Normal Hematopoietic Progenitor Subsets Have Distinct Reactive Oxygen Species, BCL2 and Cell-Cycle Profiles That Are Decoupled from Maturation in Acute Myeloid Leukemia

In acute myeloid leukemia (AML) quiescence and low oxidative state, linked to BCL2 mitochondrial regulation, endow leukemic stem cells (LSC) with treatment-resistance. LSC in CD34+ and more mature CD34− AML have heterogeneous immunophenotypes overlapping with normal stem/progenitor cells (SPC) but may be differentiated by functional markers. We therefore investigated the oxidative/reactive oxygen species (ROS) profile, its relationship with cell-cycle/BCL2 for normal SPC, and whether altered in AML and myelodysplasia (MDS). In control BM (n = 24), ROS levels were highest in granulocyte-macrophage progenitors (GMP) and CD34− myeloid precursors but megakaryocyte-erythroid progenitors had equivalent levels to CD34+CD38low immature-SPC although they were ki67high. BCL2 upregulation was specific to GMPs. This profile was also observed for CD34+SPC in MDS-without-excess-blasts (MDS-noEB, n = 12). Erythroid CD34− precursors were, however, abnormally ROS-high in MDS-noEB, potentially linking oxidative stress to cell loss. In pre-treatment AML (n = 93) and MDS-with-excess-blasts (MDS-RAEB) (n = 14), immunophenotypic mature-SPC had similar ROS levels to co-existing immature-SPC. However ROS levels varied between AMLs; Flt3ITD+/NPM1wild-type CD34+SPC had higher ROS than NPM1mutated CD34+ or CD34− SPC. An aberrant ki67lowBCL2high immunophenotype was observed in CD34+AML (most prominent in Flt3ITD AMLs) but also in CD34− AMLs and MDS-RAEB, suggesting a shared redox/pro-survival adaptation. Some patients had BCL2 overexpression in CD34+ ROS-high as well as ROS-low fractions which may be indicative of poor early response to standard chemotherapy. Thus normal SPC subsets have distinct ROS, cell-cycle, BCL2 profiles that in AML /MDS-RAEB are decoupled from maturation. The combined profile of these functional properties in AML subpopulations may be relevant to differential treatment resistance

Molecular Pathogenesis of Secondary Acute Promyelocytic Leukemia

Balanced chromosomal translocations that generate chimeric oncoproteins are considered to be initiating lesions in the pathogenesis of acute myeloid leukemia. The most frequent is the t(15;17)(q22;q21), which fuses the PML and RARA genes, giving rise to acute promyelocytic leukemia (APL). An increasing proportion of APL cases are therapy-related (t-APL), which develop following exposure to radiotherapy and/or chemotherapeutic agents that target DNA topoisomerase II (topoII), particularly mitoxantrone and epirubicin. To gain insights into molecular mechanisms underlying the formation of the t(15;17) we mapped the translocation breakpoints in a series of t-APLs, which revealed significant clustering according to the nature of the drug exposure. Remarkably, in approximately half of t-APL cases arising following mitoxantrone treatment for breast cancer or multiple sclerosis, the chromosome 15 breakpoint fell within an 8-bp “hotspot” region in PML intron 6, which was confirmed to be a preferential site of topoII-mediated DNA cleavage induced by mitoxantrone. Chromosome 15 breakpoints falling outside the “hotspot”, and the corresponding RARA breakpoints were also shown to be functional topoII cleavage sites. The observation that particular regions of the PML and RARA loci are susceptible to topoII-mediated DNA damage induced by epirubicin and mitoxantrone may underlie the propensity of these agents to cause APL

National policy development for cotrimoxazole prophylaxis in Malawi, Uganda and Zambia: the relationship between Context, Evidence and Links

BACKGROUND: Several frameworks have been constructed to analyse the factors which influence and shape the uptake of evidence into policy processes in resource poor settings, yet empirical analyses of health policy making in these settings are relatively rare. National policy making for cotrimoxazole (trimethoprim-sulfamethoxazole) preventive therapy in developing countries offers a pertinent case for the application of a policy analysis lens. The provision of cotrimoxazole as a prophylaxis is an inexpensive and highly efficacious preventative intervention in HIV infected individuals, reducing both morbidity and mortality among adults and children with HIV/AIDS, yet evidence suggests that it has not been quickly or evenly scaled-up in resource poor settings. METHODS: Comparative analysis was conducted in Malawi, Uganda and Zambia, using the case study approach. We applied the 'RAPID' framework developed by the Overseas Development Institute (ODI), and conducted a total of 47 in-depth interviews across the three countries to examine the influence of context (including the influence of donor agencies), evidence (both local and international), and the links between researcher, policy makers and those seeking to influence the policy process. RESULTS: Each area of analysis was found to have an influence on the creation of national policy on cotrimoxazole preventive therapy (CPT) in all three countries. In relation to context, the following were found to be influential: government structures and their focus, donor interest and involvement, healthcare infrastructure and other uses of cotrimoxazole and related drugs in the country. In terms of the nature of the evidence, we found that how policy makers perceived the strength of evidence behind international recommendations was crucial (if evidence was considered weak then the recommendations were rejected). Further, local operational research results seem to have been taken up more quickly, while randomised controlled trials (the gold standard of clinical research) was not necessarily translated into policy so swiftly. Finally the links between different research and policy actors were of critical importance, with overlaps between researcher and policy maker networks crucial to facilitate knowledge transfer. Within these networks, in each country the policy development process relied on a powerful policy entrepreneur who helped get cotrimoxazole preventive therapy onto the policy agenda. CONCLUSIONS: This analysis underscores the importance of considering national level variables in the explanation of the uptake of evidence into national policy settings, and recognising how local policy makers interpret international evidence. Local priorities, the ways in which evidence was interpreted, and the nature of the links between policy makers and researchers could either drive or stall the policy process. Developing the understanding of these processes enables the explanation of the use (or non-use) of evidence in policy making, and potentially may help to shape future strategies to bridge the research-policy gaps and ultimately improve the uptake of evidence in decision making

Trafficking of storage proteins in developing grain of wheat

The processing properties of the wheat flour are largely determined by the structures and interactions of the grain storage proteins (also called gluten proteins) which form a continuous visco-elastic network in dough. Wheat gluten proteins are classically divided into two groups, the monomeric gliadins and the polymeric glutenins, with the latter being further classified into low molecular weight (LMW) and high molecular weight (HMW) subunits. The synthesis, folding and deposition of the gluten proteins take place within the endomembrane system of the plant cell. However, determination of the precise routes of trafficking and deposition of individual gluten proteins in developing wheat grain has been limited in the past by the difficulty of developing monospecific antibodies. To overcome this limitation, a single gluten protein (a LMW subunit) was expressed in transgenic wheat with a C-terminal epitope tag, allowing the protein to be located in the cells of the developing grain using highly specific antibodies. This approach was also combined with the use of wider specificity antibodies to compare the trafficking and deposition of different gluten protein groups within the same endosperm cells. These studies are in agreement with previous suggestions that two trafficking pathways occur in wheat, with the proteins either being transported via the Golgi apparatus into the vacuole or accumulating directly within the lumen of the ER. They also suggest that the same individual protein could be trafficked by either pathway, possibly depending on the stage of development, and that segregation of gluten proteins both between and within protein bodies may occur

Treatment outcomes of new tuberculosis patients hospitalized in Kampala, Uganda: a prospective cohort study.

BACKGROUND: In most resource limited settings, new tuberculosis (TB) patients are usually treated as outpatients. We sought to investigate the reasons for hospitalisation and the predictors of poor treatment outcomes and mortality in a cohort of hospitalized new TB patients in Kampala, Uganda. METHODS AND FINDINGS: Ninety-six new TB patients hospitalised between 2003 and 2006 were enrolled and followed for two years. Thirty two were HIV-uninfected and 64 were HIV-infected. Among the HIV-uninfected, the commonest reasons for hospitalization were low Karnofsky score (47%) and need for diagnostic evaluation (25%). HIV-infected patients were commonly hospitalized due to low Karnofsky score (72%), concurrent illness (16%) and diagnostic evaluation (14%). Eleven HIV uninfected patients died (mortality rate 19.7 per 100 person-years) while 41 deaths occurred among the HIV-infected patients (mortality rate 46.9 per 100 person years). In all patients an unsuccessful treatment outcome (treatment failure, death during the treatment period or an unknown outcome) was associated with duration of TB symptoms, with the odds of an unsuccessful outcome decreasing with increasing duration. Among HIV-infected patients, an unsuccessful treatment outcome was also associated with male sex (P = 0.004) and age (P = 0.034). Low Karnofsky score (aHR = 8.93, 95% CI 1.88 - 42.40, P = 0.001) was the only factor significantly associated with mortality among the HIV-uninfected. Mortality among the HIV-infected was associated with the composite variable of CD4 and ART use, with patients with baseline CD4 below 200 cells/µL who were not on ART at a greater risk of death than those who were on ART, and low Karnofsky score (aHR = 2.02, 95% CI 1.02 - 4.01, P = 0.045). CONCLUSION: Poor health status is a common cause of hospitalisation for new TB patients. Mortality in this study was very high and associated with advanced HIV Disease and no use of ART

Diagnosis of acute myeloid leukemia according to the WHO classification in the Japan Adult Leukemia Study Group AML-97 protocol

We reviewed and categorized 638 of 809 patients who were registered in the Japan Adult Leukemia Study Group acute myeloid leukemia (AML)-97 protocol using morphological means. Patients with the M3 subtype were excluded from the study group. According to the WHO classification, 171 patients (26.8%) had AML with recurrent genetic abnormalities, 133 (20.8%) had AML with multilineage dysplasia (MLD), 331 (51.9%) had AML not otherwise categorized, and 3 (0.5%) had acute leukemia of ambiguous lineage. The platelet count was higher and the rate of myeloperoxidase (MPO)-positive blasts was lower in AML with MLD than in the other WHO categories. The outcome was significantly better in patients with high (≥50%) than with low (<50%) ratios of MPO-positive blasts (P < 0.01). The 5-year survival rates for patients with favorable, intermediate, and adverse karyotypes were 63.4, 39.1, and 0.0%, respectively, and 35.5% for those with 11q23 abnormalities (P < 0.0001). Overall survival (OS) did not significantly differ between nine patients with t(9;11) and 23 with other 11q23 abnormalities (P = 0.22). Our results confirmed that the cytogenetic profile, MLD phenotype, and MPO-positivity of blasts are associated with survival in patients with AML, and showed that each category had the characteristics of the WHO classification such as incidence, clinical features, and OS

Operational research in Malawi: making a difference with cotrimoxazole preventive therapy in patients with tuberculosis and HIV.

BACKGROUND: In Malawi, high case fatality rates in patients with tuberculosis, who were also co-infected with HIV, and high early death rates in people living with HIV during the initiation of antiretroviral treatment (ART) adversely impacted on treatment outcomes for the national tuberculosis and ART programmes respectively. This article i) discusses the operational research that was conducted in the country on cotrimoxazole preventive therapy, ii) outlines the steps that were taken to translate these findings into national policy and practice, iii) shows how the implementation of cotrimoxazole preventive therapy for both TB patients and HIV-infected patients starting ART was associated with reduced death rates, and iv) highlights lessons that can be learnt for other settings and interventions. DISCUSSION: District and facility-based operational research was undertaken between 1999 and 2005 to assess the effectiveness of cotrimoxazole preventive therapy in reducing death rates in TB patients and subsequently in patients starting ART under routine programme conditions. Studies demonstrated significant reductions in case fatality in HIV-infected TB patients receiving cotrimoxazole and in HIV-infected patients about to start ART. Following the completion of research, the findings were rapidly disseminated nationally at stakeholder meetings convened by the Ministry of Health and internationally through conferences and peer-reviewed scientific publications. The Ministry of Health made policy changes based on the available evidence, following which there was countrywide distribution of the updated policy and guidelines. Policy was rapidly moved to practice with the development of monitoring tools, drug procurement and training packages. National programme performance improved which showed a significant decrease in case fatality rates in TB patients as well as a reduction in early death in people with HIV starting ART. SUMMARY: Key lessons for moving this research endeavour through to policy and practice were the importance of placing operational research within the programme, defining relevant questions, obtaining "buy-in" from national programme staff at the beginning of projects and having key actors or "policy entrepreneurs" to push forward the policy-making process. Ultimately, any change in policy and practice has to benefit patients, and the ultimate judge of success is whether treatment outcomes improve or not

Human immunodeficiency virus infection and cerebral malaria in children in Uganda: a case-control study

<p>Abstract</p> <p>Background</p> <p>Human immunodeficiency virus (HIV)-1 infection increases the burden of malaria by increasing susceptibility to infection and decreasing the response to malarial treatment. HIV-1 has also been found to suppress the immune system and predispose to severe forms of malaria in adults. There is still a paucity of data on the association between HIV-1 infection and cerebral malaria in children. The aim of this study was to determine whether HIV-1 infection is a risk factor for cerebral malaria in children.</p> <p>Method</p> <p>We conducted an unmatched case-control study, in which 100 children with cerebral malaria were compared with 132 with uncomplicated malaria and 120 with no malaria. In stratified analyses we estimated odds ratios (ORs) and 95% confidence intervals (CIs) adjusted for age.</p> <p>Results</p> <p>HIV-1 infection was present in 9% of children with cerebral malaria compared to 2.3% in uncomplicated malaria (age-adjusted odds ratio (aOR) 5.94 (95% confidence interval (CI) 1.36-25.94, p = 0.012); and 2.5% in children with no malaria (aOR 3.85 (95% CI0.99-14.93, p = 0.037). The age-adjusted odds of being HIV-positive among children with cerebral malaria compared to the control groups (children with uncomplicated malaria and no malaria) was 4.98 (95% CI 1.54-16.07), p-value = 0.003.</p> <p>Conclusions</p> <p>HIV-1 infection is associated with clinical presentation of cerebral malaria in children. Clinicians should ensure that children diagnosed with HIV infection are initiated on cotrimoxazole prophylaxis as soon as the diagnosis is made and caretakers counselled on the importance of adherence to the cotrimoxazole towards reducing the risk of acquiring <it>P.falciparum </it>malaria and associated complications such as cerebral malaria. Other malaria preventive measures such as use of insecticide-treated mosquito nets should also be emphasized during counselling sessions.</p