Alzheimer's disease and related dementia (ADRD) constitutes a growing health crisis. Equally chronic metabolicdiseases such as type 2 diabetes (T2D) are increasing because of the prevalence of obesity and other riskfactors. T2D is a risk factor for ADRD and both T2D and ADRD share common causal mechanisms: insulinresistance; impaired glucose metabolism; inflammation; dyslipidemia; and impaired cholesterol mobilization. TheAPOE4 allele is the greatest genetic risk factor for AD. ApoE4 is poorly lipidated and lipidation of apoE requiredfor stability and positive function is controlled by the ATP-binding cassette transporter ABCA1. Deletion ofABCA1 in FAD mouse models exacerbates pathology and behavioral deficits; and rare human loss-of-functionmutations in ABCA1 increase ADRD risk. ABCA1 is a gene product of liver X receptor (LXR); however inductionof lipogenesis in the liver (steatosis and triglyceride elevation) by LXR agonists has hindered progress. Anonlipogenic ABCA1-inducer (NLAI) would address multiple causal factors in T2D and ADRD including APOE4risk in AD. We have optimized a phenotypic drug discovery strategy for NLAIs yielding hit series that enhancedcholesterol mobilization attenuated inflammation and improved biomarkers of glucose metabolism. One hit andan early lead derived from it (CL2-57) increased ABCA1 and APOE without upregulating lipogenic genes. CL2-57 administered orally in the high-fat diet (HFD) model of obesogenic T2D attenuated insulin resistance reducedweight gain and from full metabolomic analysis improved biomarkers and lipid profiles. Aim 1: To optimize NLAIs.Phenotypic optimization will be driven by reporter assays (induction of ABCA1 in CCF cells with minimal effectson SREBP1c in HepG2 cells) and secondary assays in the testing funnel validated in development of CL2-57.In silico and in vitro predictors of oral/brain bioavailability and SAR will guide optimization. Validation byPCR/immunoassay will extend to ABCG1/APOE and FAS/SCD1 in cell cultures. Aim 2: In vivo PK/PD and safety.NLAI treatment of mice for 3 days LPS is sufficient to assess target engagement and pharmacodynamics inthe liver and brain with a safety readout (no triglyceride elevation nor neutropenia) suitable to define PK/PD anddosing. Aim 3: In vivo efficacy will be measured A) in 5xFAD mice (A cognition and disease-associatedmicroglia) and B) in HFD-treated mice (WT hAPOE3-KI and hAPOE4-KI) to identify APOE genotype specificinteractions with HFD and NLAI treatment in vivo and ex vivo in astrocytes and neurons. These mouse modelswill establish the efficacy of an NLAI development lead. Aim 4: Pretox and target deconvolution will be used toidentify CYP liabilities and any off-targets that will inform future safety pharmacology. The following Milestonesare proposed: #1 lead NLAI meeting in vitro TPP with brain bioavailability; #2 orally bioavailable lead NLAI thatshows dose-dependent target engagement in the brain without lipogenesis elevated triglycerides or neutropenia;#3 nomination of a development lead that meets the in vivo TPP with data on targets and CYPs.