Development of a Fluorescent Ligand for the Intracellular Allosteric Binding Site of the Neurotensin Receptor 1.

The membrane protein family of G protein-coupled receptors (GPCRs) represents a major class of drug targets. Over the last years, the presence of additional intracellular binding sites besides the canonical orthosteric binding pocket has been demonstrated for an increasing number of GPCRs. Allosteric modulators harnessing these pockets may represent valuable alternatives when targeting the orthosteric pocket is not successful for drug development. Starting from SBI-553, a recently discovered intracellular allosteric modulator for neurotensin receptor subtype 1 (NTSR1), we developed the fluorescent molecular probe 14. Compound 14 binds to NTSR1 with an affinity of 0.68 µM in the presence of the agonist NT(8-13). NanoBRET-based ligand binding assays with 14 were established to derive the affinity and structure-activity relationships for allosteric NTSR1 modulators in a direct and nonisotopic manner, thereby facilitating the search for and optimization of novel allosteric NTSR1 ligands. As a consequence of cooperativity between the ligands binding to the allosteric and orthosteric pocket, compound 14 can also be used to investigate orthosteric NTSR1 agonists and antagonists. Moreover, employing 14 as a probe in a drug library screening, we identified novel chemotypes as binders for the intracellular allosteric SBI-553 binding pocket of NTSR1 with single-digit micromolar affinity. These hits may serve as interesting starting points for the development of novel intracellular allosteric ligands for NTSR1 as a highly interesting yet unexploited drug target in the fields of pain and addiction disorder therapy.

A Chemical Biology Toolbox Targeting the Intracellular Binding Site of CCR9: Fluorescent Ligands, New Drug Leads and PROTACs.

A conserved intracellular allosteric binding site (IABS) has recently been identified at several G protein-coupled receptors (GPCRs). Starting from vercirnon, an intracellular C-C chemokine receptor type 9 (CCR9) antagonist and previous phase III clinical candidate for the treatment of Crohn's disease, we developed a chemical biology toolbox targeting the IABS of CCR9. We first synthesized a fluorescent ligand enabling equilibrium and kinetic binding studies via NanoBRET as well as fluorescence microscopy. Applying this molecular tool in a membrane-based setup and in living cells, we discovered a 4-aminopyrimidine analogue as a new intracellular CCR9 antagonist with improved affinity. To chemically induce CCR9 degradation, we then developed the first PROTAC targeting the IABS of GPCRs. In a proof-of-principle study, we succeeded in showing that our CCR9-PROTAC is able to reduce CCR9 levels, thereby offering an unprecedented approach to modulate GPCR activity.

Bivalent ligands promote endosomal trafficking of the dopamine D3 receptor-neurotensin receptor 1 heterodimer.

Bivalent ligands are composed of two pharmacophores connected by a spacer of variable size. These ligands are able to simultaneously recognize two binding sites, for example in a G protein-coupled receptor heterodimer, resulting in enhanced binding affinity. Taking advantage of previously described heterobivalent dopamine-neurotensin receptor ligands, we demonstrate specific interactions between dopamine D3 (D3R) and neurotensin receptor 1 (NTSR1), two receptors with expression in overlapping brain areas that are associated with neuropsychiatric diseases and addiction. Bivalent ligand binding to D3R-NTSR1 dimers results in picomolar binding affinity and high selectivity compared to the binding to monomeric receptors. Specificity of the ligands for the D3R-NTSR1 receptor pair over D2R-NTSR1 dimers can be achieved by a careful choice of the linker length. Bivalent ligands enhance and stabilize the receptor-receptor interaction leading to NTSR1-controlled internalization of D3R into endosomes via recruitment of ß-arrestin, highlighting a potential mechanism for dimer-specific receptor trafficking and signalling.

Lessons from a community vaccination programme to control a meningococcal disease serogroup W outbreak in remote South Australia, 2017.

PROBLEM: From December 2016 to February 2017, two cases of invasive meningococcal disease and one case of meningococcal conjunctivitis, all serogroup W, occurred in Aboriginal children in the Ceduna region of South Australia. The clustering of cases in time and place met the threshold for a community outbreak. CONTEXT: The Ceduna region is a remote part of South Australia, with more than 25% of the population identifying as Aboriginal or Torres Strait Islander. ACTION: As part of the outbreak response, a community-wide meningococcal vaccination programme against serogroups A, C, W and Y was implemented in a collaboration among different agencies of the South Australia Department for Health and Well-being, Aboriginal health and community services providers, and other local service providers and government agencies. The programme comprised an outbreak vaccination schedule, targeting all people aged 3 2 months residing in the cases' places of residence or in towns with close links. OUTCOME: Between March and June 2017, 3383 persons were vaccinated, achieving an estimated coverage of 71-85% of the target population, with 31% (n = 1034) of those vaccinated identifying as Aboriginal or Torres Strait Islander. No local cases of serogroup W occurred during the vaccination programme, but two further cases were notified by the end of 2018. DISCUSSION: The participation of a large number of local and non-health-sector stakeholders in programme planning and implementation, a clear response management structure and high community acceptability were identified as key factors that contributed to the programme achieving high vaccination coverage. The need to develop standard operating procedures for community-based outbreak response interventions to ease logistical challenges was considered an important lesson learnt.

Correction to: Managing Urban Plant Invasions: a Multi-Criteria Prioritization Approach.

The original version of the article unfortunately contained an error with the figure captions. The appropriate captions for Fig. 3-6 are published accordingly. The original article has been corrected.

Managing Urban Plant Invasions: a Multi-Criteria Prioritization Approach.

Alien plant invasions in urban areas can have considerable impact on biodiversity and ecosystem services (ES). Managing urban plant invasions is particularly challenging given the complex interactions between ecological, economic and social elements that exist in the urban milieu. Strategic landscape-scale insights are crucial for guiding management, as are tactical site-scale perspectives to plan and coordinate control efforts on the ground. Integrating these requirements to enhance management efficiency is a major challenge. Decision-support models have considerable potential for guiding and informing management strategies when problems are complex. This study uses multi-criteria decision tools to develop a prioritization framework for managing invasive alien plants (IAPs) in urban areas at landscape and local scales. We used the Analytic Hierarchy Process (AHP; a multi-criteria decision support model) to develop and rank criteria for prioritising IAP management in the City of Cape Town (CoCT), South Africa. Located within a global biodiversity hotspot, Cape Town has a long history of alien plant introductions and a complex socio-political make-up, creating a useful system to explore the challenges associated with managing urban plant invasions. To guide the prioritization of areas for IAP management across the CoCT, a stakeholder workshop was held to identify a goal and criteria for consideration, and to assess the relative importance given to each criterion in IAP management. Workshop attendees were drawn from multiple disciplines involved with different aspects of IAP research and management: government departments, scientists and researchers, and managers with a diverse set of skills and interests. We selected spatial datasets and applied our multi-criteria decision analysis in a Geographic Information System (GIS) to develop a landscape-scale prioritization map. To address issues relevant in an urban setting, we also modified an existing IAP management framework to develop a tactical (site-level) prioritization scheme for guiding on-the-ground control operations. High-priority sites for IAP management were identified at landscape- and local scales across the study area. Factors related to safety and security emerged as pivotal features for setting spatially-explicit priorities for management. The approach applied in this study can be useful for managers in all urban settings to guide the selection and prioritization of areas for IAP management.