Breast cancer resistance protein (BCRP) gene expression in a cohort of adult Egyptian patients with acute myeloid leukemia.

BACKGROUND: Acute myeloid leukemia (AML), an aggressive clonal disease, is genetically heterozygous. The prognostic role of expression of Breast Cancer Resistance Protein (BCRP) gene, which behaves as a multidrug transporter, in adult AML is ambiguous. OBJECTIVE: The objective is to assess the level of mRNA expression of BCRP gene in newly diagnosed cytogenetically normal adult Egyptian AML patients; and to clarify its potential influence and association between therapeutic responsiveness and disease free survival. METHODS: The BCRP gene expression was evaluated by quantifying its mRNA using real time RT-PCR in fifty newly diagnosed cytogenetically normal adult AML patients and 20 healthy normal controls. The expression was evaluated in relation to clinical and prognostic factors, response to treatment and the survival rate. RESULTS: BCRP mRNA was over expressed in adult AML patients compared to controls. This study showed a positive statistical correlation between BCRP gene expression and the percent of CD34 expression. Statistical analysis did not reveal any association between BCRP expression level and chemotherapeutic responsiveness or disease free survival rate. CONCLUSION: The significance of BCRP gene expression and its function in AML is very complicated, therefore more standardized clinical studies are needed.

Association of DNA Methyltransferase 3B Promotor Polymorphism With Childhood Chronic Immune Thrombocytopenia.

BACKGROUND: DNA methylation is an epigenetic process that refers to chromatin-based mechanisms in the regulation of gene expression without DNA alternation. It is mediated by DNA methyltransferases (DNMTs). The DNA methyltransferase 3B (DNMT3B) gene contains a C-to-T single nucleotide polymorphism (SNP; rs2424913) in the Promotor region, 149 base pairs from the transcription start site, which is reported to significantly increase the Promotor activity. OBJECTIVE: To investigate the prevalance of rs2424913 single nucleotide polymorphism located in the DNMT3B gene Promotor. METHODS: In the present study, we investigated the prevalence of rs2424913 single nucleotide polymorphism located in DNMT3B gene Promotor by restriction fragment length polymorphism (PCR-RFLP) in Egyptian pediatric chronic immune thrombocytopenia (ITP) patients and controls. RESULTS: The homozygous genotype (TT) was significantly higher in our patient and conferred almost 3-fold increased risk of chronic ITP when compared to controls. CONCLUSION: The present study shows that DNMT3B rs2424913 promotor polymorphism represents a genetic risk factor that may play an important role in understanding the pathogenesis of chronic ITP.

Diagnostic values of CD64, C-reactive protein and procalcitonin in ventilator-associated pneumonia in adult trauma patients: a pilot study.

BACKGROUND: Ventilator-associated pneumonia (VAP) is one of the most common nosocomial infections; however, its diagnosis remains difficult to establish in the critical care setting. We investigated the potential role of neutrophil CD64 (nCD64) expression as an early marker for the diagnosis of VAP. METHODS: Forty-nine consecutive patients with clinically suspected VAP were prospectively included in a single-center study. The levels of nCD64, C-reactive protein (CRP), and serum procalcitonin (PCT) were analyzed for diagnostic evaluation at the time of intubation (baseline), at day 0 (time of diagnosis), and at day 3. The receiver operating characteristic curves were analyzed to identify the ideal cutoff values. RESULTS: VAP was confirmed in 36 of 49 cases. In patients with and without VAP, the median levels (interquartile range, IQR) of nCD64 did not differ either at baseline [2.4 (IQR, 1.8-3.1) and 2.6 (IQR, 2.3-3.2), respectively; p=0.3] or at day 0 [2 (IQR, 2.5-3.0) and 2.6 (IQR, 2.4-2.9), respectively; p=0.8]. CRP showed the largest area under the curve (AUC) at day 3. The optimum cutoff value for CRP according to the maximum Youden index was 133 mg/dL. This cutoff value had 69% sensitivity and 76% specificity for predicting VAP; the AUC was 0.73 (95% CI, 0.59-0.85). The nCD64 and PCT values could not discriminate between the VAP and non-VAP groups either at day 0 or day 3. CONCLUSIONS: The results of this pilot study suggest that neutrophil CD64 measurement has a poor role in facilitating the diagnosis of VAP and thus may not be practically recommended to guide the administration of antibiotics when VAP is suspected.

Interleukin 28B polymorphisms and therapy response in Egyptian hepatitis C genotype-4 patients.

Hepatitis C infection represents a major health problem in Egypt; only 20% of patients undergo spontaneous clearance of the virus and around 25% of all patients progress to develop cirrhosis. More than 90% of Egyptian patients have hepatitis C virus (HCV) genotype-4. Combined pegylated interferon and oral ribavirin are the current standard therapies for HCV-4. The aim of the work is to evaluate the predictive power of the rs12979860 IL28B SNP and rs12980275 IL28B SNP for treatment response in Egyptian patients infected with HCV genotype 4. One hundred eleven HCV patients receiving combined treatment were studied for rs12979860 and rs12980275 polymorphisms by the restriction fragment length polymorphism technique. The rs12979860 CC and rs12979860 AA genotypes were significantly associated with sustained virological response (p=0.001). Our results suggest that studying IL28B polymorphisms contribute to proper prediction of response to standard therapies in Egyptian patients, optimizing cost effectiveness, and minimizing unneeded adverse effect of therapy.

Frequency of expression of RHAMM/CD168 in Egyptian patients with CML.

RHAMM/CD168 is a cell surface receptor for hyaluronan, a glycoaminoglycan that plays a fundamental role in cell growth, differentiation and motility. It is one of the leukemia-associated antigens (LAA) identified in patients with myeloid leukemias. WE AIMED: at studying the frequency of expression of RHAMM/CD168 in Egyptian patients with CML, both in chronic phase and accelerated/blastic phase, as a potential target structure for cellular immunotherapies, and to compare it with available western records. PATIENTS AND METHODS: RHAMM expression was tested by RT-PCR in peripheral blood mononuclear cells of 60 CML patients divided into 2 groups, group A: 44 chronic phase CML patients, group B: 16 accelerated/ blastic phase patients as well as 15 healthy volunteers. OUR RESULTS: Demonstrated that 14/44 (31.8%) of chronic CML patients showed positive RHAMM expression in contrast to 15/16 ( 93.7%) in the accelerated/blastic phase patients. Moreover within the chronic phase patients the RHAMM positive patients had a significantly higher level of bcr-abl/abl ratio. This highlighted the contribution of RHAMM expression with CML disease progression. CONCLUSION: Our work demonstrated a similar proportion of RHAMM expression in both Egyptian and western CML patients. This may pave the way for subsequent studies suggesting the concomitant use of RHAMM R3 peptide vaccination with conventional CML therapy especially in accelerated phase, in order to achieve complete molecular remission for our patient.

Evaluation of bone marrow in 143 lymphomas: the relative frequency and pattern of involvement, secondary myelopathies, pitfalls and diagnostic validity.

UNLABELLED: The aim of the present study is to assess the frequency of bone marrow (BM) involvement by both bone marrow aspirate and biopsy (BMA and BMB, respectively) procedures in established cases of lymphomas at initial presentation, and to study the relative frequency of marrow disease in relation to lymphoma types, patterns of infiltration and the 2ry associated changes, as well as the diagnostic challenges. Moreover, the diagnostic validity of BMA is tested taking the results of the BMB as the true test results, in order to determine the role of each procedure in the diagnostic approach of marrow infiltration. PATIENTS AND METHOD: This is a retrospective study carried out on 143 nonconsecutive Egyptian patients with lymphomas obtained from a private series during the years 2005 to 2008. Criteria of inclusion included the availability of full medical records and material (medical and pathological), patient consent, nodal disease with no therapy prior to BM sampling, except in 7 patients who had another 2nd BMB following therapy. BMA and BMB were performed as part of the routine workup for diagnosis and staging of lymphoma. The patients had a male to female sex ratio of 2.6:1 and a wide age range from 4 to 74 years. RESULTS: In the present series, 64 cases out of the 143 lymphoma patients studied (44.8%) had a BM disease. Involvement was mostly bilateral (80%). Patients older than 40 years showed higher incidence of bone marrow involvement. There was complete concordance (100%) between both diagnostic procedures in the detection of 76 marrow disease-free lymphoma patients. BMA showed no false positive results and a low rate of deference that makes of it an ideal screening test. Three deferred smears of CLL for BMB diagnosis were all positive for involvement. However, in a total number of 64 BMB positive patients, aspirates could only identify lymphoma involvement in 42 lymphoma patients and missed 22 patients with a BM disease, with an overall sensitivity rate of 65.6%. BMB had a high diagnostic viability and is an easily applied reproducible procedure for diagnosis of BM involvement based on a more detailed informative analysis of both architectural and individual cytomorphologic changes. CONCLUSION: The relatively high level of BM involvement in Egyptian lymphoma patients was directly proportional to high-risk factors. The diagnostic validity of BMB is higher than that of BMA. However, BMA serves as a good positive test in screening lymphomas for marrow disease. A negative BMA does not exclude involvement. Thus, smears should be taken as a complimentary procedure.

Evaluation of MIB-1 and p53 overexpression as risk factors in large cell non-Hodgkin lymphoma in adults.

BACKGROUND: Improvement of current results of therapy for large cell non-Hodgkin lymphoma patients can be achieved by optimization of initial treatment or application of risk-adapted therapy. The international prognostic index ( IPI), introduced to identify high-risk patients, was recently criticized because it was based on clinical risk factors only, ignoring important tumor molecular risk factors and it fails to identify a sector of high-risk patients, who ultimately relapse. OBJECTIVE: The aim of this study is to evaluate the value of two tumor biomarkers:MIB-1 and p53 as potential risk factors in diffuse large cell lymphoma. MIB-1 measures tumor cell proliferation, whereas p53 is related to tumor progression and response to chemotherapy. PATIENTS AND METHODS: The study was done on 69 adult patients with diffuse large cell NHL ( 58 B-phenotype and 11 T-phenotype). Clinical risk assessment was determined by the IPI and patients with a score of 3 or more were considered high-risk. Expression of MIB-1 and p53 was determined by immunohistochemistry and nuclear staining was quantitated by image analysis. Immunoexpression was considered high for MIB-1 nuclear count 50% and p53 counts 20%. Evaluation included both response to chemotherapy ( mostly CHOP), as well as 2- year overall survival analysis. RESULTS: The IPI was the only clinical variable which had a significant impact on survival. Overexpression of both MIB-1 and p53 was associated with poor response to treatment, as well as unfavorable survival. Combined risk factor analysis revealed that only MIB-1 was an independent variable. MIB-1 could also identify some high-risk patients previously categorized in the IPI lowrisk group. CONCLUSIONS: MIB-1 is an independent biologic risk factor for large cell NHL. In order to optimize risk assessment of these patients, it is recommended to construct a new prognostic index by adding MIB-1 overexpression to the other clinical factors of standard IPI. This may allow better identification of high-risk patients and help to guide planning of effective initial treatment. Key Words:NHL - MIB-1 - p53 - CHOP - Risk factors.

Ex vivo expansion of stem cells: defining optimum conditions using various cytokines.

With the increasing information on the number, quality, and characteristics of hematopoietic stem cells (HSC) in umbilical cord and placental blood, this material has been found to be efficacious as an alternative source of HSC for transplantation in children. In this study, we sought to define the optimal conditions for ex vivo expansion of cord blood (CB) stem cells. These conditions include: the combinations and concentrations of hematopoietic growth factors (stem cell factor [SCF], granulocyte-macrophage colony-stimulating factor [GM-CSF], interleukin [IL]-3, thrombopoietin [Tpo], IL-6 and Fms-like tyrosine kinase 3 ligand [Flt-3L]), the duration of culture, and the effect of serum supplementation. In this study, 2 protocols were applied for ex vivo expansion of CB stem cells. In protocol I, 20 CB samples were expanded in a static, serum-added, liquid culture for 7 and 11 days using 5 cytokine cocktails. In protocol II, 10 CB samples were expanded for 7 days using cytokines of cocktail 1, with and without IL-6 and Flt-3L, in serum-added and serum-free culture media. This protocol was intended to verify the effect of IL-6, Flt-3L, and the role of serum supplementation in short-term liquid culture. From the present study, it can be concluded that cocktail 1 is the cocktail of choice for ex vivo expansion of CB stem cells in serum-free, liquid culture expanded for 7 days. We can also conclude that culture expanded for 7 days is better than 11 days, as the fold expansion of CD34+ cells was not significantly increased or even decreased in some of the cocktails used. Moreover, the percent of CD95+ cells (apoptotic cells) was significantly increased on day 11 compared to day 7 in the cocktails tested.