Azza M. Kamel

Peripheral blood vs bone marrow as a source for allogeneic hematopoietic stem cell transplantation.

In this randomized prospective study, we included 30 patients with different hematological diseases (acute myeloid leukemia, acute lymphoblastic leukemia, chronic myeloid leukemia, myelodysplastic syndrome or severe aplastic anemia) to compare peripheral blood stem cells (PBSC) (15 patients; mean age 23) and bone marrow (BM) (15 patients; mean age 21.8) as a source for allogeneic transplantation regarding the tempo of hematopoietic recovery and the incidence of acute graft-versus-host disease (GVHD). In the BM group, the median nucleated cell count harvested was 1.3 × 1010, while in the PBSC group, the aphereses contained a median of 4.4 × 106 CD34+/kg recipient weight. PBSC transplantation (PBSCT) was associated with faster hematopoietic reconstitution measured as absolute neutrophil count (ANC) >0.5 × 109/l (log-rank P value <0.0018) and platelet count >25 × 109/l (log-rank P value <0.0098). seven patients (46.7%) in the bm group vs only one patient (6.7%) in the PBSC group developed acute GVHD (P = 0.013). Therefore, we conclude that PBSCT is associated with faster hematopoietic recovery and the incidence of acute GVHD does not exceed that seen with BMT.

Rationale for an international consortium to study inherited genetic susceptibility to childhood acute lymphoblastic leukemia.

Acute lymphoblastic leukemia is the major pediatric cancer in developed countries. To date most association studies of acute lymphoblastic leukemia have been based on the candidate gene approach and have evaluated a restricted number of polymorphisms. Such studies have served to highlight difficulties in conducting statistically and methodologically rigorous investigations into acute lymphoblastic leukemia risk. Recent genome-wide association studies of childhood acute lymphoblastic leukemia have provided robust evidence that common variation at four genetic loci confers a modest increase in risk. The accumulated experience to date and relative lack of success of initial efforts to identify novel acute lymphoblastic leukemia predisposition loci emphasize the need for alternative study designs and methods. The International Childhood Acute Lymphoblastic Leukaemia Genetics Consortium includes 12 research groups in Europe, Asia, the Middle East and the Americas engaged in studying the genetics of acute lymphoblastic leukemia. The initial goal of this consortium is to identify and characterize low-penetrance susceptibility variants for acute lymphoblastic leukemia through association-based analyses. Efforts to develop genome-wide association studies of acute lymphoblastic leukemia, in terms of both sample size and single nucleotide polymorphism coverage, and to increase the number of single nucleotide polymorphisms taken forward to large-scale replication should lead to the identification of additional novel risk variants for acute lymphoblastic leukemia. Ethnic differences in the risk of acute lymphoblastic leukemia are well recognized and thus in assessing the interplay between inherited and non-genetic risk factors, analyses using different population cohorts with different incidence rates are likely to be highly informative. Given that the frequency of many acute lymphoblastic leukemia subgroups is small, identifying differential effects will realistically only be possible through multi-center pooled analyses. Here, we review the rationale for identifying genetic risk variants for acute lymphoblastic leukemia and our proposed strategy for establishing the International Childhood Acute Lymphoblastic Leukaemia Genetics Consortium.

Prognostic utility of routine chimerism testing at 2 to 6 months after allogeneic hematopoietic cell transplantation.

The utility of routine chimerism analysis as a prognostic indicator of subsequent outcomes after allogeneic hematopoietic cell transplantation (HCT) with myeloablative conditioning regimens remains controversial. To address this controversy, routine chimerism test results at 2 to 6 months after HCT with myeloablative conditioning regimens were evaluated for association with subsequent risk of chronic graft-versus-host disease (GVHD), nonrelapse mortality (NRM), relapse, and overall mortality. Only 70 of 1304 patients (5%) had < 95% donor-derived cells in the marrow. Low donor chimerism in the marrow occurred more often in patients with low-risk diseases compared with those with higher-risk diseases and was significantly associated with a reduced risk of chronic GVHD. Among 673 patients evaluated, 164 (24%) had < 85% donor-derived T cells in the blood. Low donor T cell chimerism was more frequent in patients with low-risk diseases compared with those with higher-risk diseases, in those who received conditioning with busulfan compared with those who received conditioning with total body irradiation, and in those with lower-grade acute GVHD. Low donor T cell chimerism in the blood was significantly associated with a reduced risk of chronic GVHD but not with a reduced risk of relapse, NRM, or overall mortality. Routine testing of chimerism in the marrow and blood at 2 to 6 months after HCT with myeloablative conditioning regimens may be helpful in documenting engraftment in clinical trials, but provides only limited prognostic information in clinical practice.

Immunological phenotypic pattern of acute lymphoblastic leukaemia in Egypt

We have performed immunophenotyping studies on 186 untreated cases of acute lymphoblastic leukemia (ALL) in an Egyptian population, using panels of monoclonal antibodies (mAb) and an avidin-biotin-immunoperoxidase detection system. Sixty-two of these cases were tested with a panel of mAb directed against the T-cell markers CD2, CD4, CD8, B-cell markers CD20, kappa and lambda, the common ALL antigen (common ALLa) and class II HLA antigens. The remaining 124 cases were also tested with additional markers of T- and B-cell precursors, namely CD7 and CD19. The common leukocyte antigen, T200, was used to exclude nonhemopoietic neoplasms. Cases that remained unclassifiable were further tested with a wider panel of T-cell markers, including CD1, CD2, CD3 and CD5. In some cases multiple mAb directed against the same antigens were used. The relative frequencies of common ALL and B ALL were calculated from the total number of cases and were found to be 39.2% and 3.2%, respectively. The proportions of T-cell and null leukemias were calculated from the better characterized subgroup of 124 cases, and were found to be 50% and 4.8%, respectively. In our series, the age distribution of common ALL revealed a peak at 2-5 yr, but this was partially obscured in the entire series by the high proportion of T-cell cases, which had an age peak between 4 and 12 yr of age. Our results demonstrate marked differences in the phenotypic pattern of ALL in Egypt compared to Western Countries, the predominant finding being a relative excess of T-cell ALL and a paucity of common ALL cases. At present it is not clear whether this results from an increased incidence of T-cell ALL or a decreased incidence of common ALL.

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.

Phenotypic analysis of T-cell acute lymphoblastic leukemia in Egypt.

Leukemic cells from 46 T ALL cases were studied with a wide panel of mAb reacting with T cells using an immunoperoxidase technique. The cases included 15 adults (16 years or over) and 31 children (less than 16 years). The mAb used in the panel were: CD1, (T6), CD2 (T11, X11, D66, clone 2), CD3 (T3/Leu4), CD4, (T4/Leu3a), CD5 (Leu1, T1, A50, I73D9), CD7 (Leu9, I21), CD8 (T8/Leu2a) and HNK1. Based on their reactivity with the mAb panel all cases were assigned to one of the intrathymic differentiation compartments. Among the adults, five cases were assigned to compartment I, six to compartment II and four to compartment III. The pediatric cases included eight in compartment I, eighteen in compartment II and five in compartment III. Fifteen L1 cases studied included four in compartment I, seven in compartment II and four in compartment III; while thirty L2 cases showed nine in compartment I, sixteen in compartment II and five cases in compartment III. The most frequently observed CD groups among T ALL cells were CD5 (100% and 88.9% in children and adults respectively), CD7 (93.1% and 84.6%) and CD2 (76.7% and 76.9%). The most frequently reactive mAb in our series was Leu1 (81.8% followed by I21 (71.1%) and Leu9 (68.9%). The most frequently reactive combination of two mAb was Leu1/I21 (100%) followed by Leu1/T11 (97.4%). Five of the 46 cases reacted with HNK1, suggesting an origin from the natural killer (NK) subset. Our results indicate that in T-cell ALL in Egypt the surface phenotype is similar to that of intermediate or late thymocytes in more than 66% of cases. CD5, CD7 and CD2 were the most frequently detected antigens. Studies of the association between T-cell phenotype and socioeconomic status are warranted.

The prognostic significance of minimal residual disease in adult Egyptian patients with precursor acute lymphoblastic leukemia

BACKGROUND Minimal residual disease (MRD) studies in adult acute lymphoblastic leukemia (ALL) give highly significant prognostic information superior to other standard criteria as age, gender and total leucocytic count (TLC) in distinguishing patients at high and low risk of relapse. OBJECTIVES We aimed to determine the value of MRD monitoring by flowcytometry (FCM) in predicting outcome in adult Precursor ALL patients. PATIENTS AND METHODS Bone marrow (BM) samples were analyzed by 4-color FCM collected at diagnosis and after induction therapy (MRD1) to correlate MRD positivity with disease free survival (DFS) and overall survival (OS). RESULTS Study included 57 adult ALL patients (44 males and 13 females) with a median age of 22 years (18-49). DFS showed no significant difference with age, gender and initial TLC (p=0.838, 0.888 and 0.743, respectively). Cumulative DFS at 2 years was 34% for B-lineage ALL (n: 35) and 57% for T-lineage ALL (n: 18) (p = 0.057). Cumulative DFS at 2 years was 7% for MRD1 positive (high risk, HR) versus 57% for MRD1 negative patients (Low risk, LR) (p < 0.001). Cumulative DFS at 2 years was 29% for HR patients (n: 26) versus 55% for LR (n: 27) according to GMALL classification (p = 0.064). Cumulative OS did not differ according to age, gender and TLC (p = 0.526, 0.594 and 0.513, respectively). Cumulative OS at 2 years was 36% for B ALL (n: 39) versus 77% for TALL (n: 18) (p = 0.016) and was 49% for Philadelphia chromosome (Ph) negative patients versus 0% for Ph-positive patients (p < 0.001). Regarding MRD1, OS at 2 years was 18% for MRD1 HR (n: 17) versus 65% for MRD1 LR (n: 38) (p < 0.001). OS was 35% for high-risk patients (n: 30) and 62% for low-risk patients (n: 27) classified according to GMALL risk stratification (p = 0.017). CONCLUSION MRD by FCM is a strong independent predictor of outcome in terms of DFS and OS and is a powerful informative parameter in guiding individual treatment in ALL patients.

Peripheral blood mammaglobin gene expression for diagnosis and prediction of metastasis in breast cancer patients.

Aim:  To evaluate the value of peripheral blood mammaglobin (MG) gene expression for diagnosis and prediction of metastasis in breast cancer patients.

The effect of killer cell immunoglobulin-like receptor genotype on outcome of hematopoietic stem cell transplantation from matched sibling

The alloreactivity of natural killer (NK) cell after allogeneic hematopoietic stem cell transplantation (AHSCT) is regulated by the interaction between donor killer immunoglobulin-like receptors (KIRs) and recipient human leukocyte antigen (HLA)-class I molecules. The aim was to identify KIR genes, haplotypes and their HLA-class I ligands and to investigate their association with transplantation outcome. The study included 65 patient/donor pairs who received AHSCT from HLA-matched identical siblings. KIR genotyping was done for donors using reverse sequence specific oligonucleotide probes (rSSO) coupled with luminex technology, while HLA-C genotyping was performed in patients using rSSO strip assay. In multivariate analysis, KIR2DS4 was associated with significant reduced incidence of relapse (p = .002). A trend towards reduced incidence of relapse was also observed with more than two KIR B motifs (p = .09), whereas a significant increased relapse was associated with homozygous HLA-C2 ligand compared to combined C1/C2 and C1/C1 (p = .04). Activating KIR2DS3 was associated with rapid leukocyte engraftment (p = .02). While, KIR 2DL5 was associated with decreased CMV infection (p = .03) and better platelets engraftment (p = .05). KIR genes, haplotypes and HLA-C alleles have an impact on HSCT outcome. Better selection of donors with favorable KIR genotype can improve HLA-matched sibling HSCT outcome especially for AML patients.

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

BACKGROUND B-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. AIM OF THE WORK This work was conducted to study adhesion molecules status in CLL and its potential impact on both hematological and clinical parameters. PATIENTS AND METHODS The 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. RESULTS Positive 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). CONCLUSION The level of different adhesion molecules expression in CLL is apparently reflected on the potential migratory behavior of the leukemic cells to different organs.