Discovery of a small-molecule inhibitor and cellular probe of Keap1–Nrf2 protein–protein interaction

A high-throughput screen (HTS) of the MLPCN library using a homogenous fluorescence polarization assay identified a small molecule as a first-in-class direct inhibitor of Keap1–Nrf2 protein–protein interaction. The HTS hit has three chiral centers; a combination of flash and chiral chromatographic separation demonstrated that Keap1-binding activity resides predominantly in one stereoisomer (SRS)-5 designated as ML334 (LH601A), which is at least 100× more potent than the other stereoisomers. The stereochemistry of the four cis isomers was assigned using X-ray crystallography and confirmed using stereospecific synthesis. (SRS)-5 is functionally active in both an ARE gene reporter assay and an Nrf2 nuclear translocation assay. The stereospecific nature of binding between (SRS)-5 and Keap1 as well as the preliminary but tractable structure–activity relationships support its use as a lead for our ongoing optimization     

Azetidines and spiro azetidines as novel P2 units in hepatitis C virus NS3 protease inhibitors

Herein, we report the synthesis and structure–activity relationship studies of new analogs of boceprevir 1 and telaprevir 2. Introduction of azetidine and spiroazetidines as a P2 substituent that replaced the pyrrolidine moiety of 1 and 2 led to the discovery of a potent hepatitis C protease inhibitor 37c (EC₅₀ = 0.8 μM).     

Synthesis and evaluation of Janus type nucleosides as potential HCV NS5B polymerase inhibitors

The synthesis of new ribo and 2′-β-C-methyl ribo Janus type nucleosides J-AA, J-AG and J-AU is reported along with their ability to block HCV and HIV replication. Their toxicity was also assessed in Huh7, human lymphocytes, CEM and Vero cells.     

Phylogeography of the widespread African puff adder (Bitis arietans) reveals multiple Pleistocene refugia in southern Africa

Evidence from numerous Pan‐African savannah mammals indicates that open‐habitat refugia existed in Africa during the Pleistocene, isolated by expanding tropical forests during warm and humid interglacial periods. However, comparative data from other taxonomic groups are currently lacking. We present a phylogeographic investigation of the African puff adder (Bitis arietans), a snake that occurs in open‐habitat formations throughout sub‐Saharan Africa. Multiple parapatric mitochondrial clades occur across the current distribution of B. arietans, including a widespread southern African clade that is subdivided into four separate clades. We investigated the historical processes responsible for generating these phylogeographic patterns in southern Africa using species distribution modelling and genetic approaches. Our results show that interior regions of South Africa became largely inhospitable for B. arietans during glacial maxima, whereas coastal and more northerly areas remained habitable. This corresponds well with the locations of refugia inferred from mitochondrial data using a continuous phylogeographic diffusion model. Analysis of data from five anonymous nuclear loci revealed broadly similar patterns to mtDNA. Secondary admixture was detected between previously isolated refugial populations. In some cases, this is limited to individuals occurring near mitochondrial clade contact zones, but in other cases, more extensive admixture is evident. Overall, our study reveals a complex history of refugial isolation and secondary expansion for puff adders and a mosaic of isolated refugia in southern Africa. We also identify key differences between the processes that drove isolation in B. arietans and those hypothesized for sympatric savannah mammals.     

Phenotypic changes associated with RNA interference silencing of chalcone synthase in apple (Malus × domestica)

We have identified in apple (Malus × domestica) three chalcone synthase (CHS) genes. In order to understand the functional redundancy of this gene family RNA interference knockout lines were generated where all three of these genes were down‐regulated. These lines had no detectable anthocyanins and radically reduced concentrations of dihydrochalcones and flavonoids. Surprisingly, down‐regulation of CHS also led to major changes in plant development, resulting in plants with shortened internode lengths, smaller leaves and a greatly reduced growth rate. Microscopic analysis revealed that these phenotypic changes extended down to the cellular level, with CHS‐silenced lines showing aberrant cellular organisation in the leaves. Fruit collected from one CHS‐silenced line was smaller than the ‘Royal Gala’ controls, lacked flavonoids in the skin and flesh and also had changes in cell morphology. Auxin transport experiments showed increased rates of auxin transport in a CHS‐silenced line compared with the ‘Royal Gala’ control. As flavonoids are well known to be key modulators of auxin transport, we hypothesise that the removal of almost all flavonoids from the plant by CHS silencing creates a vastly altered environment for auxin transport to occur and results in the observed changes in growth and development.     

Photoluminescent Gold Nanoclusters as Sensing Probes for Uropathogenic Escherichia coli

Glycan-bound nanoprobes have been demonstrated as suitable sensing probes for bacteria containing glycan binding sites. In this study, we demonstrated a facile approach for generating glycan-bound gold nanoclusters (AuNCs). The generated AuNCs were used as sensing probes for corresponding target bacteria. Mannose-capped AuNCs (AuNCs@Mann) were generated and used as the model sensors for target bacteria. A one-step synthesis approach was employed to generate AuNCs@Mann. In this approach, an aqueous solution of tetrachloroauric acid and mannoside that functionized with a thiol group (Mann-SH) was stirred at room temperature for 48 h. The mannoside functions as reducing and capping agent. The size of the generated AuNCs@Mann is 1.95±0.27 nm, whereas the AuNCs with red photoluminescence have a maximum emission wavelength of ∼630 nm (λ_excitation = 375 nm). The synthesis of the AuNCs@Mann was accelerated by microwave heating, which enabled the synthesis of the AuNCs@Mann to complete within 1 h. The generated AuNCs@Mann are capable of selectively binding to the urinary tract infection isolate Escherichia coli J96 containing the mannose binding protein FimH expressed on the type 1 pili. On the basis of the naked eye observation, the limit of detection of the sensing approach is as low as ∼2×10⁶ cells/mL.     

TB Incidence in an Adolescent Cohort in South Africa

Background

Tuberculosis (TB) is a major public health problem globally. Little is known about TB incidence in adolescents who are a proposed target group for new TB vaccines. We conducted a study to determine the TB incidence rates and risk factors for TB disease in a cohort of school-going adolescents in a high TB burden area in South Africa.

Methods

We recruited adolescents aged 12 to 18 years from high schools in Worcester, South Africa. Demographic and clinical information was collected, a tuberculin skin test (TST) performed and blood drawn for a QuantiFERON TB Gold assay at baseline. Screening for TB cases occurred at follow up visits and by surveillance of registers at public sector TB clinics over a period of up to 3.8 years after enrolment.

Results

A total of 6,363 adolescents were enrolled (58% of the school population targeted). During follow up, 67 cases of bacteriologically confirmed TB were detected giving an overall incidence rate of 0.45 per 100 person years (95% confidence interval 0.29–0.72). Black or mixed race, maternal education of primary school or less or unknown, a positive baseline QuantiFERON assay and a positive baseline TST were significant predictors of TB disease on adjusted analysis.

Conclusion

The adolescent TB incidence found in a high burden setting will help TB vaccine developers plan clinical trials in this population. Latent TB infection and low socio-economic status were predictors of TB disease.     

Science Educational Outreach Programs That Benefit Students and Scientists

Both scientists and the public would benefit from improved communication of basic scientific research and from integrating scientists into education outreach, but opportunities to support these efforts are limited. We have developed two low-cost programs—"Present Your PhD Thesis to a 12-Year-Old" and "Shadow a Scientist”—that combine training in science communication with outreach to area middle schools. We assessed the outcomes of these programs and found a 2-fold benefit: scientists improve their communication skills by explaining basic science research to a general audience, and students'' enthusiasm for science and their scientific knowledge are increased. Here we present details about both programs, along with our assessment of them, and discuss the feasibility of exporting these programs to other universities.