prethymic Cytoplasmic CD3 Negative Acute Lymphoblastic Leukemia or Acute Undifferentiated Leukemia: A Case Report

Acute undiffentiated leukemia (AUL) is an acute leukemia with no more than one membrane marker of any given lineage. Blasts often express HLA-DR, CD34, and/or CD38 and may be positive for terminal deoxynucleotidyl transferase (TdT). The expression of CD34, HLA-DR, and CD38 has been shown in pro-T-ALL, although in this case, blasts should also express CD7 and cyCD3. However, some cases of T-ALL without CD3 in the cytoplasm and all TCR chain genes in germ line configuration are reported, features that fit well with a very early hematopoietic cell. We report a case of acute leukemia CD34+/−HLADR+CD7+CD38+cyCD3− in which a diagnosis of AUL was considered. However the blasts were also positive for CD99 and TCR delta gene rearrangement which was found on molecular studies. Therefore a differential diagnosis between AUL and an early cyCD3 negative T-ALL was debated

Carcinoma and multiple lymphomas in one patient: establishing the diagnoses and analyzing risk factors

Multiple malignancies may occur in the same patient, and a few reports describe cases with multiple hematologic and non-hematologic neoplasms. We report the case of a patient who showed the sequential occurrence of four different lymphoid neoplasms together with a squamous cell carcinoma of the lung. A 62-year-old man with adenopathy was admitted to the hospital, and lymph node biopsy was positive for low-grade follicular lymphoma. He achieved a partial remission with chemotherapy. Two years later, a PET-CT scan showed a left hilar mass in the lung; biopsy showed a squamous cell carcinoma. Simultaneously, he was diagnosed with diffuse large B cell lymphoma in a neck lymph node; after chemo- and radiotherapy, he achieved a complete response. A restaging PET-CT scan 2 years later revealed a retroperitoneal nodule, and biopsy again showed a low-grade follicular lymphoma, while a biopsy of a cutaneous scalp lesion showed a CD30-positive peripheral T cell lymphoma. After some months, a liver biopsy and a right cervical lymph node biopsy showed a CD30-positive peripheral T cell lymphoma consistent with anaplastic lymphoma kinase-negative anaplastic large cell lymphoma. Flow cytometry and cytogenetic and molecular genetic analysis performed at diagnosis and during the patient’s follow-up confirmed the presence of two clonally distinct B cell lymphomas, while the two T cell neoplasms were confirmed to be clonally related. We discuss the relationship between multiple neoplasms occurring in the same patient and the various possible risk factors involved in their development

Acute Delta Hepatitis in Italy spanning three decades (1991–2019): Evidence for the effectiveness of the hepatitis B vaccination campaign

Updated incidence data of acute Delta virus hepatitis (HDV) are lacking worldwide. Our aim was to evaluate incidence of and risk factors for acute HDV in Italy after the introduction of the compulsory vaccination against hepatitis B virus (HBV) in 1991. Data were obtained from the National Surveillance System of acute viral hepatitis (SEIEVA). Independent predictors of HDV were assessed by logistic-regression analysis. The incidence of acute HDV per 1-million population declined from 3.2 cases in 1987 to 0.04 in 2019, parallel to that of acute HBV per 100,000 from 10.0 to 0.39 cases during the same period. The median age of cases increased from 27 years in the decade 1991-1999 to 44 years in the decade 2010-2019 (p < .001). Over the same period, the male/female ratio decreased from 3.8 to 2.1, the proportion of coinfections increased from 55% to 75% (p = .003) and that of HBsAg positive acute hepatitis tested for by IgM anti-HDV linearly decreased from 50.1% to 34.1% (p < .001). People born abroad accounted for 24.6% of cases in 2004-2010 and 32.1% in 2011-2019. In the period 2010-2019, risky sexual behaviour (O.R. 4.2; 95%CI: 1.4-12.8) was the sole independent predictor of acute HDV; conversely intravenous drug use was no longer associated (O.R. 1.25; 95%CI: 0.15-10.22) with this. In conclusion, HBV vaccination was an effective measure to control acute HDV. Intravenous drug use is no longer an efficient mode of HDV spread. Testing for IgM-anti HDV is a grey area requiring alert. Acute HDV in foreigners should be monitored in the years to come

Prethymic Cytoplasmic CD3 Negative Acute Lymphoblastic Leukemia or Acute Undifferentiated Leukemia: A Case Report

Acute undiffentiated leukemia (AUL) is an acute leukemia with no more than one membrane marker of any given lineage. Blasts often express HLA-DR, CD34, and/or CD38 and may be positive for terminal deoxynucleotidyl transferase (TdT). The expression of CD34, HLA-DR, and CD38 has been shown in pro-T-ALL, although in this case, blasts should also express CD7 and cyCD3. However, some cases of T-ALL without CD3 in the cytoplasm and all TCR chain genes in germ line configuration are reported, features that fit well with a very early hematopoietic cell. We report a case of acute leukemia CD34+/−HLADR+CD7+CD38+cyCD3− in which a diagnosis of AUL was considered. However the blasts were also positive for CD99 and TCR delta gene rearrangement which was found on molecular studies. Therefore a differential diagnosis between AUL and an early cyCD3 negative T-ALL was debated

Tailoring p-Type Behavior in ZnO Quantum Dots through Enhanced Sol-Gel Synthesis: Mechanistic Insights into Zinc Vacancies.

The synthesis and control of properties of p-type ZnO is crucial for a variety of optoelectronic and spintronic applications; however, it remains challenging due to the control of intrinsic midgap (defect) states. In this study, we demonstrate a synthetic route to yield colloidal ZnO quantum dots (QD) via an enhanced sol-gel process that effectively eliminates the residual intermediate reaction molecules, which would otherwise weaken the excitonic emission. This process supports the creation of ZnO with p-type properties or compensation of inherited n-type defects, primarily due to zinc vacancies under oxygen-rich conditions. The in-depth analysis of carrier recombination in the midgap across several time scales reveals microsecond carrier lifetimes at room temperature which are expected to occur via zinc vacancy defects, supporting the promoted p-type character of the synthesized ZnO QDs

Abnormal phenotype of bone marrow plasma cells in patients with chronic myeloid leukemia undergoing therapy with Imatinib

Imatinib induces several effects on the immune system, including hypogammaglobulinemia and has been associated with multiple myeloma in some patients. We studied the phenotype of plasma cells from patients with chronic myeloid leukemia (CML) undergoing therapy with Imatinib mesylate (Glivec). Bone marrow samples from 30 CML patients were evaluated and plasma cells were identified by multiparametric flow cytometry. In 21 patients an abnormal plasma cell phenotype, characterized by the absence of CD19, was registered, with 12 patients expressing also the CD56 molecule. A significant correlation between abnormal plasma cell phenotype and reduced gamma-globulin levels was found. Immunofixation was always negative. Therapy with Imatinib for CML seems to induce a plasma cell phenotype with the same characteristics as monoclonal gammapathies. These findings deserve further studies and suggest to monitor plasma protein electrophoresis and gamma-globulin levels in all patients treated with Imatinib