ABSTRACT. Cardiovascular disease is the leading cause of death worldwide. A major goal of pharmacologic therapy is to lower plasma lipids, especially low-density lipoprotein cholesterol (LDL), the cardinal risk factor for coronary artery disease (CAD). Patients with familial hypercholesterolemia (FH) have increased risk of developing early-onset CAD due to dysfunctional clearance of LDL from circulation and increased plasma levels of this atherogenic lipoprotein. Underdiagnosis and undertreatment of FH are exacerbated by limited therapeutic options. However, one strategy has been successful at lowering LDL in FH patients: Inhibition of the hepatically secreted protein Angiopoietin-like protein 3 (ANGPTL3). A monoclonal antibody (evinacumab) inhibiting circulating ANGPTL3 recently obtained FDA approval for treatment of FH due to substantial reductions in LDL, even in patients with complete deficiency of the LDL receptor (LDLR). In contrast, existing lipid-lowering agents largely rely on LDLR function to remove this atherogenic lipoprotein from the blood. In accordance with the NHLBI Research Priorities, it is important for researchers to define molecular characteristics that can predict meaningful or inadequate responses to therapy in different populations with cardiovascular disease. Despite promising clinical trial data, the LDLR-independent mechanisms by which ANGPTL3 inhibition lowers LDL have not been fully characterized. This is an important question to address not only to describe how drugs inhibiting ANGPTL3 work but also to recommend targets for reducing plasma lipids via a novel LDLR-independent pathway of lipoprotein metabolism. The main hypothesis of this proposal is that ANGPTL3 deficiency lowers LDL in part by modulating hepatic lipoprotein assembly and lipid metabolism while simultaneously altering characteristics of secreted VLDL particles. This hypothesis will be addressed by achieving the goals outlined in this proposal. The proposed experiments employ a combination of hepatocyte cell culture systems, including immortalized cancer cells and an advanced cell culture model that produces hepatocyte-like cells (HLCs) from induced pluripotent stem cells (iPSCs). These iPSCs are derived from a unique population of human subjects with complete genetic ANGPTL3 deficiency. Aim 1 will use these models to test whether changes in lipoprotein secretory transit and/or lipid metabolism contribute to LDL lowering in ANGPTL3 deficiency. Aim 2 will test whether lipoprotein particle secretion kinetics, clearance, size, and/or lipid composition could also be contributing to LDL lowering. These experiments will be carried out as part of a rigorous fellowship training plan in a well-resourced and highly collaborative environment. Consistent with the NHLBI Strategic Plan, the proposed studies will help future clinician-investigators and the patients they serve by improving understanding of the functions of ANGPTL3- inhibiting drugs; describing a role f...