Project Summary/Abstract Lung diseases are a global public health problem with significant morbidity, inadequate therapies and a very high burden to society. Of these, idiopathic pulmonary fibrosis (IPF) is particularly challenging to treat as existing therapeutics, Nintedanib and Pirfenidone, only slow progression but do not reverse fibrosis. One of the reasons is in part due to the fact that the vascular endothelium represents a key barrier to effective delivery of anti-fibrotic drugs into lung tissue. Current systemic lung therapies rely on passive transvascular transport to move circulating drugs from the bloodstream and into the target tissue. Hoverer, only minute fractions of the injected doses actually reach the lung tissue, diminishing efficacy and often necessitating dose escalation. The ideal goal is to deliver the entire dose of therapeutic into the lung with the minimal exposure of other tissues. We attempt to approach this ideal goal by pumping unique immunotherapeutics precisely into lung tissue to comprehensively block activation and differentiation of MF, the key driver of profibrotic extracellular matrix (ECM) deposition and activation of survival cascades, leading to progressive scarring. The goal of this project is to develop and test novel lung precision drug delivery system (LPDDS) targeting endothelial cells caveolae that are actively and specifically pump myofibroblasts (MF)-directed antifibrotics into lungs at microdoses after intravenous (iv) injection. We have identified the cell surface protein Thy-1 a precision therapeutic and a major modulator of MF activation and differentiation and characterized its role in resolving fibrosis via synergistic engagement of multiple profibrotic signaling pathways. Based on this, we engineered the first “dual precision” bispecific therapeutic mAPP:Thy-1 with antibody mediating precise binding/delivery to and penetration of lung tissue via caveolae transcytosis and the other – mediating Thy-1’s fibrosis-resolving activity in the lung interstitium where the loss of Thy-1 promotes a dysregulated MF phenotype driving fibrosis. To further potentiate antifibrotic effect, we designed mAPP:FAP-Nintedanib immunoconjugates, where concerted action of Thy-1 and Nintedanib in the lung parenchyma has even greater power to concentrate their fibrosis-resolving activities at the site of action, potentially reversing fibrosis. Our goal is to explore transendothelial pumping via caveolae to improve therapy of lung diseases at various stages from very early inflammatory to fibrotic. We will optimize lung targeting of our LPDDS through engineering and chemistry and will study their specific lung delivery, penetration, accumulation, and therapeutic impact using multiple imaging techniques (SPECT-CT, IVM, EM, and IHC). Therapeutic effects will be determined in two rat models that can reproduce many pathological hallmarks of inflammatory and fibrotic disease stages. Our specific aims are 1) to generate and te...