Phenotypic screening reveals TNFR2 as a promising target for cancer immunotherapy.

Antibodies that target cell-surface molecules on T cells can enhance anti-tumor immune responses, resulting in sustained immune-mediated control of cancer. We set out to find new cancer immunotherapy targets by phenotypic screening on human regulatory T (Treg) cells and report the discovery of novel activators of tumor necrosis factor receptor 2 (TNFR2) and a potential role for this target in immunotherapy. A diverse phage display library was screened to find antibody mimetics with preferential binding to Treg cells, the most Treg-selective of which were all, without exception, found to bind specifically to TNFR2. A subset of these TNFR2 binders were found to agonise the receptor, inducing iκ-B degradation and NF-κB pathway signalling in vitro. TNFR2 was found to be expressed by tumor-infiltrating Treg cells, and to a lesser extent Teff cells, from three lung cancer patients, and a similar pattern was also observed in mice implanted with CT26 syngeneic tumors. In such animals, TNFR2-specific agonists inhibited tumor growth, enhanced tumor infiltration by CD8+ T cells and increased CD8+ T cell IFN-γ synthesis. Together, these data indicate a novel mechanism for TNF-α-independent TNFR2 agonism in cancer immunotherapy, and demonstrate the utility of target-agnostic screening in highlighting important targets during drug discovery.GW, BM, SG, JC-U, AS, AG-M, CB, JJ, RL, AJL, SR, RS, LJ, VV-A, RM and RWW were funded by MedImmune; JP and VB were funded by AstraZeneca PLC; JW, RSA-L and JB were funded by NIHR Cambridge Biomedical Research Centre and Kidney Research UK; JS and JF were funded by Retrogenix Ltd

Evaluation of a pro-recovery training intervention (REFOCUS-RETAFORM) in specialist mental health services across France: stepped-wedge cluster randomised controlled trial protocol

BackgroundWhile recovery orientation is national policy in many countries, evidence remains limited for the effectiveness at a service level. This paper describes the protocol for implementing a pro-recovery training intervention (REFOCUS-RETAFORM) in specialist mental health services across France. The aim is to evaluate whether REFOCUS-RETAFORM plus usual care leads to improved outcomes for adolescent and adult mental health service users compared with usual care alone.MethodsA two-step stepped wedge cluster randomised controlled trial will be conducted, with a nested qualitative sub-study exploring stakeholders’ views on changes in staff-user relationships and implementation influences. The REFOCUS-RETAFORM intervention is a training intervention for mental health staff, to develop recovery-promoting relationships and pro-recovery working practices. Clusters are services, which transition sequentially from control to intervention condition in a randomised order. Eight clusters are randomised to deliver REFOCUS-RETAFORM in year one and eight clusters in year two. Each cluster delivers REFOCUS-RETAFORM to two teams from their organisation (32 teams in total). Participants are a) service users aged 13–65 years attending services implementing REFOCUS-RETAFORM, and b) staff receiving the intervention. The primary outcome is the Questionnaire about the Process of Recovery. Secondary outcomes include perceived stigma and coercion, self-stigma and wellbeing for service users, and recovery-orientation for staff. Data will be collected from 540 service users (180 at baseline, 180 at month 12, 180 at month 24) and 220 staff. We will use multilevel mixed-effects models, adjusting for secular trends and thematic analysis for the qualitative interview data.DiscussionFindings will inform the continued transformation of French specialist mental health services toward a recovery orientation

Exo1 and Rad24 Differentially Regulate Generation of ssDNA at Telomeres of Saccharomyces cerevisiae cdc13-1 Mutants

Cell cycle arrest in response to DNA damage depends upon coordinated interactions between DNA repair and checkpoint pathways. Here we examine the role of DNA repair and checkpoint genes in responding to unprotected telomeres in budding yeast cdc13-1 mutants. We show that Exo1 is unique among the repair genes tested because like Rad9 and Rad24 checkpoint proteins, Exo1 inhibits the growth of cdc13-1 mutants at the semipermissive temperatures. In contrast Mre11, Rad50, Xrs2, and Rad27 contribute to the vitality of cdc13-1 strains grown at permissive temperatures, while Din7, Msh2, Nuc1, Rad2, Rad52, and Yen1 show no effect. Exo1 is not required for cell cycle arrest of cdc13-1 mutants at 36° but is required to maintain arrest. Exo1 affects but is not essential for the production of ssDNA in subtelomeric Y′ repeats of cdc13-1 mutants. However, Exo1 is critical for generating ssDNA in subtelomeric X repeats and internal single-copy sequences. Surprisingly, and in contrast to Rad24, Exo1 is not essential to generate ssDNA in X or single-copy sequences in cdc13-1 rad9Δ mutants. We conclude that Rad24 and Exo1 regulate nucleases with different properties at uncapped telomeres and propose a model to explain our findings

MRX protects telomeric DNA at uncapped telomeres of budding yeast cdc13-1 mutants

MRX, an evolutionally conserved DNA damage response complex composed of Mre11, Rad50 and Xrs2, is involved in DNA double strand break (DSB) repair, checkpoint activation and telomere maintenance. At DSBs, MRX plays a role in generating single stranded DNA (ssDNA) and signalling cell cycle arrest. Here we investigated whether MRX also contributes to generating ssDNA or signalling cell cycle arrest at uncapped telomeres. To investigate the role of MRX, we generated a conditionally degradable Rad50 protein and combined this with cdc13-1, a temperature sensitive mutation in the Cdc13 telomere capping protein. We show that Rad50 does not contribute to ssDNA generation or cell cycle arrest in response to cdcl3-1 uncapped telomeres. Instead, we find that Rad50 inhibits ssDNA accumulation and promotes cdc13-1 cell viability, consistent with a major role for MRX in telomere capping