From Native to Amyloid in the Test Tube and in Cells: A Journey of Misbehaving Antibodies

Abstract

Light chain (AL) amyloidosis is an incurable disease characterized by the misfolding, aggregation, and systemic deposition of amyloid composed of immunoglobulin light chains. We have conducted thermodynamic and fibril formation studies of both variable domain and full length immunoglobulin proteins involved in AL amyloidosis. We have found that there is thermodynamic range -under physiological conditions- in which these proteins populate partially folded states that favor amyloid formation.We have studied the internalization of these same AL proteins in soluble form and as amyloid fibrils into human cardiomyocytes. Our results show how external amyloid aggregates rapidly surround the cells and act as a recruitment point for soluble protein, triggering amyloid fibril elongation. A fraction of aggregates surrounding the cardiomyocytes is internalized via micropinocytosis (same mechanism followed by soluble proteins). AL amyloid fibrils are shown to cause cell growth arrest at low concentration. Soluble proteins induce apoptosis, demonstrating different cytotoxic mechanisms between soluble protein and amyloid aggregates.In vitro amyloid formation experiments show that heterologous recruitment of light chains is kinetically determined by the conformation of the amyloidogenic conformational precursor and modulated by the differential ability of each protein to either nucleate or elongate fibrils.Studies on the early events of fibril formation followed by dynamic light scattering, chromatography and electron microscopy have shown differences in the species formed by amyloidogenic proteins (stable small oligomers) compared to control proteins (large oligomers observed).Overall, our studies emphasize the complex interactions between light chain and cells that result in fibril internalization, protein recruitment, and cytotoxicity that may occur in AL amyloidosis.