A constant domain mutation in a patient-derived antibody light chain reveals principles of AL amyloidosis.

Light chain (AL) amyloidosis is a debilitating disease in which mutant antibody light chains (LC), secreted by aberrant plasma cell clones, misfold and form insoluble fibrils, which can be deposited in various organs. In the majority of cases, the fibrillar deposits consist of LC variable domains (VL) containing destabilizing mutations compared to their germline counterparts. This is also true for the patient LC FOR005. However, this pathogenic LC sequence contains an additional mutation in the constant domain (CL). The mechanistic impact of CL mutations is not yet understood in the context of AL amyloidosis. Our analysis reveals that the FOR005 CL mutation influences the amyloid pathway in specific ways: (1) folding and stability of the patient CL domain are strongly impaired; (2) the mutation disrupts the LC dimer interface and weakens dimerization; (3) the CL mutation promotes proteolytic cleavage of the LC monomers resulting in an isolated, amyloidogenic VL domain while dimeric LCs are not cleaved. The enhanced proteolysis rates and the inability of full-length LCs to form amyloid fibrils even in the presence of a destabilized CL domain support a model for AL amyloidosis in which the CL domain plays a protective role and in which proteolytic cleavage precedes amyloid formation.

Antibodies gone bad - the molecular mechanism of light chain amyloidosis.

Light chain amyloidosis (AL) is a systemic disease in which abnormally proliferating plasma cells secrete large amounts of mutated antibody light chains (LCs) that eventually form fibrils. The fibrils are deposited in various organs, most often in the heart and kidney, and impair their function. The prognosis for patients diagnosed with AL is generally poor. The disease is set apart from other amyloidoses by the huge number of patient-specific mutations in the disease-causing and fibril-forming protein. The molecular mechanisms that drive the aggregation of mutated LCs into fibrils have been enigmatic, which hindered the development of efficient diagnostics and therapies. In this review, we summarize our current knowledge on AL amyloidosis and discuss open issues.

MALDI mass spectrometry imaging unravels organ and amyloid-type specific peptide signatures in pulmonary and gastrointestinal amyloidosis.

PURPOSE: Amyloidosis is a disease group caused by pathological aggregation and deposition of peptides in diverse tissue sites. Recently, matrix-assisted laser desorption/ionization mass spectrometry imaging coupled with ion mobility separation (MALDI-IMS MSI) was introduced as a novel tool to identify and classify amyloidosis using single sections from formalin-fixed and paraffin-embedded cardiac biopsies. Here, we tested the hypothesis that MALDI-IMS MSI can be applied to lung and gastrointestinal specimens. EXPERIMENTAL DESIGN: Forty six lung and 65 gastrointestinal biopsy and resection specimens with different types of amyloid were subjected to MALDI-IMS MSI. Ninety three specimens included tissue areas without amyloid as internal negative controls. Nine cases without amyloid served as additional negative controls. RESULTS: Utilizing a peptide filter method and 21 known amyloid specific tryptic peptides we confirmed the applicability of a universal peptide signature with a sensitivity of 100% and a specificity of 100% for the detection of amyloid deposits in the lung and gastrointestinal tract. Additionally, the frequencies of individual m/z-values of the 21 tryptic marker peptides showed organ- and tissue-type specific differences. CONCLUSIONS AND CLINICAL RELEVANCE: MALDI-IMS MSI adds a valuable analytical approach to diagnose and classify amyloid and the detection frequency of individual tryptic peptides is organ-/tissue-type specific.

Molecular mechanism of amyloidogenic mutations in hypervariable regions of antibody light chains.

Systemic light chain (AL) amyloidosis is a fatal protein misfolding disease in which excessive secretion, misfolding, and subsequent aggregation of free antibody light chains eventually lead to deposition of amyloid plaques in various organs. Patient-specific mutations in the antibody VL domain are closely linked to the disease, but the molecular mechanisms by which certain mutations induce misfolding and amyloid aggregation of antibody domains are still poorly understood. Here, we compare a patient VL domain with its nonamyloidogenic germline counterpart and show that, out of the five mutations present, two of them strongly destabilize the protein and induce amyloid fibril formation. Surprisingly, the decisive, disease-causing mutations are located in the highly variable complementarity determining regions (CDRs) but exhibit a strong impact on the dynamics of conserved core regions of the patient VL domain. This effect seems to be based on a deviation from the canonical CDR structures of CDR2 and CDR3 induced by the substitutions. The amyloid-driving mutations are not necessarily involved in propagating fibril formation by providing specific side chain interactions within the fibril structure. Rather, they destabilize the VL domain in a specific way, increasing the dynamics of framework regions, which can then change their conformation to form the fibril core. These findings reveal unexpected influences of CDR-framework interactions on antibody architecture, stability, and amyloid propensity.