The overall goal of this proposal is to identify the misfolded cytotoxic species populated by light chain amyloidosis (AL) proteins. AL is a rare, fatal misfolding disease characterized by the proliferation of monoclonal plasma cells that secrete immunoglobulin light chains that misfold as amyloid deposits in vital organs. Current treatments reduce the population of the malignant monoclonal plasma cells. Little is known about the misfolding process converting soluble proteins into insoluble AL fibrils and the nature of the toxic species generated during this process;identifying the toxic species is crucial to developing targeted therapies for AL. We have determined that the protein AL-09 forms an altered dimer interface. A single somatic mutation is responsible for the change in the dimer interface, causing loss of thermodynamic stability, and promoting amyloid formation with respect to its germline ('wild type') protein. Our preliminary data show that AL-09 is the most amyloidogenic AL protein studied in our laboratory, it is highly toxic to cardiomyocytes in its soluble form, and its amyloid fibril core is consistent with the altered dimer interface structure. In addition, we want to explore alternative mechanisms of cytotoxicity that would explain the phenomenon observed with some patients who have achieved significant reduction of circulating light chain post-treatment but still have disease progression. Based on this information, our central hypothesis is that soluble AL light chain species are highly toxic. There are different mechanisms that generate AL toxic species. Specific somatic mutations in AL light chains cause the protein to favor one or more cytotoxic mechanisms.
In aim 1, we will test the hypothesis that protein internalization is required for cytotoxicity using toxicity assays on cardiomyocytes and human arteriole dilation assays.
For aim 2, we will test the hypothesis that low thermodynamic stability of some AL fibrils could cause disaggregation that may play a role in cytotoxicity. In addition, we will identify alternative mechanisms of cytotoxicity by testing the hypothesis that some AL proteins are able to cross seed with non-pathogenic light chains present in the normal repertoire of immunoglobulin molecules.
For aim 3, we will test the hypothesis that AL fibril core composition is mostly driven by the predominant dimer structure populated by each AL protein and will correlate to protein cytotoxicity. Our results will have a collective impact because they will provide new knowledge into the mechanism of AL by identifying common cytotoxic species and AL fibril core regions. This knowledge will be essential for the development of rational drug design to ultimately eliminate the devastating consequences of this disease.

Public Health Relevance

Light chain Amyloidosis (AL) is a rare, incurable, and devastating disease that affects vital organs. In this application, Mechanisms of Aggregation in Light Chain Amyloidosis, we propose to study the toxic species that damage these organs to better design future therapeutic approaches for AL.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM071514-08
Application #
8471712
Study Section
Biophysics of Neural Systems Study Section (BPNS)
Program Officer
Wehrle, Janna P
Project Start
2006-05-01
Project End
2015-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
8
Fiscal Year
2013
Total Cost
$300,187
Indirect Cost
$109,834
Name
Mayo Clinic, Rochester
Department
Type
DUNS #
006471700
City
Rochester
State
MN
Country
United States
Zip Code
55905
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Truran, Seth; Weissig, Volkmar; Ramirez-Alvarado, Marina et al. (2014) Nanoliposomes protect against AL amyloid light chain protein-induced endothelial injury. J Liposome Res 24:69-73
Poshusta, Tanya L; Katoh, Nagaaki; Gertz, Morie A et al. (2013) Thermal stability threshold for amyloid formation in light chain amyloidosis. Int J Mol Sci 14:22604-17
Blancas-Mejia, Luis M; Ramirez-Alvarado, Marina (2013) Systemic amyloidoses. Annu Rev Biochem 82:745-74
Levinson, Rebecca T; Olatoye, Oludare O; Randles, Edward G et al. (2013) Role of mutations in the cellular internalization of amyloidogenic light chains into cardiomyocytes. Sci Rep 3:1278
Martin, Douglas J; Ramirez-Alvarado, Marina (2011) Glycosaminoglycans promote fibril formation by amyloidogenic immunoglobulin light chains through a transient interaction. Biophys Chem 158:81-9
Chadli, Ahmed; Felts, Sara J; Wang, Qin et al. (2010) Celastrol inhibits Hsp90 chaperoning of steroid receptors by inducing fibrillization of the Co-chaperone p23. J Biol Chem 285:4224-31
Peterson, Francis C; Baden, Elizabeth M; Owen, Barbara A L et al. (2010) A single mutation promotes amyloidogenicity through a highly promiscuous dimer interface. Structure 18:563-70
Martin, Douglas J; Ramirez-Alvarado, Marina (2010) Comparison of amyloid fibril formation by two closely related immunoglobulin light chain variable domains. Amyloid 17:129-36
Randles, Edward G; Thompson, James R; Martin, Douglas J et al. (2009) Structural alterations within native amyloidogenic immunoglobulin light chains. J Mol Biol 389:199-210

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