Secondary education reform in Lesotho and Zimbabwe and the needs of rural girls: Pronouncements, policy and practice

Analysis of the educational needs of rural girls in Lesotho and Zimbabwe suggests a number of shortcomings in the current form of secondary education, and ways in which it might be modified so as to serve this sizeable group of students better. Several of the shortcomings, notably in relation to curricular irrelevance and excessive focus on examinations, have long been recognised, including by politicians. Yet political pronouncements are seldom translated into policy, and even where policy is formulated, reforms are seldom implemented in schools. This paper makes use of interviews with educational decision-makers in the two southern African countries and a range of documentary sources to explore why, despite the considerable differences between the two contexts, much needed educational reforms have been implemented in neither

One Scaffold, Three Binding Modes: Novel and Selective Pteridine Reductase 1 Inhibitors Derived from Fragment Hits Discovered by Virtual Screening†

The enzyme pteridine reductase 1 (PTR1) is a potential target for new compounds to treat human African trypanosomiasis. A virtual screening campaign for fragments inhibiting PTR1 was carried out. Two novel chemical series were identified containing aminobenzothiazole and aminobenzimidazole scaffolds, respectively. One of the hits (2-amino-6-chloro-benzimidazole) was subjected to crystal structure analysis and a high resolution crystal structure in complex with PTR1 was obtained, confirming the predicted binding mode. However, the crystal structures of two analogues (2-amino-benzimidazole and 1-(3,4-dichloro-benzyl)-2-amino-benzimidazole) in complex with PTR1 revealed two alternative binding modes. In these complexes, previously unobserved protein movements and water-mediated protein-ligand contacts occurred, which prohibited a correct prediction of the binding modes. On the basis of the alternative bindingmode of 1-(3,4-dichloro-benzyl)-2-amino-benzimidazole, derivatives were designed and selective PTR1 inhibitors with low nanomolar potency and favorable physicochemical properties were obtained

2,4-Diaminopyrimidines as Potent Inhibitors of Trypanosoma brucei and Identification of Molecular Targets by a Chemical Proteomics Approach

The protozoan parasite Trypanosoma brucei is the causative agent of human African trypanosomiasis (HAT) or sleeping sickness, a fatal disease affecting nearly half a million people in sub-Saharan Africa. Current treatments for HAT have very poor safety profiles and are difficult to administer. There is an urgent need for new, safe and effective treatments for sleeping sickness. This work describes the discovery of 2,4-diaminopyrimidines, exemplified by 4-[4-amino-5-(2-methoxy-benzoyl)-pyrimidin-2-ylamino]-piperidine-1-carboxylic acid phenylamide or SCYX-5070, as potent inhibitors of T. brucei growth in vitro and also in animal models for HAT. To determine the parasite proteins responsible for interaction with SCYX-5070 and related compounds, affinity pull-downs were performed followed by sequence analysis and parasite genome database searching. The work revealed that mitogen-activated protein kinases (MAPKs) and cdc2-related kinases (CRKs) are the major proteins specifically bound to the immobilized compound, suggesting their potential participation in the pharmacological effects of 2,4-diaminopyrimidines against trypanosomatid protozoan parasites. These data strongly support the use of 2,4-diminipyrimidines as leads for the development of new drug candidates for the treatment of HAT

Tolerance to drug-induced cell death favours the acquisition of multidrug resistance in Leishmania

The control of the protozoan parasite Leishmania relies on few drugs with unknown cellular targets and unclear mode of action. Several antileishmanials, however, were shown to induce apoptosis in Leishmania and this death mechanism was further studied in drug-sensitive and drug-resistant Leishmania infantum. In sensitive parasites, antimonials (SbIII), miltefosine (MF) and amphotericin B (AMB), but not paromomycin (PARO), triggered apoptotic cell death associated with reactive oxygen species (ROS). In contrast, Leishmania mutants resistant to SbIII, MF or AMB not only failed to undergo apoptosis following exposure to their respective drugs, but also were more tolerant towards apoptosis induced by other antileishmanials, provided that these killed Leishmania via ROS production. Such tolerance favored the rapid acquisition of multidrug resistance. PARO killed Leishmania in a non-apoptotic manner and failed to produce ROS. PARO resistance neither protected against drug-induced apoptosis nor provided an increased rate of acquisition of resistance to other antileishmanials. However, the PARO-resistant mutant, but not SbIII-, MF- or AMB-resistant mutants, became rapidly cross-resistant to methotrexate, a model drug also not producing ROS. Our results therefore link the mode of killing of drugs to tolerance to cell death and to a facilitated emergence of multidrug resistance. These findings may have fundamental implications in the field of chemotherapeutic interventions

<i>Trypanosoma brucei</i> DHRF-TS revisited:characterisation of a bifunctional and highly unstable recombinant dihydrofolate reductase-thymidylate synthase

<div><p>Bifunctional dihydrofolate reductase–thymidylate synthase (DHFR-TS) is a chemically and genetically validated target in African trypanosomes, causative agents of sleeping sickness in humans and nagana in cattle. Here we report the kinetic properties and sensitivity of recombinant enzyme to a range of lipophilic and classical antifolate drugs. The purified recombinant enzyme, expressed as a fusion protein with elongation factor Ts (Tsf) in ThyA^- <i>Escherichia coli</i>, retains DHFR activity, but lacks any TS activity. TS activity was found to be extremely unstable (half-life of 28 s) following desalting of clarified bacterial lysates to remove small molecules. Stability could be improved 700-fold by inclusion of dUMP, but not by other pyrimidine or purine (deoxy)-nucleosides or nucleotides. Inclusion of dUMP during purification proved insufficient to prevent inactivation during the purification procedure. Methotrexate and trimetrexate were the most potent inhibitors of DHFR (<i>K</i>_i 0.1 and 0.6 nM, respectively) and FdUMP and nolatrexed of TS (<i>K</i>_i 14 and 39 nM, respectively). All inhibitors showed a marked drop-off in potency of 100- to 1,000-fold against trypanosomes grown in low folate medium lacking thymidine. The most potent inhibitors possessed a terminal glutamate moiety suggesting that transport or subsequent retention by polyglutamylation was important for biological activity. Supplementation of culture medium with folate markedly antagonised the potency of these folate-like inhibitors, as did thymidine in the case of the TS inhibitors raltitrexed and pemetrexed.</p></div

Provider imposed restrictions to clients’ access to family planning in urban Uttar Pradesh, India: a mixed methods study

BACKGROUND: Medical barriers refer to unnecessary policies or procedures imposed by health care providers that are not necessarily medically advised; these restrictions impede clients’ access to family planning (FP). This mixed methods study investigates provider imposed barriers to provision of FP using recent quantitative and qualitative data from urban Uttar Pradesh, India. METHODS: Baseline quantitative data were collected in six cities in Uttar Pradesh, India from service delivery points (SDP), using facility audits, exit interviews, and provider surveys; for this study, the focus is on the provider surveys. More than 250 providers were surveyed in each city. Providers were asked about the FP methods they provide, and if they restrict clients’ access to each method based on age, parity, partner consent, or marital status. For the qualitative research, we conducted one-on-one interviews with 21 service providers in four of the six cities in Uttar Pradesh. Each interview lasted approximately 45 minutes. RESULTS: The quantitative findings show that providers restrict clients’ access to spacing and long-acting and permanent methods of FP based on age, parity, partner consent and marital status. Qualitative findings reinforce that providers, at times, make judgments about their clients’ education, FP needs and ability to understand FP options thereby imposing unnecessary barriers to FP methods. CONCLUSIONS: Provider restrictions on FP methods are common in these urban Uttar Pradesh sites. This means that women who are young, unmarried, have few or no children, do not have the support of their partner, or are less educated may not be able to access or use FP or their preferred method. These findings highlight the need for in-service training for staff, with a focus on reviewing current guidelines and eligibility criteria for provision of methods

Genomic and Proteomic Studies on the Mode of Action of Oxaboroles against the African Trypanosome

SCYX-7158, an oxaborole, is currently in Phase I clinical trials for the treatment of human African trypanosomiasis. Here we investigate possible modes of action against Trypanosoma brucei using orthogonal chemo-proteomic and genomic approaches. SILAC-based proteomic studies using an oxaborole analogue immobilised onto a resin was used either in competition with a soluble oxaborole or an immobilised inactive control to identify thirteen proteins common to both strategies. Cell-cycle analysis of cells incubated with sub-lethal concentrations of an oxaborole identified a subtle but significant accumulation of G2 and >G2 cells. Given the possibility of compromised DNA fidelity, we investigated long-term exposure of T. brucei to oxaboroles by generating resistant cell lines in vitro. Resistance proved more difficult to generate than for drugs currently used in the field, and in one of our three cell lines was unstable. Whole-genome sequencing of the resistant cell lines revealed single nucleotide polymorphisms in 66 genes and several large-scale genomic aberrations. The absence of a simple consistent mechanism among resistant cell lines and the diverse list of binding partners from the proteomic studies suggest a degree of polypharmacology that should reduce the risk of resistance to this compound class emerging in the field. The combined genetic and chemical biology approaches have provided lists of candidates to be investigated for more detailed information on the mode of action of this promising new drug clas

Do schistosome vaccine trials in mice have an intrinsic flaw that generates spurious protection data?

The laboratory mouse has been widely used to test the efficacy of schistosome vaccines and a long list of candidates has emerged from this work, many of them abundant internal proteins. These antigens do not have an additive effect when co-administered, or delivered as SWAP homogenate, a quarter of which comprises multiple candidates; the observed protection has an apparent ceiling of 40–50 %. We contend that the low level of maturation of penetrating cercariae (~32 % for Schistosoma mansoni) is a major limitation of the model since 68/100 parasites fail to mature in naïve mice due to natural causes. The pulmonary capillary bed is the obstacle encountered by schistosomula en route to the portal system. The fragility of pulmonary capillaries and their susceptibility to a cytokine-induced vascular leak syndrome have been documented. During lung transit schistosomula burst into the alveolar spaces, and possess only a limited capacity to re-enter tissues. The acquired immunity elicited by the radiation attenuated (RA) cercarial vaccine relies on a pulmonary inflammatory response, involving cytokines such as IFNγ and TNFα, to deflect additional parasites into the alveoli. A principal difference between antigen vaccine protocols and the RA vaccine is the short interval between the last antigen boost and cercarial challenge of mice (often two weeks). Thus, after antigen vaccination, challenge parasites will reach the lungs when both activated T cells and cytokine levels are maximal in the circulation. We propose that “protection” in this situation is the result of physiological effects on the pulmonary blood vessels, increasing the proportion of parasites that enter the alveoli. This hypothesis will explain why internal antigens, which are unlikely to interact with the immune response in a living schistosomulum, plus a variety of heterologous proteins, can reduce the level of maturation in a non-antigen-specific way. These proteins are “successful” precisely because they have not been selected for immunological silence. The same arguments apply to vaccine experiments with S. japonicum in the mouse model; this schistosome species seems a more robust parasite, even harder to eliminate by acquired immune responses. We propose a number of ways in which our conclusions may be tested

Leishmania infantum Asparagine Synthetase A Is Dispensable for Parasites Survival and Infectivity

A growing interest in asparagine (Asn) metabolism has currently been observed in cancer and infection fields. Asparagine synthetase (AS) is responsible for the conversion of aspartate into Asn in an ATP-dependent manner, using ammonia or glutamine as a nitrogen source. There are two structurally distinct AS: the strictly ammonia dependent, type A, and the type B, which preferably uses glutamine. Absent in humans and present in trypanosomatids, AS-A was worthy of exploring as a potential drug target candidate. Appealingly, it was reported that AS-A was essential in Leishmania donovani, making it a promising drug target. In the work herein we demonstrate that Leishmania infantum AS-A, similarly to Trypanosoma spp. and L. donovani, is able to use both ammonia and glutamine as nitrogen donors. Moreover, we have successfully generated LiASA null mutants by targeted gene replacement in L. infantum, and these parasites do not display any significant growth or infectivity defect. Indeed, a severe impairment of in vitro growth was only observed when null mutants were cultured in asparagine limiting conditions. Altogether our results demonstrate that despite being important under asparagine limitation, LiAS-A is not essential for parasite survival, growth or infectivity in normal in vitro and in vivo conditions. Therefore we exclude AS-A as a suitable drug target against L. infantum parasites