Fibril-Targeting Nanobodies for Restoring Mitochondrial Stress and Cardiac Function in ATTR-CM - Project Summary Transthyretin amyloidosis cardiomyopathy (ATTR-CM) is a progressive, life-threatening disease caused by extracellular deposition of misfolded transthyretin (TTR) fibrils in the myocardium. These aggregates impair mitochondrial function, calcium signaling, and cardiomyocyte contractility, leading to heart failure. While stabilizers and RNAi therapies limit TTR production/stability loss, they do not remove deposited fibrils, which is the ongoing source of cardiac dysfunction. This project will define how fibrils drive mitochondrial and Ca²⁺ failure, identify host-directed drivers to nominate a co-therapy target, and develop nanobody therapeutics to clear fibrils and restore cardiac function. Using patient-anchored resources, ATTR-CM cardiac tissue, patient iPSC lines, and vascularized cardiac organoids (vCOs) benchmarked to adult human heart, with established fibril-deposition protocols, Dr. Zehra Yildirim will model disease phenotypes in a human-relevant system. Aim 1 will define how fibril deposition drives mitochondrial and Ca²⁺ dysfunction using Seahorse metabolic analysis, Cryo-FIB/SEM, optical mapping, and MEA electrophysiology, and will integrate multi-omics and quantitative proteomics to map time-dependent, TTR-linked niche maturation (apolipoprotein coat, complement/ECM, lysosome) and associated respiratory-chain attrition and oxidative/iron stress consistent with functional readouts. Aim 2 will leverage AI-guided protein engineering combined with a pre-existing nanobody repertoire and yeast-display selection to generate high-specificity anti-fibril nanobodies with multiple high-priority binders readily identified and are advancing through expression and developability testing, de-risking translation. Aim 3 will evaluate safety and efficacy of the top candidate in preclinical organoid and mouse models. Dr. Yildirim’s career development plan integrates training in computational biology, AI-driven drug discovery, and structural biology to complement her expertise in cardiovascular molecular biology and bioengineering. Under the primary mentorship of Dr. Joseph Wu (iPSC disease modeling) and co-mentorship of Dr. Possu Huang (AI-driven protein engineering), she will gain expertise in protein engineering, structural proteomics, and therapeutic development. Collaboration with Dr. Wah Chiu at SLAC will provide hands-on Cryo-FIB/SEM training. Externships in Dr. Huang’s lab (AI nanobody design) and Dr. Chiu’s lab (mitochondrial structural analysis), together with coursework in bioinformatics, structural biology, and translational research, will position her to launch an independent research program. Successful completion will clarify ATTR-CM pathogenesis, deliver a translatable nanobody platform for amyloidosis, and accelerate clinical translation of AI-designed therapeutics, improving patient outcomes and advancing precision cardiovascular medicine. This integrated, human-anchored platform and validated binder pipeline directly addresses the unmet need for fibril clearance in ATTR-CM and de-risks the K99 to R00 transition with clear, measurable milestones.
