AT1R-Selective Negative Allosteric Modulator Targeting the Epitope for Pathogenic Autoantibody Activation

A major therapeutic strategy for managing cardiovascular and renal diseases is the use of angiotensin receptor blockers (ARBs). These drugs block both pathogenic and physiological signaling by the angiotensin II type 1 receptor (AT1R) by preventing angiotensin II (AngII) binding to orthosteric pocket in transmembrane portion of AT1R. However, in many autoimmune diseases, AT1R activation is chronically driven by pathogenic autoantibody binding to a distinct extracellular surface located allosteric site. Currently, there are no allosteric inhibitors of AT1R available to mechanistically prevent the autoantibodies and preserve the orthosteric hormonal functions of AngII. Here, we report the structure-guided hit-to-lead optimization of TP1, a first-in-class AT1R allosteric chemotype and negative allosteric modulator (NAM) with improved potency and drug-like properties. Computational docking and site-directed mutagenesis identified a TP1 binding mode that is distinct from the orthosteric-site binding of ARBs and AngII. Synthetically pure TP1 significantly reduced the capacity of AT1R to bind AngII without affecting AngII binding to AT2R, demonstrating an AT1R selective effect. TP1 reduced AngII-induced calcium signaling and migration in AT1R-expressing cells, arterial contraction, and increases in blood pressure in AngII-infused mice, demonstrating its activity as a NAM. Crucially, TP1 blocked binding of pathogenic autoantibodies to AT1R-expressing cells from the serum of patients with preeclampsia and dilated cardiomyopathy, a function distinct from ARBs. Together, these findings establish TP1 as a proof-of-concept, AT1R-selective NAM chemotype with therapeutic potential for AT1R autoantibody-driven conditions such as preeclampsia, where ARBs are strictly contraindicated. Developing AT1R-targeted NAM antihypertensive drugs may offer therapeutic advantages over ARBs and expand the pharmacological toolkit for this important CVD receptor.