Nahla El-Sharkawy

Impact of CD34 subsets on engraftment kinetics in allogeneic peripheral blood stem cell transplantation

Summary:Our objective was to evaluate, probably for the first time, the impact of CD34 subsets on engraftment kinetics in allogeneic PBSC transplantation (PBSCT). PBSC graft components were analyzed in 62 cases for the absolute count/kg of total CD34+ and the following subsets: DR− and +, CD71+/−, CD38+/−, CD33+/− and CD61+/−. Time to ANC >0.5 and >1 × 109/l and platelets >20 and >50 × 109/l was reported. The median value for each parameter was used to discriminate rapid from slow engraftment. Four parameters showed significant predictive power of early neutrophil engraftment, namely CD34+/DR− (P=0.002), CD34+/38− (P=0.02), CD34+/CD61− (P=0.04) and total CD34+ cell dose (P=0.04). Four parameters showed significant predictive power of early platelet engraftment, namely CD34+/CD61+ (P=0.02), CD34+/CD38− and total CD34+ cell dose (P=0.04) and CD34+/CD71− (P=0.05). Comparing patients who received >to those who received < the threshold dose(s), only CD34+/CD38− lost its significance for neutrophil engraftment; and only CD34+/CD61+ retained its significance for platelet engraftment (P=0.03); furthermore, the former group required significantly fewer platelet transfusions (P=0.018). We concluded that in allogeneic PBSCT, the best predictor of early neutrophil engraftment is the absolute CD34+/DR− and for early platelet engraftment is the absolute CD34+/CD61+ cell dose.

Flow cytometric platelet cross-matching to predict platelet transfusion in acute leukemia

A great variety of patient‐ and product‐related factors influence the outcome of platelet transfusions. Our study assessed the predictive value of a flow cytometric platelet cross match test for the outcome of HLA matched and unmatched platelet transfusions in patients with acute leukemia. Thirty nine patients (26 adults and 13 children) received 60 ABO compatible apheresis platelet unites ranging from 1 to 4 per patient (mean = 1.54; median = 2). We performed flowcytometric platelet cross‐matching, HLA Class I typing by sequence‐specific primer (SSP) for patients and complement‐dependent cytotoxicity (CDC) for donors and screening of HLA Class I antibodies by ELISA. Effectiveness of platelet transfusion was evaluated using the corrected count increment which was calculated at 60 min and 18‐ to 24‐h posttransfusion. Multivariate analysis was performed to detect which variable can predict transfusion response more than others. Cross‐matched platelet transfusions associated with good response in 51.4% of transfusion events in adults and 73.3% in children. The noncrossmatched platelet transfusions associated with poor response in 83.3% in adults and 100% in children (P‐values 0.143, 0.041, respectively). In the presence of clinical factors or HLA alloimmunization in adults, cross‐matched platelets were associated with good response in 29.6 and 22.2% respectively. In children this occurred in 81.8 and 66.7%, respectively. In presence or absence of HLA matching, flow cytometry platelet cross‐matching was the most predictor for transfusion response (P = 0.05). Because of the difficulties to find frequent HLA matched donors for acute leukemia patients; flow cytometric platelet cross‐matching may provide the most useful way for selecting donors. It is useful even in the presence of alloimunization in children. J. Clin. Apheresis, 2011.

Induction and repair of DNA double-strand breaks using constant-field gel electrophoresis and apoptosis as predictive markers for sensitivity of cancer cells to cisplatin

This study was designed to evaluate some parameters that may play a role in the prediction of cancer cells sensitivity to cisplatin (CIS). Sensitivity, induction and repair of DNA double-strand breaks (DSB), cell cycle regulation and induction of apoptosis were measured in four cancer cell lines with different sensitivities to CIS. Using a sulphorhodamine-B assay, the cervical carcinoma cells (HeLa) were found to be the most sensitive to CIS followed by breast carcinoma cells (MCF-7) and liver carcinoma cells (HepG2). Colon carcinoma HCT116 cells were the most resistant. As measured by constant-field gel electrophoresis (CFGE), DSB induction, but not residual DSB exhibited a significant correlation with the sensitivity of cells to CIS. Flow cytometric DNA ploidy analysis revealed that 67% of HeLa cells and 10% of MCF-7 cells shift to sub-G1 phase after incubation with CIS. Additionally, CIS induced the arrest of MCF-7 cells in S-phase and the arrest of HepG2 and HCT116 cells in both S phase and G2/M phase. Determination of the Fas-L level and Caspase-9 activity indicated that CIS-induced apoptosis results from the mitochondrial (intrinsic) pathway. These results, if confirmed using clinical samples, indicate that the induction of DNA DSB as measured by CFGE and the induction of apoptosis should be considered, along with other predictive markers, in future clinical trials to develop predictive assays for platinum -based therapy.

Adhesion molecules expression in CLL: Potential impact on clinical and hematological parameters.

BACKGROUNDnB-cell chronic lymphocytic leukemia (CLL) is marked by the accumulation of CD5+ B lymphocytes within the blood, bone marrow (BM), and secondary lymphoid tissues. Abnormalities in the expression and function of cell adhesion molecules may account for the patterns of intra-nodal growth and hematogenous spread of the malignant cells. Chemokines and integrin-mediated adhesion and trans-endothelial migration (TEM) are central aspects in trafficking and retention of hematopoietic cells in the BM and lymphoid organs.nnnAIM OF THE WORKnThis work was conducted to study adhesion molecules status in CLL and its potential impact on both hematological and clinical parameters.nnnPATIENTS AND METHODSnThe study included 78 newly diagnosed CLL patients. Immunophenotyping was performed on peripheral blood using the chronic lymphoid panel. Adhesion molecules (CD11a, CD11b, CD49d, CD49C, CD29 and CD38) were tested using monoclonal antibodies and analyzed by Flow Cytometry.nnnRESULTSnPositive correlation was encountered between adhesion molecules: CD38 with CD49d (r=0.25, p=0.028), CD11a with CD11b, CD49d and CD29 (r=0.394, p=0.001; r=0.441, p=<0.01 and r=0.446, p<0.01 respectively) and CD29 with CD49c and CD49d (r=0.437, p<0.01; r=0.674, p<0.01 respectively). CD49c showed negative correlation with Rai staging (r=-0.269, p=0.033). CD11a and CD29 showed a significant relation with splenomegaly (p=0.04 and 0.03 respectively) and CD49d showed a significant relation with lymphadenopathy (p=0.02).nnnCONCLUSIONnThe level of different adhesion molecules expression in CLL is apparently reflected on the potential migratory behavior of the leukemic cells to different organs.

Impact of FLT3 Receptor (CD135) Detection by Flow Cytometry on Clinical Outcome of Adult Acute Myeloid Leukemia Patients

Micro‐Abstract In a large study focusing on the impact of CD135 on clincal presentation and its relation to other prognostic parameters, we addressed whether CD135 could be predictor of FMS‐like tyrosine kinase 3 (FLT3) mutation. We found an association between CD135 expression and persistence of positive minimal residual disease postinduction status. CD135 is a crucial indicator of unfavorable outcome that influences complete remission status, disease‐free survival, and overall survival in adult Egyptian acute myeloid leukeia patients. Background The significance of FMS‐like tyrosine kinase 3 (FLT3)‐ITD mutation in acute myeloid leukemia (AML) prognosis has been well established. The aims of this study were to investigate the prognostic impact of the FLT3 protein (CD135) expression and its association with FLT3‐ITD mutation, and to identify its role in minimal residual disease. Patients and Methods CD135 was measured by flow cytometry on leukemic blasts of 257 adults with de novo AML. High expression of CD135 ≥ 20% was correlated with clinical, laboratory, and other prognostic factors that influenced treatment outcome. FLT3‐ITD mutation was tested by PCR. Results The frequency of CD135 expression was 138 (53.7%) of 257. FLT3‐ITD was detected in (21.4%). Positive CD135 expression was associated with high total leukocyte count (P = .006), platelet count (P = .003), monocytic leukemia (P < .001), and CD34 (P = .008) and CD117 (P = .006) expression. CD135 expression ≥ 25% was a predictor of FLT3‐ITD mutation (P = .03). CD135 overexpression was a negative predictor of complete remission and of postinduction minimal residual disease at days 14 and 28 (P < .001). CD135 had an adverse impact on overall and disease‐free survival (68.5% vs. 15%, P = .002). Multivariate analysis indicated CD135 was the sole independent prognostic factor for overall survival (hazard ratio = 2.49; 95% confidence interval, 1.855‐3.345; P < .001). Conclusion CD135 is emerging as a prognostic factor, a new marker for minimal residual disease, and a potential novel therapeutic target of AML.

Increased expression of brother of the regulator of imprinted sites in peripheral blood neutrophils is associated with both benign and malignant breast lesions

BORIS, a paralog of the multifunctional CCCTC‐binding factor (CTCF) gene is restricted to testis and normally not present in females. It is aberrantly activated in various human cancers including cancer breast. Using immunohistochemistry, western blot and/or RT‐PCR, significantly higher levels of BORIS expression were reported in the neutrophils of cancer breast patients. We hypothesized that Flow Cytometry might be a better technique for objective quantitative evaluation of BORIS in neutrophils and we wanted to investigate if BORIS would discriminate between benign and malignant breast lesions.

Hematogones as a Prognostic Indicator in Allogeneic Hematopoietic Stem Cell Transplantation: Single Center Experience

withdrawal and early start of empirical antibiotics, receive antifungal and antiviral or G-CSF according to guidelines. Main Outcomes Measures: To study the important factors that predicts the risk of mortality. Results: The different modalities used for the diagnosis of infection, factors affecting prognosis were analyzed and response to antibiotics were analysed. The most frequent diagnosis in our study were AML, followed by ALL, NHL, and others (HL, MM, CLL, CML bc). The disease status of hematological malignancy patient was found highly significant and affects the control of NF episode. The more the patient develop neutropenic episodes the more the risk of mortality increase. Our study had proven the significance of MASCC score for predicting mortality in hemato-oncology patients. Sixty-two % of the identified pathogens were gram positive while gram negative bacteria represented 38%. In the blood culture, gram positive bacteria were identified in 80% of cases. Gram negative bacteria were the commonest pathogens identified by other diagnostic modalities (19/28, 68%). To develop an Egyptian prognostic system for NF patient a multivariate analysis were conducted between the most significant identified risk factors as shown in table 1. Conclusions: The management of febrile neutropenia in hematological malignant patients requires further attention to many other risk factors. Prophylactic and empiric antibiotics should be modified according to the pattern of infection, common pathogens and drug sensitivity. The choice of antibiotics should depend on local department policy. Several procedures should be done to avoid emergence of drug resistance organism.