Paraneoplastic neurological disorders (PNS) are inflammatory conditions that can affect both the central and peripheral nervous system and are generally associated with the presence of an underlying tumour. The neurological manifestations are caused by an aberrant immune response that attacks both the tumour and the nervous system. Even though the immune attack to the nervous system is likely to be mainly mediated by cytotoxic T-cells, autoantibodies against intracellular neuronal proteins (onconeural antibodies) are often present in PNS, and represent a crucial diagnostic marker. In addition, specific onconeural antibodies are associated with certain tumours, thus orienting the diagnostic course in these patients. The laboratory detection of onconeural antibodies is complex, and can rely on dot blot using recombinant neuronal proteins, or tissue based assays (TBAs), that exploit animal brain slices as a substrate for antibody binding. Most laboratories rely on commercial kits, and limit their tests to easy-to-perform line blots. However, it has been suggested that the combination of line blots and TBAs provides the highest accuracy, and some studies suggested that the performance of in-house TBAs is superior to commercial TBAs. The first part of this thesis will focus on the optimization of an in-house TBA for the detection of onconeural antibodies. In the second part, we will focus on the validation of the in-house TBA and on the comparison with the currently used commercial line blots and TBAs.
Le sindromi neurologiche paraneoplastiche (PNS) rappresentano delle condizioni infiammatorie che colpiscono sia il sistema nervoso centrale che quello periferico e sono generalmente associate alla presenza di un tumore. Le manifestazioni neurologiche sono causate da un aberrante risposta del sistema immunitario che attacca sia il tumore che il sistema nervoso. Anche se è probabile che l'attacco immunitario al sistema nervoso sia principalmente mediato dai linfociti T citotossici, gli autoanticorpi contro le proteine neuronali intracellulari (anticorpi onconeurali) sono spesso presenti nel SNP e rappresentano un marker diagnostico cruciale. Inoltre, ad alcuni tumori sono associati specifici anticorpi onconeurali, orientando così l’esito diagnostico in questi pazienti. Il rilevamento in laboratorio degli anticorpi onconeurali è complesso e può fare affidamento su dot blot utilizzando proteine neuronali ricombinanti o saggi basati sui tessuti (TBA), che sfruttano fette di cervello animale come substrato per il legame dell'anticorpo. La maggior parte dei laboratori si affida a kit commerciali e limita i propri test all’utilizzo di line-blot di facile esecuzione. Tuttavia, è stato suggerito che la combinazione di line blot e TBA fornisca la massima precisione e alcuni studi hanno suggerito che le prestazioni dei TBA home-made siano superiori ai TBA commerciali. La prima parte di questa tesi si concentrerà sull'ottimizzazione di un TBA home-made per la rilevazione di anticorpi onconneurali. Nella seconda parte ci concentreremo sulla validazione del TBA interno e sul confronto con i line blot e TBA commerciali attualmente utilizzate.
Strategie diagnostiche per la rilevazione di anticorpi anti-neuronali nelle sindromi neurologiche paraneoplastiche: uno studio comparativo
FERRERO, CLELIA
2021/2022
Abstract
Paraneoplastic neurological disorders (PNS) are inflammatory conditions that can affect both the central and peripheral nervous system and are generally associated with the presence of an underlying tumour. The neurological manifestations are caused by an aberrant immune response that attacks both the tumour and the nervous system. Even though the immune attack to the nervous system is likely to be mainly mediated by cytotoxic T-cells, autoantibodies against intracellular neuronal proteins (onconeural antibodies) are often present in PNS, and represent a crucial diagnostic marker. In addition, specific onconeural antibodies are associated with certain tumours, thus orienting the diagnostic course in these patients. The laboratory detection of onconeural antibodies is complex, and can rely on dot blot using recombinant neuronal proteins, or tissue based assays (TBAs), that exploit animal brain slices as a substrate for antibody binding. Most laboratories rely on commercial kits, and limit their tests to easy-to-perform line blots. However, it has been suggested that the combination of line blots and TBAs provides the highest accuracy, and some studies suggested that the performance of in-house TBAs is superior to commercial TBAs. The first part of this thesis will focus on the optimization of an in-house TBA for the detection of onconeural antibodies. In the second part, we will focus on the validation of the in-house TBA and on the comparison with the currently used commercial line blots and TBAs.È consentito all'utente scaricare e condividere i documenti disponibili a testo pieno in UNITESI UNIPV nel rispetto della licenza Creative Commons del tipo CC BY NC ND.
Per maggiori informazioni e per verifiche sull'eventuale disponibilità del file scrivere a: unitesi@unipv.it.
https://hdl.handle.net/20.500.14239/15759