A Randomized Trial Examining the Effectiveness of Switching From Olanzapine, Quetiapine, or Risperidone to Aripiprazole to Reduce Metabolic Risk: Comparison of Antipsychotics for Metabolic Problems (CAMP)

We conducted a multi-site, randomized controlled trial examining the strategy of switching from olanzapine, quetiapine, or risperidone to aripiprazole to ameliorate metabolic risk factors for cardiovascular disease

Genomic profile of advanced breast cancer in circulating tumour DNA.

The genomics of advanced breast cancer (ABC) has been described through tumour tissue biopsy sequencing, although these approaches are limited by geographical and temporal heterogeneity. Here we use plasma circulating tumour DNA sequencing to interrogate the genomic profile of ABC in 800 patients in the plasmaMATCH trial. We demonstrate diverse subclonal resistance mutations, including enrichment of HER2 mutations in HER2 positive disease, co-occurring ESR1 and MAP kinase pathway mutations in HR + HER2- disease that associate with poor overall survival (p = 0.0092), and multiple PIK3CA mutations in HR + disease that associate with short progression free survival on fulvestrant (p = 0.0036). The fraction of cancer with a mutation, the clonal dominance of a mutation, varied between genes, and within hotspot mutations of ESR1 and PIK3CA. In ER-positive breast cancer subclonal mutations were enriched in an APOBEC mutational signature, with second hit PIK3CA mutations acquired subclonally and at sites characteristic of APOBEC mutagenesis. This study utilises circulating tumour DNA analysis in a large clinical trial to demonstrate the subclonal diversification of pre-treated advanced breast cancer, identifying distinct mutational processes in advanced ER-positive breast cancer, and novel therapeutic opportunities

Comparative transcriptome profiling of amyloid precursor protein family members in the adult cortex

<p>Abstract</p> <p>Background</p> <p>The β-amyloid precursor protein (APP) and the related β-amyloid precursor-like proteins (APLPs) undergo complex proteolytic processing giving rise to several fragments. Whereas it is well established that Aβ accumulation is a central trigger for Alzheimer's disease, the physiological role of APP family members and their diverse proteolytic products is still largely unknown. The secreted APPsα ectodomain has been shown to be involved in neuroprotection and synaptic plasticity. The γ-secretase-generated APP intracellular domain (AICD) functions as a transcriptional regulator in heterologous reporter assays although its role for endogenous gene regulation has remained controversial.</p> <p>Results</p> <p>To gain further insight into the molecular changes associated with knockout phenotypes and to elucidate the physiological functions of APP family members including their proposed role as transcriptional regulators, we performed DNA microarray transcriptome profiling of prefrontal cortex of adult wild-type (WT), APP knockout (APP^-/-), APLP2 knockout (APLP2^-/-) and APPsα knockin mice (APP^α/α) expressing solely the secreted APPsα ectodomain. Biological pathways affected by the lack of APP family members included neurogenesis, transcription, and kinase activity. Comparative analysis of transcriptome changes between mutant and wild-type mice, followed by qPCR validation, identified co-regulated gene sets. Interestingly, these included heat shock proteins and plasticity-related genes that were both down-regulated in knockout cortices. In contrast, we failed to detect significant differences in expression of previously proposed AICD target genes including <it>Bace1</it>, <it>Kai1</it>, <it>Gsk3b</it>, <it>p53</it>, <it>Tip60</it>, and <it>Vglut2</it>. Only <it>Egfr </it>was slightly up-regulated in APLP2^-/-mice. Comparison of APP^-/-and APP^α/αwith wild-type mice revealed a high proportion of co-regulated genes indicating an important role of the C-terminus for cellular signaling. Finally, comparison of APLP2^-/-on different genetic backgrounds revealed that background-related transcriptome changes may dominate over changes due to the knockout of a single gene.</p> <p>Conclusion</p> <p>Shared transcriptome profiles corroborated closely related physiological functions of APP family members in the adult central nervous system. As expression of proposed AICD target genes was not altered in adult cortex, this may indicate that these genes are not affected by lack of APP under resting conditions or only in a small subset of cells.</p

Pathogenic Variant Filtering for Mitochondrial Genome Haplotype Reporting

Given the enhanced discriminatory power of the mitochondrial DNA (mtDNA) genome (mitogenome) over the commonly sequenced control region (CR) portion, the scientific merit of mitogenome sequencing is generally accepted. However, many laboratories remain beholden to CR sequencing due to privacy policies and legal requirements restricting the use of disease information or coding region (codR) information. In this report, we present an approach to obviate the reporting of sensitive codR data in forensic haplotypes. We consulted the MitoMap database to identify 92 mtDNA codR variants with confirmed pathogenicity. We determined the frequencies of these pathogenic variants in literature-quality and forensic-quality databases to be very low, at 1.2% and 0.36%, respectively. The observed effect of pathogenic variant filtering on random match statistics in 2488 forensic-quality mitogenome haplotypes from four populations was nil. We propose that pathogenic variant filtering should be incorporated into variant calling algorithms for mitogenome haplotype reporting to maximize the discriminatory power of the locus while minimizing the reveal of sensitive genetic information

The Value of Whole-Genome Sequencing for Mitochondrial DNA Population Studies : Strategies and Criteria for Extracting High-Quality Mitogenome Haplotypes

Whole-genome sequencing (WGS) data present a readily available resource for mitochondrial genome (mitogenome) haplotypes that can be utilized for genetics research including population studies. However, the reconstruction of the mitogenome is complicated by nuclear mitochondrial DNA (mtDNA) segments (NUMTs) that co-align with the mtDNA sequences and mimic authentic heteroplasmy. Two minimum variant detection thresholds, 5% and 10%, were assessed for the ability to produce authentic mitogenome haplotypes from a previously generated WGS dataset. Variants associated with NUMTs were detected in the mtDNA alignments for 91 of 917 (~8%) Swedish samples when the 5% frequency threshold was applied. The 413 observed NUMT variants were predominantly detected in two regions (nps 12,612–13,105 and 16,390–16,527), which were consistent with previously documented NUMTs. The number of NUMT variants was reduced by ~97% (400) using a 10% frequency threshold. Furthermore, the 5% frequency data were inconsistent with a platinum-quality mitogenome dataset with respect to observed heteroplasmy. These analyses illustrate that a 10% variant detection threshold may be necessary to ensure the generation of reliable mitogenome haplotypes from WGS data resources

Platinum-Quality Mitogenome Haplotypes from United States Populations

A total of 1327 platinum-quality mitochondrial DNA haplotypes from United States (U.S.) populations were generated using a robust, semi-automated next-generation sequencing (NGS) workflow with rigorous quality control (QC). The laboratory workflow involved long-range PCR to minimize the co-amplification of nuclear mitochondrial DNA segments (NUMTs), PCR-free library preparation to reduce amplification bias, and high-coverage Illumina MiSeq sequencing to produce an average per-sample read depth of 1000 &times; for low-frequency (5%) variant detection. Point heteroplasmies below 10% frequency were confirmed through replicate amplification, and length heteroplasmy was quantitatively assessed using a custom read count analysis tool. Data analysis involved a redundant, dual-analyst review to minimize errors in haplotype reporting with additional QC checks performed by EMPOP. Applying these methods, eight sample sets were processed from five U.S. metapopulations (African American, Caucasian, Hispanic, Asian American, and Native American) corresponding to self-reported identity at the time of sample collection. Population analyses (e.g., haplotype frequencies, random match probabilities, and genetic distance estimates) were performed to evaluate the eight datasets, with over 95% of haplotypes unique per dataset. The platinum-quality mitogenome haplotypes presented in this study will enable forensic statistical calculations and thereby support the usage of mitogenome sequencing in forensic laboratories

Abstract PS5-02: Assessment of early ctDNA dynamics to predict efficacy of targeted therapies in metastatic breast cancer: Results from plasmaMATCH trial

Abstract Background: Early changes in circulating tumour DNA (ctDNA) levels may identify which patients respond to therapy earlier than imaging, with ctDNA levels falling rapidly in patients who respond to therapy. The plasmaMATCH trial assessed the utility of ctDNA testing with an error-corrected 73-gene targeted panel (Guardant360, Guardant Health) to allocate patients to four mutation matched therapy cohorts. ESR1-extended fulvestrant (A), HER2-neratinib +/- fulvestrant (B), AKT1-capivasertib + fulvestrant (C), AKT basket-capivasertib (D). Here, we report paired baseline and early on treatment ctDNA analysis from plasmaMATCH, to establish the optimal criteria for predicting progression free survival (PFS). Methods: In plasmaMATCH treatment cohorts, plasma samples were collected for ctDNA analysis pre-treatment at cycle 1-day 1 (C1D1) and cycle 2-day 1 (C2D1) timepoints, and sequenced with the Guardant 360 assay. Patients were included if they had a minimum of 14 days of treatment in the first cycle. Multiple different methods were investigated to integrate variant allele fractions (VAF) of mutations identified at each timepoint to estimate the level of ctDNA, including maximum VAF, mean VAF and weighted mean VAF, and weighted mean VAF of clonal mutations at C1D1. Variants with a VAF &amp;lt;0.3%, set as the limit of detection, in C1D1 were excluded. Genes frequently mutated in CHIP were excluded (GNAS, JAK2, IDH1, IDH2 and ATM) from the weighted mean VAF of clonal mutations method. The circulating DNA ratio (CDR) was calculated as the ratio of C2D1 level relative to C1D1 level. The optimal cut-point for predicting PFS was assessed by fitting a range of cutpoints for each VAF integration method, identifying the cut-point with the highest Harrell’s C-index. Results: A total of 142 patients were enrolled into plasmaMATCH cohorts A-D, 79 patients had samples sent for paired C1D1-C2D1 plasma ctDNA sequencing, 1 failed sequencing and 1 insufficient treatment, and 77 (54%) patients had assessable C1D1-C2D1 plasma ctDNA sequencing results (45 cohort A, 12 cohort B, 12 cohort C, 8 cohort D). A weighted mean of clonal mutations in C1D1 ctDNA sequencing was the optimal method for integrating VAF, with peak C-Index 0.67. At the optimal C-index cutoff of 0.132, median PFS with high ctDNA CDR was 2.4 months (95% CI 2.0-3.7) and with suppressed ctDNA CDR was 9.9 months (95% CI 7.0-13.7) (HR 4.3, 95% CI 2.4-7.6, p&amp;lt;0.0001). Early changes in ctDNA level were also predictive in cohorts A extended dose fulvestrant alone (HR 5.8, 95% CI 2.2-16, p=0.0001) and cohorts B-D of targeted therapy (HR 3.8, 95% CI 1.7-8.6, p=0.00063). In analysis that was not pre-planned, patients with undetectable ctDNA at C2D1 had a particularly good outcome (p&amp;lt;0.0001, table 1). Conclusions: We identify an optimal methodology for assessing early dynamic changes in ctDNA that predicts treatment efficacy in patients with metastatic breast cancer. This methodology will require validation in independent data-sets, and if validated would allow trials of adapting therapy on the basis of early ctDNA dynamics. Table 1ctDNA dynamics categoryMedian PFS months (95%CI)6-month PFSORRUndetectable (N=11) CDR=018.2 (10.2-NA)91%9/11 (82%)Suppressed (N=14) CDR &amp;lt;0.132 and &amp;gt;05.4 (4.6-NA)48%6/14 (43%)High (N=52) CDR &amp;gt;=0.1322.4 (2.0-3.7)8%4/52 (8%) Citation Format: Javier Pascual, Rosalind J Cutts, Belinda Kingston, Sarah Hrebien, Lucy S Kilburn, Sarah Kernaghan, Laura Moretti, Katie Wilkinson, Andrew M Wardley, Iain R Macpherson, Richard D Baird, Rebecca Roylance, Michael Hubank, Giselle Walsh, Iris Faull, Kimberly C Banks, Richard B Lanman, Isaac Garcia-Murillas, Judith M Bliss, Alistair Ring, Nicholas C Turner. Assessment of early ctDNA dynamics to predict efficacy of targeted therapies in metastatic breast cancer: Results from plasmaMATCH trial [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS5-02.</jats:p

SARS-CoV-2 mitochondriopathy in COVID-19 pneumonia exacerbates hypoxemia

Rationale: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes COVID-19 pneumonia. We hypothesize that SARS-CoV-2 causes alveolar injury and hypoxemia by damaging mitochondria in airway epithelial cells (AEC) and pulmonary artery smooth muscle cells (PASMC), triggering apoptosis and bioenergetic impairment, and impairing hypoxic pulmonary vasoconstriction (HPV), respectively. Objectives: We examined the effects of: A) human betacoronaviruses, SARS-CoV-2 and HCoV-OC43, and individual SARS-CoV-2 proteins on apoptosis, mitochondrial fission, and bioenergetics in AEC; and B) SARS-CoV-2 proteins and mouse hepatitis virus (MHV-1) infection on HPV. Methods: We used transcriptomic data to identify temporal changes in mitochondrial-relevant gene ontology (GO) pathways post-SARS-CoV-2 infection. We also transduced AECs with SARS-CoV-2 proteins (M, Nsp7 or Nsp9) and determined effects on mitochondrial permeability transition pore (mPTP) activity, relative membrane potential, apoptosis, mitochondrial fission, and oxygen consumption rates (OCR). In human PASMC, we assessed the effects of SARS-CoV-2 proteins on hypoxic increases in cytosolic calcium, an HPV proxy. In MHV-1 pneumonia, we assessed HPV via cardiac catheterization and apoptosis using the TUNEL assay. Results: SARS-CoV-2 regulated mitochondrial apoptosis, mitochondrial membrane permeabilization and electron transport chain (ETC) GO pathways within 2 hours of infection. SARS-CoV-2 downregulated ETC Complex I and ATP synthase genes, and upregulated apoptosis-inducing genes. SARS-CoV-2 and HCoV-OC43 upregulated and activated dynamin-related protein 1 (Drp1) and increased mitochondrial fission. SARS-CoV-2 and transduced SARS-CoV-2 proteins increased apoptosis inducing factor (AIF) expression and activated caspase 7, resulting in apoptosis. Coronaviruses also reduced OCR, decreased ETC Complex I activity and lowered ATP levels in AEC. M protein transduction also increased mPTP opening. In human PASMC, M and Nsp9 proteins inhibited HPV. In MHV-1 pneumonia, infected AEC displayed apoptosis and HPV was suppressed. BAY K8644, a calcium channel agonist, increased HPV and improved SpO2. Conclusions: Coronaviruses, including SARS-CoV-2, cause AEC apoptosis, mitochondrial fission, and bioenergetic impairment. SARS-CoV-2 also suppresses HPV by targeting mitochondria. This mitochondriopathy is replicated by transduction with SARS-CoV-2 proteins, indicating a mechanistic role for viral-host mitochondrial protein interactions. Mitochondriopathy is a conserved feature of coronaviral pneumonia that may exacerbate hypoxemia and constitutes a therapeutic target