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Abstract

Immunoglobulins and their component light chains are essential parts of the human immune system. Even though, in the context of an AL amyloidosis disease, light chains can be pathogenic. This pathogenicity arises from at least one malignant plasma cell, which infiltrates the bone marrow and produces an excess of free light chains – which misfold and ultimately deposit as fibrils in the extracellular space. Why and how these naturally soluble light chains transit into fibrils is still unknown. One research approach relies on the natural function of the light chains themselves: antigen recognition and underlying sequence diversity. The analysis of this sequence diversity and corresponding mutations may allow making inferences about potential pathogenic mechanisms in AL amyloidosis. Nevertheless, a control group is needed to draw distinct conclusions. Here, light chains from the related plasma cell disorder multiple myeloma, which do not form fibrils, offer a suitable option. This work compared full-length lambda light chain sequences from 82 AL amyloidosis and 52 multiple myeloma patients and revealed several striking aspects – such as, a significant association between IGLV6-57 and AL amyloidosis as well as between IGLV2-23 and multiple myeloma. Overall, the results of this work are well in line with previously published studies and strengthen the assumption that IGLV subfamilies present a difference in their amyloidogenic potential. Intuitively, mutations may alter this inherent amyloidogenic potential. In the context of this work, mutation analyses showed that not the quantity but rather the distinct exchange pattern and location might be crucial. This hypothesis rests upon the findings that the multiple myeloma sequences always displayed a significantly higher mutation count than the respective AL amyloidosis sequences and the detection of different exchange patterns. In more detail, these patterns mostly concerned charged amino acids and even differed between the analyzed IGLV subfamilies. Therefore, individual pathological sequence-based mechanisms for each IGLV subfamily are hypothesized. In addition to the characteristic of light chain fibril formation, AL amyloidosis patients often show a distinct organ tropism. To further analyze this aspect, the AL amyloidosis patients were stratified accordingly beforehand. In the end, this study included 61 patients with a dominant heart and 21 patients with a heart and kidney involvement. When comparing these subgroups, it was possible to detect differences concerning the IGLV subfamily assignment, mutation counts, and again most prominently, charged amino acids. In the context of charged amino acids, the IGLV6-57 assigned sequences stood out. Here, it was possible to detect diverging patterns between the two AL amyloidosis subgroups in several regions. Taken together, these results suggest that not exclusively the light chains themselves, but also the tissue environment might influence their amyloidogenic potential which are mirrored in certain sequence characteristics. Besides the already mentioned high levels of free light chains, AL amyloidosis patients frequently display a loss of the clonal heavy chain. To further investigate these parameters and to find out if they reflect in certain light chain sequence characteristics, the AL amyloidosis and multiple myeloma cohort were accordingly stratified. Addressing the level of free light chains, the most prominent difference concerned the molecular weight of the light chains. IGLV6-57 and IGLV2-14 AL amyloidosis sequences from patients with a high level of free light chains displayed a higher molecular weight than the other subgroup. The second analysis concerning the presence or absence of a clonal heavy chain revealed an increased mutation count in the IGLV and the IGLC segment of light chains with potential heavy chain binding partners. These results led to the hypothesis that mismatched heavy chains and light chains might influence the excess of free light chains. In this context, there might also be a positive selection for light chains that exhibit a higher aggregation tendency and isoelectric point close to the physiological pH of the blood – which would render them insoluble within the patient’s bloodstream. In conclusion, the results of this study lead to several intertwining hypotheses concerning the pathogenic mechanisms in AL amyloidosis: Certain IGLV subfamilies innately present with favorable properties for amyloid formation. Here, individual pathological sequence-based mechanisms, which frequently involve alterations of charged amino acids, can be assumed. Further, not only the light chains themselves but also external factors, like the tissue environment and a potential clonal heavy chain binding partner, might influence their amyloidogenity. In this context, there might also be a positive selection for light chains which exhibit favorable characteristics like an increased aggregation tendency or a favorable isoelectric point. Besides the biological questions concerning the pathogenic mechanisms, there is an urgent need for a screening option for patients with a monoclonal gammopathy of undefined significance on the clinical/medical side. Identifying patients at high risk for disease progression towards AL amyloidosis would possibly prevent end-organ damage due to earlier diagnosis. Based on the results of this work, a risk stratification mainly based on the IGLV subfamily usage was established. Keeping the previously mentioned IGLV associations in mind, it is reasonable that the highest significant relative risk, 11.63, was calculated for IGLV6-57. In contrast, the lowest significant relative risk, 0.04, was detected for IGLV2-23. In summary, this work contributes to resolving the pathogenic mechanisms of AL amyloidosis and provides a starting point for clinical risk assessment based on light chain sequencing.