Abstract: The hallmarks of chronic kidney disease (CKD) include fibrosis, increased inflammatory responses, and dysregulated mineral metabolism, which are all especially problematic in aged patients and strongly associated with morbidity and mortality. Indeed, the fibrotic lesions arising in humans as well as CKD rodent models cause dramatic decreases in renal function, leading to further cycles of inflammation and activation of myofibroblasts with collagen accumulation. The bone-produced hormone fibroblast growth factor- 23 (FGF23) controls phosphate and vitamin D metabolism, and is markedly increased in late-stage CKD (>1000-fold), in part due to loss of its co-receptor, Klotho (KL). The dysregulation of FGF23 results in severe endocrine bone disease in aged patients whose musculoskeletal system is already compromised, and it is not known whether this pathologic phenotype can be ameliorated with improvement of kidney function. There are no direct therapies for CKD fibrosis, and it remains unclear whether targeting fibrosis using broad spectrum drugs is feasible in clinical practice because most drugs that decrease fibrosis in pre-clinical models target multiple essential pathogenic pathways (eg, renin-angiotensin-aldosterone system blockade). Fibroblast activation protein (FAP) is a transmembrane serine protease expressed by activated fibroblasts in wound healing and pathological conditions such as fibrosis and cancer. Our collaborator, Dr. Jonathan Epstein, developed a transient anti-FAP Chimeric Antigen Receptor (CAR)-T cell mRNA therapy (or ‘FAPCAR’) which reduced cardiac fibrosis following infarction. FAP is an optimal target for CAR-T cell targeting as it is a cell surface molecule only expressed during pathology and not at high levels under normal conditions. To treat fibrosis, lipid nanoparticles (LNPs) designed to target CD5-positive T cells are endocytosed, and the mRNA encoding FAPCAR translated within the cell. The advantage of this approach is that the generated in vivo fibrosis-targeted CAR-T cells do not require individualized patient derived cells, and there is no danger of random T-cell gene interruption as in standard CAR-T therapy. Our central hypothesis is: CD5/LNP-FAPCAR pre-clinical therapy will directly target renal fibrosis and improve renal function and mineral metabolism either alone or in combination treatment in aged mice with CKD. Considering the magnitude of the negative effects of fibrosis on CKD outcomes in the aging population, and the fact there are no FDA-approved therapies directly resolving fibrosis, our studies are important for the CKD field. Thus we will address our hypothesis under Specific Aims: 1. To test preclinical CD5/LNP-FAPCAR therapy in prevention and treatment of CKD fibrosis in aged and adult mice; and 2.To test FAP-expressing fibroblast ablation as a combination therapy. Thus, our proposed studies are innovative as we will test for the first time whether a novel transient LNP-FAPCAR therapy can...