CLEAR - clozapine in early psychosis: study protocol for a multi-centre, randomised controlled trial of clozapine vs other antipsychotics for young people with treatment resistant schizophrenia in real world settings.

BACKGROUND: Clozapine is an antipsychotic drug with unique efficacy, and it is the only recommended treatment for treatment-resistant schizophrenia (TRS: failure to respond to at least two different antipsychotics). However, clozapine is also associated with a range of adverse effects which restrict its use, including blood dyscrasias, for which haematological monitoring is required. As treatment resistance is recognised earlier in the illness, the question of whether clozapine should be prescribed in children and young people is increasingly important. However, most research to date has been in older, chronic patients, and evidence regarding the efficacy and safety of clozapine in people under age 25 is lacking. The CLEAR (CLozapine in EARly psychosis) trial will assess whether clozapine is more effective than treatment as usual (TAU), at the level of clinical symptoms, patient rated outcomes, quality of life and cost-effectiveness in people below 25 years of age. Additionally, a nested biomarker study will investigate the mechanisms of action of clozapine compared to TAU. METHODS AND DESIGN: This is the protocol of a multi-centre, open label, blind-rated, randomised controlled effectiveness trial of clozapine vs TAU (any other oral antipsychotic monotherapy licenced in the British National Formulary) for 12 weeks in 260 children and young people with TRS (12-24 years old). AIM AND OBJECTIVES: The primary outcome is the change in blind-rated Positive and Negative Syndrome Scale scores at 12 weeks from baseline. Secondary outcomes include blind-rated Clinical Global Impression, patient-rated outcomes, quality of life, adverse effects, and treatment adherence. Patients will be followed up for 12 months and will be invited to give consent for longer term follow-up using clinical records and potential re-contact for further research. For mechanism of action, change in brain magnetic resonance imaging (MRI) biomarkers and peripheral inflammatory markers will be measured over 12 weeks. DISCUSSION: The CLEAR trial will contribute knowledge on clozapine effectiveness, safety and cost-effectiveness compared to standard antipsychotics in young people with TRS, and the results may guide future clinical treatment recommendation for early psychosis. TRIAL REGISTRATION: ISRCTN Number: 37176025, IRAS Number: 1004947. TRIAL STATUS: In set-up. Protocol version 4.0 01/08/23. Current up to date protocol available here: https://fundingawards.nihr.ac.uk/award/NIHR131175# /.

Enhancers associated with unstable RNAs are rare in plants.

Unstable transcripts have emerged as markers of active enhancers in vertebrates and shown to be involved in many cellular processes and medical disorders. However, their prevalence and role in plants is largely unexplored. Here, we comprehensively captured all actively initiating ("nascent") transcripts across diverse crops and other plants using capped small (cs)RNA-seq. We discovered that unstable transcripts are rare, unlike in vertebrates, and often originate from promoters. Additionally, many "distal" elements in plants initiate tissue-specific stable transcripts and are likely bone fide promoters of yet-unannotated genes or non-coding RNAs, cautioning against using genome annotations to infer "enhancers" or transcript stability. To investigate enhancer function, we integrated STARR-seq data. We found that annotated promoters, and other regions that initiate stable transcripts rather than unstable transcripts, function as stronger enhancers in plants. Our findings underscore the blurred line between promoters and enhancers and suggest that cis-regulatory elements encompass diverse structures and mechanisms in eukaryotes.

Exhaust the exhausters: Targeting regulatory T cells in the tumor microenvironment.

The concept of cancer immunotherapy has gained immense momentum over the recent years. The advancements in checkpoint blockade have led to a notable progress in treating a plethora of cancer types. However, these approaches also appear to have stalled due to factors such as individuals' genetic make-up, resistant tumor sub-types and immune related adverse events (irAE). While the major focus of immunotherapies has largely been alleviating the cell-intrinsic defects of CD8+ T cells in the tumor microenvironment (TME), amending the relationship between tumor specific CD4+ T cells and CD8+ T cells has started driving attention as well. A major roadblock to improve the cross-talk between CD4+ T cells and CD8+ T cells is the immune suppressive action of tumor infiltrating T regulatory (Treg) cells. Despite their indispensable in protecting tissues against autoimmune threats, Tregs have also been under scrutiny for helping tumors thrive. This review addresses how Tregs establish themselves at the TME and suppress anti-tumor immunity. Particularly, we delve into factors that promote Treg migration into tumor tissue and discuss the unique cellular and humoral composition of TME that aids survival, differentiation and function of intratumoral Tregs. Furthermore, we summarize the potential suppression mechanisms used by intratumoral Tregs and discuss ways to target those to ultimately guide new immunotherapies.

Radiation dosimetry by automatic image analysis of dicentric chromosomes.

A system for scoring dicentric chromosomes by image analysis comprised fully automatic location of mitotic cells, automatic retrieval, focus and digitization at high resolution, automatic rejection of nuclei and debris and detection and segmentation of chromosome clusters, automatic centromere location, and subsequent rapid interactive visual review of potential dicentric chromosomes to confirm positives and reject false positives. A calibration set of about 15,000 cells was used to establish the quadratic dose response for 60Co gamma-irradiation. The dose-response function parameters were established by a maximum likelihood technique, and confidence limits in the dose response and in the corresponding inverse curve, of estimated dose for observed dicentric frequency, were established by Monte Carlo techniques. The system was validated in a blind trial by analysing a test set comprising a total of about 8000 cells irradiated to 1 of 10 dose levels, and estimating the doses from the observed dicentric frequency. There was a close correspondence between the estimated and true doses. The overall sensitivity of the system in terms of the proportion of the total population of dicentrics present in the cells analysed that were detected by the system was measured to be about 40%. This implies that about 2.5 times more cells must be analysed by machine than by visual analysis. Taking this factor into account, the measured review time and false positive rates imply that analysis by the system of sufficient cells to provide the equivalent of a visual analysis of 500 cells would require about 1 h for operator review.

Automated densitometry of cell populations in a continuous-motion imaging cell scanner.

Continuous-motion imaging provides a method for the rapid quantitative analysis of slide-mounted cell preparations. Densitometric and morphometric cellular features can be measured and used to classify individual cells. Tests on the densitometric accuracy of the CERVIFIP CMI system show a c.v. of 6.5% for diploid mouse hepatocytes and a measuring rate of up to 500 cells/sec. CMI is particularly useful for problems requiring the interactive analysis of rare cell types and for the measurement of morphological and densitometric characteristics of cell populations. Typical applications include the prescreening of cervical cytology specimens and the detection of rare autoradiographically labeled lymphocytes.

A fast interval processor (FIP) for cervical prescreening.

A cervical prescreening device (FIP: fast interval processor) designed to scan and classify a slide-mounted specimen within two minutes is described. The image analysis techniques are based directly on the MRC Cerviscan equipment with the minimal conversion needed to adapt these techniques for interval processing. A high scanning rate is achieved by scanning with a charge-coupled diode linear image sensor along one axis and by stepping the microscope stage continuously along the other axis. High processing rates are achieved using an asynchronous pipeline approach. Operations on pixels are carried out by parallel dedicated hardware units, while operations on intervals (segments) are carrie out using a dual microprocessing configuration. A determined attempt has been made to minimize the cost of the components required. Preliminary results showing scanning and processing performance are given.

A cytogeneticist's microscope.

It is demonstrated that there are a number of advantages in using a mechanised microscope for scoring a large number of metaphase cells from human blood lymphocyte preparations. Following the development of an automatic metaphase spread finding machine based upon a large motorised microscope and a synchronous closed circuit television camera and flashing light source, a much smaller machine which is more appropriate to the cytogenetics laboratory, but with a similar metaphase finding performance has been constructed. The new machine which consists of a Cambridge Instruments 1 micron stepping microscope stage, a linear diode array scanner and a computer is described in detail. Metaphase finding performance figures for various orcein stained human blood lymphocyte preparations are given.