The PRESIDE (PhaRmacogEnomicS In DEpression) Trial: a double-blind randomised controlled trial of pharmacogenomic-informed prescribing of antidepressants on depression outcomes in patients with major depressive disorder in primary care.

BACKGROUND: The evidence for the clinical utility of pharmacogenomic (PGx) testing is growing, and guidelines exist for the use of PGx testing to inform prescribing of 13 antidepressants. Although previous randomised controlled trials of PGx testing for antidepressant prescribing have shown an association with remission of depression in clinical psychiatric settings, few trials have focused on the primary care setting, where most antidepressant prescribing occurs. METHODS: The PRESIDE Trial is a stratified double-blinded randomised controlled superiority trial that aims to evaluate the impact of a PGx-informed antidepressant prescribing report (compared with standard prescribing using the Australian Therapeutic Guidelines) on depressive symptoms after 12 weeks, when delivered in primary care. Six hundred seventy-two patients aged 18-65 years of general practitioners (GPs) in Victoria with moderate to severe depressive symptoms, measured using the Patient Health Questionnaire-9 (PHQ-9), will be randomly allocated 1:1 to each arm using a computer-generated sequence. Participants and GPs will be blinded to the study arm. The primary outcome is a difference between arms in the change of depressive symptoms, measured using the PHQ-9 after 12 weeks. Secondary outcomes include a difference between the arms in change in PHQ-9 score at 4, 8 and 26 weeks, proportion in remission at 12 weeks, a change in side effect profile of antidepressant medications, adherence to antidepressant medications, change in quality of life and cost-effectiveness of the intervention. DISCUSSION: This trial will provide evidence as to whether PGx-informed antidepressant prescribing is clinically efficacious and cost-effective. It will inform national and international policy and guidelines about the use of PGx to select antidepressants for people with moderate to severe depressive symptoms presenting in primary care. TRIAL REGISTRATION: Australian and New Zealand Clinical Trial Registry ACTRN12621000181808. Registered on 22 February 2021.

Application of Whole Genome Sequencing Technology in the Investigation of Genetic Causes of Fetal, Perinatal, and Early Infant Death.

Death in the fetal, perinatal, and early infant age-group has a multitude of causes, a proportion of which is presumed to be genetic. Defining a specific genetic aberration leading to the death is problematic at this young age, due to limited phenotype-genotype correlation inherent in the underdeveloped phenotype, the inability to assess certain phenotypic traits after death, and the problems of dealing with rare disorders. In this study, our aim was to increase the yield of identification of a defined genetic cause of an early death. Therefore, we employed whole genome sequencing and bioinformatic filtering techniques as a comprehensive, unbiased genetic investigation into 16 fetal, perinatal, and early infant deaths, which had undergone a full autopsy. A likely genetic cause was identified in two cases (in genes; COL2A1 and RYR1) and a speculative genetic cause in a further six cases (in genes: ARHGAP35, BBS7, CASZ1, CRIM1, DHCR7, HADHB, HAPLN3, HSPG2, MYO18B, and SRGAP2). This investigation indicates that whole genome sequencing is a significantly enabling technology when determining genetic causes of early death.

Nuclear export and centrosome targeting of the protein phosphatase 2A subunit B56alpha: role of B56alpha in nuclear export of the catalytic subunit.

Protein phosphatase (PP) 2A is a heterotrimeric enzyme regulated by specific subunits. The B56 (or B'/PR61/PPP2R5) class of B-subunits direct PP2A or its substrates to different cellular locations, and the B56alpha, -beta, and -epsilon isoforms are known to localize primarily in the cytoplasm. Here we studied the pathways that regulate B56alpha subcellular localization. We detected B56alpha in the cytoplasm and nucleus, and at the nuclear envelope and centrosomes, and show that cytoplasmic localization is dependent on CRM1-mediated nuclear export. The inactivation of CRM1 by leptomycin B or by siRNA knockdown caused nuclear accumulation of ectopic and endogenous B56alpha. Conversely, CRM1 overexpression shifted B56alpha to the cytoplasm. We identified a functional nuclear export signal at the C terminus (NES; amino acids 451-469), and site-directed mutagenesis of the NES (L461A) caused nuclear retention of full-length B56alpha. Active NESs were identified at similar positions in the cytoplasmic B56-beta and epsilon isoforms, but not in the nuclear-localized B56-delta or gamma isoforms. The transient expression of B56alpha induced nuclear export of the PP2A catalytic (C) subunit, and this was blocked by the L461A NES mutation. In addition, B56alpha co-located with the PP2A active (A) subunit at centrosomes, and its centrosome targeting involved sequences that bind to the A-subunit. Fluorescence Recovery after Photobleaching (FRAP) assays revealed dynamic and immobile pools of B56alpha-GFP, which was rapidly exported from the nucleus and subject to retention at centrosomes. We propose that B56alpha can act as a PP2A C-subunit chaperone and regulates PP2A activity at diverse subcellular locations.

Functional effects of intramembranous proline substitutions in the staphylococcal multidrug transporter QacA.

The QacA multidrug transporter is encoded on Staphylococcus aureus multidrug resistance plasmids and confers broad-range antimicrobial resistance to more than 30 monovalent and bivalent lipophilic, cationic compounds from at least 12 different chemical classes. QacA contains 10 proline residues predicted to be within transmembrane regions, several of which are conserved in related export proteins. Proline residues are classically known as helix-breakers and are highly represented within the transmembrane helices of membrane transport proteins, where they can mediate the formation of structures essential for protein stability and transport function. The importance of these 10 intramembranous proline residues for QacA-mediated transport function was determined by examining the functional effect of substituting these residues with glycine, alanine or serine. Several proline-substituted QacA mutants failed to confer high-level resistance to selected QacA substrates. However, no single proline mutation, including those at conserved positions, significantly disrupted QacA protein expression or QacA-mediated resistance to all representative substrates, suggesting that these residues are not essential for the formation of structures requisite to the QacA substrate transport mechanism.

Lymphoid enhancer factor-1 blocks adenomatous polyposis coli-mediated nuclear export and degradation of beta-catenin. Regulation by histone deacetylase 1.

The oncogenic protein beta-catenin is overexpressed in many cancers, frequently accumulating in nuclei where it forms active complexes with lymphoid enhancer factor-1 (LEF-1)/T-cell transcription factors, inducing genes such as c-myc and cyclin D1. In normal cells, nuclear beta-catenin levels are controlled by the adenomatous polyposis coli (APC) protein through nuclear export and cytoplasmic degradation. Transient expression of LEF-1 is known to increase nuclear beta-catenin levels by an unknown mechanism. Here, we show that APC and LEF-1 compete for nuclear beta-catenin with opposing consequences. APC can export nuclear beta-catenin to the cytoplasm for degradation. In contrast, LEF-1 anchors beta-catenin in the nucleus by blocking APC-mediated nuclear export. LEF-1 also prevented the APC/CRM1-independent nuclear export of beta-catenin as revealed by in vitro assays. Importantly, LEF-1-bound beta-catenin was protected from degradation by APC and axin in SW480 colon cancer cells. The ability of LEF-1 to trap beta-catenin in the nucleus was down-regulated by histone deacetylase 1, and this correlated with a decrease in LEF1 transcription activity. Our findings identify LEF-1 as key regulator of beta-catenin nuclear localization and stability and suggest that overexpression of LEF-1 in colon cancer and melanoma cells may contribute to the accumulation of oncogenic beta-catenin in the nucleus.