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1.
Int J Biol Macromol ; 270(Pt 2): 132393, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38761898

RESUMO

Light chain amyloidosis is a conformational disease caused by the abnormal proliferation and deposition of antibody light chains as amyloid fibers in organs and tissues. The effect of Cu(II) binding to the model recombinant protein 6aJL2-R24G was previously characterized in our group, and we found an acceleration of the aggregation kinetics of the protein. In this study, in order to confirm the Cu(II) binding sites, histidine variants of 6aJL2-R24G were prepared and the effects of their interaction with Cu(II) were analyzed by circular dichroism, fluorescence spectroscopy, isothermal calorimetry titrations, and molecular dynamics simulations. Confirming our earlier work, we found that His8 and His99 are the highest affinity Cu(II) binding sites, and that Cu(II) binding to both sites is a cooperative event.


Assuntos
Cobre , Histidina , Ligação Proteica , Cobre/metabolismo , Cobre/química , Histidina/química , Histidina/metabolismo , Humanos , Sítios de Ligação , Simulação de Dinâmica Molecular , Cadeias Leves de Imunoglobulina/metabolismo , Cadeias Leves de Imunoglobulina/genética , Cadeias Leves de Imunoglobulina/química , Amiloidose de Cadeia Leve de Imunoglobulina/metabolismo , Amiloidose de Cadeia Leve de Imunoglobulina/genética , Amiloidose/metabolismo , Amiloidose/genética , Cinética
3.
Biochemistry ; 62(5): 1000-1011, 2023 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-36802343

RESUMO

Light chain amyloidosis is the most common form of systemic amyloidosis. This disease is caused by the formation and deposition of amyloid fibers made from immunoglobulin light chains. Environmental conditions such as pH and temperature can affect protein structure and induce the development of these fibers. Several studies have shed light on the native state, stability, dynamics, and final amyloid state of these proteins; however, the initiation process and the fibril formation pathway remain poorly understood structurally and kinetically. To study this, we analyzed the unfolding and aggregation process of the 6aJL2 protein under acidic conditions, with temperature changes, and upon mutation, using biophysical and computational techniques. Our results suggest that the differences in amyloidogenicity displayed by 6aJL2 under these conditions are caused by traversing different aggregation pathways, including unfolded intermediates and the formation of oligomers.


Assuntos
Amiloidose , Cadeias Leves de Imunoglobulina , Humanos , Cadeias Leves de Imunoglobulina/química , Amiloide/química , Amiloidose/metabolismo , Proteínas Amiloidogênicas/genética , Mutação
4.
Rev. Soc. Cardiol. Estado de Säo Paulo ; 33(3): 294-303, 2023. ilus, Tab
Artigo em Português | CONASS, Sec. Est. Saúde SP, SESSP-IDPCPROD, Sec. Est. Saúde SP | ID: biblio-1518214

RESUMO

Amiloidose se refere a um conjunto de doenças causadas pelo depósito de fragmentos anômalos de diferentes proteínas. O coração é um dos principais órgãos afetados, nas formas mais comuns de amiloidose, com os pacientes apresentando mau prognóstico e sobrevida curta, conforme o grau de acometimento. Embora considerada uma afecção rara, vários trabalhos têm demonstrado um aumento expressivo no número de diagnósticos nos últimos anos. Tem contribuído para isso o maior arsenal diagnóstico não invasivo, com novas ferramentas de avaliação mais específicas em exames já consagrados na prática clínica diária, como o ecocardiograma, a cintilografia com marcadores ósseos e a ressonância magnética cardíaca. A disseminação do conhecimento da doença e dos seus sinais de alerta, e o surgimento de diretrizes da doença, associados à chegada de novos tratamentos medicamentosos, têm mudado a história natural desses pacientes. A diminuição da jornada até o diagnóstico e tratamento tem permitido aumento da sobrevida e da qualidade de vida desses pacientes.


Assuntos
Cardiomiopatia Hipertrófica , Cadeias Leves de Imunoglobulina , Insuficiência Cardíaca
5.
Braz J Med Biol Res ; 55: e12284, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36197414

RESUMO

Amyloidoses are a group of disorders in which soluble proteins aggregate and deposit extracellularly in tissues as insoluble fibrils, causing organ dysfunction. Clinical management depends on the subtype of the protein deposited and the affected organs. Systemic amyloidosis may stem from anomalous proteins, such as immunoglobulin light chains or serum amyloid proteins in chronic inflammation or may arise from hereditary disorders. Hereditary amyloidosis consists of a group of rare conditions that do not respond to chemotherapy, hence the identification of the amyloid subtype is essential for diagnosis, prognosis, and treatment. The kidney is the organ most frequently involved in systemic amyloidosis. Renal amyloidosis is characterized by acellular pathologic Congo red-positive deposition of amyloid fibrils in glomeruli, vessels, and/or interstitium. This disease manifests with heavy proteinuria, nephrotic syndrome, and progression to end-stage kidney failure. In some situations, it is not possible to identify the amyloid subtype using immunodetection methods, so the diagnosis remains indeterminate. In cases where hereditary amyloidosis is suspected or cannot be excluded, genetic testing should be considered. Of note, laser microdissection/mass spectrometry is currently the gold standard for accurate diagnosis of amyloidosis, especially in inconclusive cases. This article reviews the clinical manifestations and the current diagnostic landscape of renal amyloidosis.


Assuntos
Amiloidose Familiar , Amiloidose , Amiloide , Proteínas Amiloidogênicas , Amiloidose/diagnóstico , Amiloidose/patologia , Vermelho Congo/uso terapêutico , Humanos , Cadeias Leves de Imunoglobulina/uso terapêutico
8.
Med. lab ; 26(2): 119-139, 2022. ilus, Grafs, tabs
Artigo em Espanhol | LILACS | ID: biblio-1371154

RESUMO

Las amiloidosis sistémicas constituyen un grupo de enfermedades con diversas etiologías, caracterizadas por la síntesis de proteínas con plegado defectuoso, capaces de agregarse y depositarse en el medio extracelular de diferentes órganos y tejidos, alterando su estructura y función. Se conocen más de 14 formas de amiloidosis sistémica, de las cuales la más frecuente es la amiloidosis AL, objeto de esta revisión, en la que las proteínas precursoras son cadenas ligeras de inmunoglobulina inestables, secretadas por un clon de células plasmáticas o, con menor frecuencia, por un linfoma linfoplasmocítico o de células del manto. La amiloidosis AL puede llevar a una amplia gama de manifestaciones clínicas y compromiso de órganos, como el corazón y el riñón. El reconocimiento temprano de la enfermedad y el diagnóstico oportuno son determinantes para mejorar la supervivencia de los pacientes. El tratamiento deberá ser individualizado de acuerdo con la condición de cada paciente, lo que hace necesaria una correcta clasificación de los individuos según su pronóstico. La terapia dirigida a la amiloidosis está enfocada esencialmente en disminuir el compromiso orgánico, y por ende, prolongar la supervivencia con mejoría en los síntomas. En esta revisión se discutirán aspectos importantes de la fisiopatología, epidemiología, manifestaciones clínicas, diagnóstico y tratamiento de la amiloidosis AL


Systemic amyloidosis constitutes a group of diseases with diverse etiologies characterized by the synthesis of proteins with defective folding, capable of aggregating and depositing in the extracellular matrix of different organs and tissues, altering their structure and function. More than 14 forms of systemic amyloidosis are known, of which the most frequent is AL amyloidosis, the subject of this review, in which the precursor proteins are unstable immunoglobulin light chains, secreted by a clone of plasma cells or, to a lesser extent, often due to lymphoplasmacytic or mantle cell lymphoma. AL amyloidosis can lead to a wide range of clinical manifestations and organ involvement, such as the heart and kidney. Early recognition of the disease and timely diagnosis are crucial to improve patient survival. Treatment should be individualized according to the condition of each patient, which requires a properly classification of individuals according to their prognosis. Amyloidosis-targeted therapy is essentially focused on reducing organ involvement, and therefore prolonging survival with improvement in symptoms. In this review, important aspects of the pathophysiology, epidemiology, clinical manifestations, diagnosis, and treatment of AL amyloidosis are discussed


Assuntos
Amiloidose de Cadeia Leve de Imunoglobulina , Proteínas , Cadeias Leves de Imunoglobulina , Dobramento de Proteína , Proteólise , Mutação
9.
J. bras. nefrol ; 43(3): 410-416, July-Sept. 2021. tab, graf
Artigo em Inglês, Português | LILACS | ID: biblio-1340128

RESUMO

Abstract In the past decade, a new class of hemodialysis (HD) membranes (high retention onset class) became available for clinical use. The high cutoff (HCO) and the medium cutoff (MCO) membranes have wider pores and more uniformity in pore size, allowing an increased clearance of uremic toxins. Owing to the mechanism of backfiltration/internal filtration, middle molecules are dragged by the convective forces, and no substitution solution is needed. The HCO dialyzer is applied in septic patients with acute kidney injury requiring continuous kidney replacement therapy. The immune response is modulated thanks to the removal of inflammatory mediators. Another current application for the HCO dialyzer is in hematology, for patients on HD secondary to myeloma-kidney, since free light chains are more efficiently removed with the HCO membrane, reducing their deleterious effect on the renal tubules. In its turn, the MCO dialyzer is used for maintenance HD patients. A myriad of clinical trials published in the last three years consistently demonstrates the ability of this membrane to remove uremic toxins more efficiently than the high-flux membrane, an evolutionary disruption in the HD standard of care. Safety concerns regarding albumin loss as well as blood contamination from pyrogens in the dialysate have been overcome. In this update article, we explore the rise of new dialysis membranes in the light of the scientific evidence that supports their use in clinical practice.


Resumo Na última década, uma nova classe de membranas de hemodiálise (HD) (classe de início de alta retenção) tornou-se disponível para uso clínico. As membranas de ponto de corte alto (HCO) e ponto de corte médio (MCO) têm poros mais largos e maior uniformidade no tamanho dos poros, permitindo uma maior depuração de toxinas urêmicas. Devido ao mecanismo de retrofiltração/filtração interna, as moléculas médias são arrastadas pelas forças convectivas, não sendo necessária uma solução de substituição. O dialisador de HCO é aplicado em pacientes sépticos com lesão renal aguda que requerem terapia renal substitutiva contínua. A resposta imunológica é modulada graças à remoção de mediadores inflamatórios. Outra aplicação atual para o dialisador de HCO é em hematologia, para pacientes em HD secundária ao rim do mieloma, uma vez que as cadeias leves livres são removidas mais eficientemente com a membrana de HCO, reduzindo seu efeito deletério sobre os túbulos renais. Por sua vez, o dialisador de MCO é utilizado para pacientes em HD de manutenção. Uma miríade de ensaios clínicos publicados nos últimos três anos demonstra consistentemente a capacidade desta membrana de remover toxinas urêmicas de forma mais eficiente do que a membrana de alto fluxo, uma ruptura evolutiva no padrão de cuidado em HD. As preocupações de segurança em relação à perda de albumina, bem como a contaminação do sangue por pirogênios no dialisato foram superadas. Neste artigo de atualização, exploramos o surgimento de novas membranas de diálise à luz das evidências científicas que apoiam seu uso na prática clínica.


Assuntos
Humanos , Tecnologia Disruptiva , Soluções para Diálise , Diálise Renal , Cadeias Leves de Imunoglobulina , Membranas Artificiais
10.
J Bras Nefrol ; 43(3): 410-416, 2021.
Artigo em Inglês, Português | MEDLINE | ID: mdl-33836041

RESUMO

In the past decade, a new class of hemodialysis (HD) membranes (high retention onset class) became available for clinical use. The high cutoff (HCO) and the medium cutoff (MCO) membranes have wider pores and more uniformity in pore size, allowing an increased clearance of uremic toxins. Owing to the mechanism of backfiltration/internal filtration, middle molecules are dragged by the convective forces, and no substitution solution is needed. The HCO dialyzer is applied in septic patients with acute kidney injury requiring continuous kidney replacement therapy. The immune response is modulated thanks to the removal of inflammatory mediators. Another current application for the HCO dialyzer is in hematology, for patients on HD secondary to myeloma-kidney, since free light chains are more efficiently removed with the HCO membrane, reducing their deleterious effect on the renal tubules. In its turn, the MCO dialyzer is used for maintenance HD patients. A myriad of clinical trials published in the last three years consistently demonstrates the ability of this membrane to remove uremic toxins more efficiently than the high-flux membrane, an evolutionary disruption in the HD standard of care. Safety concerns regarding albumin loss as well as blood contamination from pyrogens in the dialysate have been overcome. In this update article, we explore the rise of new dialysis membranes in the light of the scientific evidence that supports their use in clinical practice.


Assuntos
Tecnologia Disruptiva , Soluções para Diálise , Humanos , Cadeias Leves de Imunoglobulina , Membranas Artificiais , Diálise Renal
11.
Sci Rep ; 11(1): 1832, 2021 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-33469111

RESUMO

The clinical utility of serum immunoglobulin free light chains (sFLC) in IgG4-related disease (IgG4-RD) is unknown. Herein we evaluated their association with clinical phenotypes, serology and activity in patients with IgG4-RD. Cross-sectional study that included 45 patients with IgG4-RD, and as controls 25 with Sjögren's syndrome (SS) and 15 with sarcoidosis. IgG4-RD patients were classified in clinical phenotypes: pancreato-hepato-biliary, retroperitoneum/aorta, head/neck-limited and Mikulicz/systemic; as well as proliferative vs. fibrotic phenotypes. We assessed the IgG4-RD Responder Index (IgG4-RD RI) at recruitment and measured IgG1, IgG4, κ and λ sFLC serum levels by turbidometry. sFLC levels were similar among IgG4-RD, SS and sarcoidosis groups. Regarding the IgG4-RD patients, the mean age was 49 years, 24 (53.3%) were men and 55.5% had activity. Eight (17.7%) belonged to pancreato-hepato-biliary, 6 (13.3%) to retroperitoneum/aorta, 14 (31.1%) to head/neck-limited, 16 (35.5%) to Mikulicz/systemic phenotypes, whereas 36 (80%) to proliferative and 9 (20%) to fibrotic phenotypes. High κ sFLC, λ sFLC and κ/λ ratio were present in 29 (64.4%), 13 (28.9%) and 13 (28.9%) of IgG4-RD patients, respectively. There were no differences in sFLC among IgG4-RD phenotypes. κ sFLC and κ/λ ratio correlated positively with the number of involved organs and IgG4-RD RI. Patients with renal involvement had higher κ sFLC and λ sFLC. The AUC for κ sFLC and λ sFLC, for renal involvement was 0.78 and 0.72, respectively. Active IgG4-RD had higher levels of κ sFLC and more frequently a high κ/λ ratio. The AUC for κ sFLC and κ/λ ratio for predicting active IgG4-RD was 0.67 and 0.70, respectively. sFLC correlated positively with IgG1 and IgG4 levels. sFLC may be useful as a biomarker of disease activity as well as multiorgan and renal involvement. In particular, a high κ/λ ratio may identify patients with active disease.


Assuntos
Doença Relacionada a Imunoglobulina G4/sangue , Cadeias Leves de Imunoglobulina/sangue , Adulto , Biomarcadores/sangue , Feminino , Humanos , Imunoglobulina G/sangue , Doença Relacionada a Imunoglobulina G4/tratamento farmacológico , Doença Relacionada a Imunoglobulina G4/imunologia , Imunossupressores/uso terapêutico , Masculino , Pessoa de Meia-Idade , Fenótipo
13.
J Appl Lab Med ; 5(1): 29-40, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-32445341

RESUMO

BACKGROUND: The treatment of multiple myeloma (MM) has been revolutionized by the introduction of therapeutic monoclonal antibodies (tmAbs). Daratumumab, a human IgG1/κ tmAb against CD38 on plasma cells, has improved overall survival in refractory MM and was recently approved as a frontline therapy for MM. Work on tmAb interference with serum protein electrophoresis (SPE) during MM monitoring has failed to provide information for laboratories on incidence of interference and effective methods of managing the interference at a practicable level. We aimed to evaluate daratumumab and elotuzumab interference in a large academic hospital setting and implement immediate solutions. METHODS: We identified and chart reviewed all cases of possible daratumumab interference by electrophoretic pattern (120 of 1317 total cases over 3 months). We retrospectively reviewed SPE cases in our laboratory to assess clinical implications of tmAb interference before the laboratory was aware of tmAb treatment. We supplemented samples with daratumumab and elotuzumab to determine the limits of detection and run free light chain analysis. RESULTS: Approximately 9% (120 of 1317) of tested cases have an SPE and/or immunofixation electrophoresis (IFE) pattern consistent with daratumumab, but only approximately 47% (56) of these cases were associated with daratumumab therapy. Presence of daratumumab led to physician misinterpretation of SPE/IFE results. Limits of daratumumab detection varied with total serum gammaglobulin concentrations, but serum free light chain analysis was unaffected. CONCLUSIONS: Clinical laboratories currently rely on interference identification by electrophoretic pattern, which may be insufficient and is inefficient. Critical tools in preventing misinterpretation efficiently include physician education, pharmacy notifications, separate order codes, and interpretive comments.


Assuntos
Anticorpos Monoclonais Humanizados , Anticorpos Monoclonais , Erros de Diagnóstico/prevenção & controle , Cadeias Leves de Imunoglobulina/análise , Mieloma Múltiplo , Anticorpos Monoclonais/análise , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais Humanizados/análise , Anticorpos Monoclonais Humanizados/farmacocinética , Anticorpos Monoclonais Humanizados/uso terapêutico , Eletroforese das Proteínas Sanguíneas/métodos , Humanos , Imunoeletroforese/métodos , Fatores Imunológicos/análise , Fatores Imunológicos/farmacocinética , Fatores Imunológicos/uso terapêutico , Limite de Detecção , Mieloma Múltiplo/sangue , Mieloma Múltiplo/diagnóstico , Mieloma Múltiplo/tratamento farmacológico , Reprodutibilidade dos Testes
14.
Protein Expr Purif ; 170: 105596, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32036001

RESUMO

Antibodies that block interaction of immune checkpoint receptors with its ligands have revolutionized the treatment of several cancers. Despite the success of this approach, the high cost has been restricted the use of this class of drugs. In this context, the development of biosimilar can be an important strategy for reducing prices and expanding access after patent has been dropped. Here, we evaluated the use of HEK293 cells for transient expression of an immune checkpoint-blocking antibody as a first step for biosimilar development. Antibody light and heavy chain genes were cloned into pCI-neo vector and transiently expressed in HEK293 cells. The culture supernatant was then subjected to protein A affinity chromatography, which allowed to obtain the antibody with high homogeneity. For physicochemical comparability, biosimilar antibody and reference drug were analyzed by SDS-PAGE, isoelectric focusing, circular dichroism and fluorescence spectroscopy. The results indicated that the both antibodies have a high degree of structural similarity. Lastly, the biosimilar antibody binding capacity to target receptor was shown to be similar to reference product in ELISA and flow cytometry assays. These data demonstrate that the HEK293 system can be used as an important tool for candidate selection and early development of biosimilar antibodies.


Assuntos
Anticorpos Monoclonais/farmacologia , Medicamentos Biossimilares/farmacologia , Inibidores de Checkpoint Imunológico/farmacologia , Proteínas de Checkpoint Imunológico/genética , Cadeias Pesadas de Imunoglobulinas/farmacologia , Cadeias Leves de Imunoglobulina/farmacologia , Anticorpos Monoclonais/biossíntese , Afinidade de Anticorpos , Especificidade de Anticorpos , Medicamentos Biossimilares/metabolismo , Cromatografia de Afinidade , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Células HEK293 , Humanos , Inibidores de Checkpoint Imunológico/imunologia , Proteínas de Checkpoint Imunológico/imunologia , Cadeias Pesadas de Imunoglobulinas/biossíntese , Cadeias Leves de Imunoglobulina/biossíntese , Focalização Isoelétrica
15.
Transgenic Res ; 29(2): 171-186, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31919795

RESUMO

The expression of recombinant proteins in plants is a valuable alternative to bioreactors using mammalian cell systems. Ease of scaling, and their inability to host human pathogens, enhance the use of plants to generate complex therapeutic products such as monoclonal antibodies. However, stably transformed plants expressing antibodies normally have a poor accumulation of these proteins that probably arise from the negative positional effects of their flanking chromatin. The induction of boundaries between the transgenes and the surrounding DNA using matrix attachment regions (MAR) and insulator elements may minimize these effects. With the PHB-01 antibody as a model, we demonstrated that the insertion of DNA elements, the TM2 (MAR) and M4 insulator, flanking the transcriptional cassettes that encode the light and heavy chains of the PHB-01 antibody, increased the protein accumulation that remained stable in the first plant progeny. The M4 insulator had a stronger effect than the TM2, with over a twofold increase compared to the standard construction. This effect was probably associated with an enhancer-promoter interference. Moreover, transgenic plants harboring two transcriptional units encoding for the PHB-01 heavy chain combined with both TM2 and M4 elements enhanced the accumulation of the antibody. In summary, the M4 combined with a double transcriptional unit of the heavy chain may be a suitable strategy for potentiating PHB-01 production in tobacco plants.


Assuntos
Anticorpos/metabolismo , Cadeias Pesadas de Imunoglobulinas/metabolismo , Elementos Isolantes , Regiões de Interação com a Matriz/genética , Nicotiana/genética , Proteínas Recombinantes/metabolismo , Transgenes/genética , Anticorpos/genética , Regulação da Expressão Gênica de Plantas , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Leves de Imunoglobulina/genética , Cadeias Leves de Imunoglobulina/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Proibitinas , Regiões Promotoras Genéticas , Proteínas Recombinantes/genética , Nicotiana/crescimento & desenvolvimento
16.
Int J Mol Sci ; 20(17)2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31438515

RESUMO

Light-chain amyloidosis (AL) is the most common systemic amyloidosis and is caused by the deposition of mainly insoluble immunoglobulin light chain amyloid fibrils in multiple organs, causing organ failure and eventually death. The germ-line λ6a has been implicated in AL, where a single point mutant at amino acid 24 (6aJL2-R24G) has been observed in around 25% of patient samples. Structural analysis has shown only subtle differences between both proteins; nevertheless, 6aJL2-R24G is more prone to form amyloid fibrils. To improve our understanding of the role of protein flexibility in amyloid fibril formation, we have used a combination of solution nuclear magnetic resonance spectroscopy and molecular dynamics simulations to complement the structural insight with dynamic knowledge. Fast timescale dynamics (ps-ns) were equivalent for both proteins, but suggested exchange events for some residues. Even though most of the intermediate dynamics (µs-ms) occurred at a similar region for both proteins, the specific characteristics are very different. A minor population detected in the dispersion experiments could be associated with the formation of an off-pathway intermediate that protects from fiber formation more efficiently in the germ-line protein. Moreover, we found that the hydrogen bond patterns for both proteins are similar, but the lifetime for the mutant is significantly reduced; as a consequence, there is a decrease in the stability of the tertiary structure that extends throughout the protein and leads to an increase in the propensity to form amyloid fibers.


Assuntos
Amiloidose/metabolismo , Humanos , Cadeias Leves de Imunoglobulina/química , Cadeias Leves de Imunoglobulina/metabolismo , Espectroscopia de Ressonância Magnética , Simulação de Dinâmica Molecular , Dobramento de Proteína , Estrutura Secundária de Proteína
17.
Sci Rep ; 9(1): 3123, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30816248

RESUMO

Immunoglobulin light chain-derived (AL) amyloidosis is a debilitating disease without known cure. Almost nothing is known about the structural factors driving the amyloidogenesis of the light chains. This study aimed to identify the fibrillogenic hotspots of the model protein 6aJL2 and in pursuing this goal, two complementary approaches were applied. One of them was based on several web-based computational tools optimized to predict fibrillogenic/aggregation-prone sequences based on different structural and biophysical properties of the polypeptide chain. Then, the predictions were confirmed with an ad-hoc synthetic peptide library. In the second approach, 6aJL2 protein was proteolyzed with trypsin, and the products incubated in aggregation-promoting conditions. Then, the aggregation-prone fragments were identified by combining standard proteomic methods, and the results validated with a set of synthetic peptides with the sequence of the tryptic fragments. Both strategies coincided to identify a fibrillogenic hotspot located at the CDR1 and ß-strand C of the protein, which was confirmed by scanning proline mutagenesis analysis. However, only the proteolysis-based strategy revealed additional fibrillogenic hotspots in two other regions of the protein. It was shown that a fibrillogenic hotspot associated to the CDR1 is also encoded by several κ and λ germline variable domain gene segments. Some parts of this study have been included in the chapter "The Structural Determinants of the Immunoglobulin Light Chain Amyloid Aggregation", published in Physical Biology of Proteins and Peptides, Springer 2015 (ISBN 978-3-319-21687-4).


Assuntos
Amiloide/metabolismo , Regiões Determinantes de Complementaridade , Cadeias Leves de Imunoglobulina/metabolismo , Amiloidose de Cadeia Leve de Imunoglobulina/metabolismo , Agregação Patológica de Proteínas/metabolismo , Sequência de Aminoácidos , Amiloide/química , Humanos , Cadeias Leves de Imunoglobulina/química , Modelos Moleculares , Conformação Proteica em Folha beta , Multimerização Proteica
18.
Int. j. cardiovasc. sci. (Impr.) ; 32(2): 177-189, mar.-abr. 2019. ilus
Artigo em Inglês | LILACS | ID: biblio-988239

RESUMO

Amyloidosis is a disease caused by extracellular deposition of insoluble protein fibrils, that results in changes in tissue architecture and consequently modification of the organ structure. Cardiac involvement is common in amyloidosis. Two major types of systemic amyloidosis affect the myocardium ­ immunoglobulin light chain and transthyretin amyloidosis ­ each leading to different prognosis. Early detection and diagnosis of cardiac amyloidosis are the main objectives in the assessment of the disease. New techniques of magnetic resonance imaging have minimized the need for biopsies for the diagnosis. Late gadolinium enhancement technique, and more recently T1 mapping, have allowed a simplified evaluation of amyloid deposits and extracellular volume. The aim of this review was to describe basic concepts and updates of the use of magnetic resonance imaging for the diagnosis amyloidosis and evaluation of its severity


Assuntos
Humanos , Masculino , Feminino , Imageamento por Ressonância Magnética/métodos , Amiloidose/diagnóstico , Amiloidose/terapia , Prognóstico , Diagnóstico por Imagem/métodos , Ecocardiografia/métodos , Biomarcadores , Cadeias Leves de Imunoglobulina , Meios de Contraste , Placa Amiloide/diagnóstico por imagem , Eletrocardiografia/métodos , Gadolínio , Ventrículos do Coração , Miocardite/patologia
19.
Clin Lymphoma Myeloma Leuk ; 18(12): e515-e519, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30201257

RESUMO

BACKGROUND: Multiple myeloma (MM) is characterized by the secretion of monoclonal protein by malignant plasma cells in the vast majority of cases. We identified and analyzed patterns of disease relapse and progression associated with disappearance of the paraprotein ("nonsecretory [NS] escape"), or conversion from production of intact Ig molecule to its associated light chain ("LC escape"). PATIENTS AND METHODS: We retrospectively reviewed medical records and a database of 791 consecutive patients with symptomatic MM. RESULTS: Twenty-eight (3.5%) patients had disease evolution associated with either NS (n = 13) or LC (n = 15) escape. The event occurred at a median of 37 months (range, 3-156 months) after the diagnosis of MM, and after a median of 3 chemotherapy regimens (range, 1-8 regimens). Presence of extramedullary disease at progression was detected in 8 (29%) patients. Sensitivity to chemotherapy before and after escape was present in 21 (75%) and 14 (50%) patients, respectively. After a median follow-up of 55 months, 19 (68%) patients died, and progressive MM was the cause of death in 18 patients. The median overall survival after escape was 20 months (95% confidence interval, 9-25 months), and no significant difference was found between the NS and LC groups (P = .44). The median overall survival after diagnosis of MM was worse in patients with NS/LC escape than in those without escape (52 vs. 94 months; P = .018). CONCLUSIONS: Our study describes the largest series of NS and LC escape in MM to date. The development of this phenomenon is associated with more aggressive clinical features, frequent resistance to chemotherapy, and worse clinical outcome.


Assuntos
Cadeias Leves de Imunoglobulina/sangue , Mieloma Múltiplo/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Imunoglobulina A/sangue , Imunoglobulina G/sangue , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/diagnóstico , Mieloma Múltiplo/mortalidade , Mieloma Múltiplo/terapia , Proteínas do Mieloma , Plasmócitos/metabolismo , Estudos Retrospectivos
20.
Rev Med Chil ; 146(1): 64-67, 2018 Jan.
Artigo em Espanhol | MEDLINE | ID: mdl-29806679

RESUMO

BACKGROUND: International guidelines suggest a screening panel for monoclonal gammopathies that contains serum protein electrophoresis (SPE), free light chain (FLC) measurements and immunofixation. This combination provides the possibility of a timely accurate diagnosis. AIM: To evaluate the sensibility of a simple screening panel (SPE + FLC). MATERIAL AND METHODS: We analyzed 191 consecutive serum samples of patients with a suspected monoclonal gammopathy (MG). RESULTS: Seventy five patients were diagnosed with MG. The sensitivity and specificity of the combination of SPE + FLC for the diagnosis of monoclonal gammopathy were 95% (95% confidence intervals 89-99) and 99% (95% confidence intervals 96-100), respectively. CONCLUSIONS: We were able to validate the international recommendations on the diagnostic accuracy of this simple combination of two tests in serum for monoclonal gammopathy.


Assuntos
Eletroforese das Proteínas Sanguíneas/métodos , Cadeias Leves de Imunoglobulina/sangue , Paraproteinemias/diagnóstico , Biomarcadores/sangue , Humanos , Estudos Prospectivos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
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