Immunoglobulin (Ig) proteins, also known as antibodies, are important molecules with great variability in their amino acid sequences. Aberrantly overproduced monoclonal Ig light chain (LC) proteins may aggregate into a β-sheet featured structure, and deposit in the extracellular space; this pathologic process, called primary amyloidosis or Ig LC amyloidosis (AL) causes problems to multiple organs during the course of the disease. Post-translational modifications (PTMs), which remain to be explored, are likely an important factor affecting the formation of AL fibrils. In addition, therapeutic monoclonal antibodies (mAbs) are widely employed because of their high specificity and low side effects. Using plants as the expression platform is commercially attractive although this approach has been hampered by low protein expression yield and undesired glycosylation patterns. The investigations detailed in this dissertation focus on the analyses of Ig proteins derived from several human and plant sources. A method combining 2D SDS-PAGE separation and mass spectrometry (MS) analysis was used for de novo sequencing of Ig in a fat biopsy for which the corresponding DNA was unavailable, and for characterizing the LC proteins found in autopsied kidney, serum and urine samples from patients with AL amyloidosis whose LC-DNA was sequenced. The PTMs of each LC were extensively characterized with different enzymes and various tandem MS techniques including collision-induced dissociation (CID), higher-energy collisional dissociation (HCD) and electron transfer dissociation (ECD). PTMs observed include truncations, mono-/di-chlorination of the tyrosine residues and a nitrile group formed from the primary amine on lysine side chains. All these may play critical roles in the fibrillogenesis and/or the disease pathogenesis. Experimental evidence supports the hypothesis that the proteolytic processing of amyloidogenic LCs occurs after deposition in the organ. Characterization of a plant-derived HSV8 mAb was accomplished using high-performance liquid chromatography separation and gel display followed by various MS methods. Three N-terminal and one C-terminal truncations were found. The N-glycan moiety attached to the heavy chain was also analyzed. The MS method established helps to elucidate important structural information on therapeutic mAbs in complex mixtures, potentially contributing to optimization of plant systems that may produce more stable mAbs.

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Introduction: Waldenström's macroglobulinemia is a hematological neoplasm belonging to the group of monoclonal gammopathies, which includes systemic symptoms and those related to an increase in M paraprotein. Objective: To describe a case of cardiac amyloidosis associated with macroglobulinemia. Clinical case: Male patient who was admitted for asthenia, dysphonia, and who, during his evolution, developed progressive dyspnea, heart failure and pleural effusion. Additionally, echocardiography showed myocardial granular pattern, while pleural biopsy was positive for Congo red staining. Subsequently, he received treatment with bortezomib, dexamethasone and rituximab, with favorable evolution. Conclusions: In this disease, early diagnosis is an important advantage for survival. Therefore, its management is palliative of cardiac manifestations. The present case shows a diagnostic challenge, in which the less frequent etiologies of heart failure must be taken into account. / Revisión por pares

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