{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Mzml":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_8Fractions.mzML"],"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F8.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F6.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F4.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F5.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_F7.raw"],"Mztab":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/03/PXD060171/wc_QE_053023_John_Whitaker_DDL357_InVivo_TMT_8Fractions.mzTab"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"labhead_mail":["VJohn@mednet.ucla.edu"],"submitter":["Jesus Campagna"],"technology_type":["Mass Spectrometry","Bottom-up proteomics"],"software":[""],"submitter_keywords":["Memory improvement","Secreted clustering enhancer","Epigenetic modulator","Alzheimers disease"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD060171"],"tissue":["Brain"],"sample_protocol":["Hippocampus were homogenized in lysis buffer (12 mM sodium lauroyl sarcosine, 0.5% sodium deoxycholate, 50 mM triethylammonium bicarbonate (TEAB), Halt™ Protease and Phosphatase Inhibitor Cocktail (Thermo Fisher Scientific)), the samples were reduced and alkylated with tris(2-carboxyethyl)phosphine (10 mM) and chloroacetamide (40 mM) for 30 minutes at 95 o C, and then digested with trypsin (20 ug) for 16 hours at 37 °C. The samples were then desalted using a modified version of Rappsilber's protocol [Ref. 1] and were isotopically labeled (TMT18plex Isobaric Label Reagent Set, Thermo Fisher Scientific) according to the manufacturer’s protocol to provide relative quantitation between samples. The samples were then fractionated separately via high pH reversed-phase chromatography (Pierce™ High pH Reversed-Phase Peptide Fractionation Kit) as per manufacturer’s protocol for increased proteome coverage.  Aliquots of each fraction were injected onto a reverse phase nanobore HPLC column (AcuTech Scientific, C18, 1.8um particle size, 360 um x 20 cm, 150 um ID), equilibrated in solvent A (water/acetonitrile/FA, 98/2/0.1, v/v/v) and eluted (300 nL/min) with an increasing concentration of solvent B (acetonitrile/water/FA, 98/2/0.1, v/v/v: min/% F; 0/0, 5/3, 18/7, 74/12, 144/24, 153/27, 162/40, 164/80, 174/80, 176/0, 180/0) using an EASY-nLC II (Thermo Fisher Scientific). The effluent from the column was directed to a nanospray ionization source connected to a hybrid quadrupole-Orbitrap mass spectrometer (Q Exactive Plus, Thermo Fisher Scientific) acquiring mass spectra in a data-dependent mode alternating between a full scan (m/z 350-1700, automated gain control (AGC) target 3 x 106, 50 ms maximum injection time, FWHM resolution 70,000 at m/z 200) and up to 15 MS/MS scans (quadrupole isolation of charge states 2-7, isolation window 0.7 m/z) with previously optimized fragmentation conditions (normalized collision energy of 32, dynamic exclusion of 30 s, AGC target 1 x 105, 100 ms maximum injection time, FWHM resolution 35,000 at m/z 200)."],"repository":["Pride"],"quantification_method":[""],"modification":[""],"data_protocol":["Raw proteomic data were searched against a Uniprot database containing the complete human proteome using SEQUEST-HT (including dynamic modifications: oxidation (+15.995) on M, deamidation (+0.984) on N/Q, and carbamidomethyl (+57.021), phosphorylation (+79.966) on S/T/Y) in Proteome Discoverer (Version 2,4, Thermo Scientific), which provided measurements of relative abundance of the identified peptides. Decoy database searching was used to generate high confidence tryptic peptides (FDR < 1%). Tryptic peptides containing amino acid sequences unique to individual proteins were used to identify and provide relative quantification between different proteins in each sample. Post-translationally modified peptides from each protein were normalized to protein abundance and peptides exhibiting a p-value ≤ 0.05 with a log2-fold change ≥ 0.5 were analyzed using a series of bioinformatics tools including functional protein association network analysis, comprehensive gene set enrichment gene ontology (GO) classification and pathway analysis, as well as kinase substrate enrichment analyses"],"omics_type":["Proteomics"],"labhead":["Varghese John"],"instrument_platform":[""],"labhead_affiliation":["The Drug Discovery Lab, Mary S. Easton Center for Alzheimer’s Disease Research, Department of Neurology, David Geffen School of Medicine, 710 Westwood Plaza, University of California Los Angeles, Los Angeles, CA 90095, USA"],"submission_type":["COMPLETE"],"species":["Mus Musculus (mouse)"],"publication":["40322539 Cohn W, Campagna J, Wi D, Lee JT, Beniwal S, Elezi G, Zhu C, Jagodzinska B, Whitelegge J, Damoiseaux R, John V. Discovery of a small molecule secreted clusterin enhancer that improves memory in Alzheimer's disease mice. NPJ Drug Discov. 2025 2(1):7 10.1038/s44386-025-00009-2"],"submitter_mail":["jcampagna@mednet.ucla.edu"],"submitter_affiliation":["UCLA"],"submitter_country":["United States"],"doi":["10.6019/PXD060171"],"pubmed_abstract":["Despite substantial research and drug discovery efforts, Alzheimer's Disease (AD) remains the sixth leading cause of death in the United States, underscoring the urgent need for novel therapeutic targets. A mutation in the clusterin (CLU) gene that hinders expression of the cyto-protective secreted isoform of clusterin (sCLU) that affects the aggregation and clearance of two key proteins implicated in AD, Aβ and tau, is the third most significant genetic risk factor for late-onset AD. Here, we present findings from our drug discovery program to identify small molecules that enhance sCLU levels and assess their impact on AD pathology and cognition in a murine model of AD. A high-throughput screening campaign identified two classes of epigenetic modulators that increase sCLU levels with subsequent medicinal chemistry efforts leading to bromodomain and extra-terminal (BET) inhibitor new chemical entities (NCEs) with enhanced potency, drug-like properties, and oral brain bioavailability. The lead candidate NCE, DDL-357, increased brain sCLU in the murine ApoE4TR-5XFAD model of AD in a subchronic study. In a follow-up chronic study in the murine 3xTg-AD model, DDL-357 reduced phospho-tau in brain and led to improvements in mouse performance and memory in the Barnes maze testing paradigm. Proteomic analysis of brain tissue from both AD models revealed changes in proteins involved in mitochondrial function and synaptic plasticity. These findings reveal the potential of sCLU enhancement as a target for therapeutic development in AD and support the continued development of the preclinical lead candidate."],"pubmed_title":["Discovery of a small molecule secreted clusterin enhancer that improves memory in Alzheimer's disease mice."],"pubmed_authors":["Cohn Whitaker W, Campagna Jesus J, Wi Dongwook D, Lee Jessica T JT, Beniwal Sahiba S, Elezi Gazmend G, Zhu Chunni C, Jagodzinska Barbara B, Whitelegge Julian J, Damoiseaux Robert R, John Varghese V"],"additional_accession":[]},"is_claimable":false,"name":"Discovery of a Small Molecule Secreted Clustering Enhancer that Improves Memory in Alzheimer's Disease Mice","description":"Here, we present findings from a drug discovery program aimed at developing small molecules that enhance brain sCLU levels and assessed their impact on AD pathology and cognition. A high-throughput screening campaign identified two classes of epigenetic modulators to increase sCLU levels, and medicinal chemistry efforts led to the development BET inhibitor new chemical entities with enhanced potency, drug-like properties, and oral brain bioavailability. Lead candidate, DDL-357, increased brain sCLU as well as proteins crucial for mitochondrial function, synaptic plasticity, and disease relevant protein aggregate clearance in AD mouse models. DDL-357 reduced p-tau and improved memory, presenting a promising multifaceted therapeutic approach for Alzheimer's 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