Small molecule stabilization of diverse amyloidogenic immunoglobulin light chains revealed by hydrogen-deuterium exchange mass spectrometry

Abstract Immunoglobulin light chains, a component of antibodies, can misfold and aggregate to cause systemic AL amyloidosis. Aggregation, including amyloid fibril formation, requires unfolding of the full-length light chain from its native state, and in most cases aberrant proteolysis. Small molecules that bind to the native state of light chains to stabilize them against conformational excursions and proteolysis are under development as drug candidates for AL amyloidosis. Since each patient has a unique light chain sequence, a challenge for candidate stabilizer drugs is to bind multiple light chains and suppress their dynamics. Here, we used hydrogen-deuterium exchange measured by mass spectrometry to characterize the binding of six small molecule stabilizers to eleven different λ light chain proteins. Despite structural and dynamic differences among the light chains, the binding of the most efficacious stabilizer molecule led to increased protection from hydrogen exchange, consistent with reduced local and global unfolding. Protection upon binding was most prominent in residues within complementarity determining region 3 and framework region 4 of the light chain variable domains, which undergo major conformational changes enabling amyloid formation. Stabilizer binding also reduced the rate at which all light chains were cleaved by protease. These data show that these stabilizers suppress the range of conformational dynamics associated with light chain aggregation, supporting their therapeutic potential. Highlights Small-molecule kinetic stabilizers suppress conformational dynamics and aberrant proteolysis of diverse amyloidogenic full-length λ light chains. HDX-MS indicates conserved protection in regions forming the interface between the two variable domains of the light chain dimer, corresponding to residues that form the core of patient-derived amyloid fibrils. PLIMSTEX binding titrations show coupled binding, dimerization and stabilization at sub-micromolar kinetic stabilizer concentrations. Small molecule kinetic stabilization across different λ sequences supports kinetic stabilizers as a promising therapeutic strategy for AL amyloidosis.