Neuropathic pain is a frequent feature in peripheral neuropathy in particular when small nerve fibers, which convey thermal and nociceptive sensations, are involved. Excruciating burning pain at feet and hand is the most common feature of small fiber neuropathy (SFN) which represents a good model for studying neuropathic pain. Voltage gated sodium channel (VGSCs) genes mutations have been found in rare familial painful disorders and more recently, variants in the same genes have been identified in idiopathic and diabetic painful neuropathies, thus widening the spectrum of genetic pain disorders. This PhD thesis aimed at investigating the risk for neuropathic pain in a well-phenotyped cohort of SFN and diabetic neuropathy patients in order to provide a clinical and genetic characterization of patients. The first section focused on the deep-phenotyping of patients with suspected SFN or neuropathic pain through the development of a database for systematic data collection, integration and sharing among clinicians and researchers. Collected data have been used to conduct two retrospective studies. The first study aimed at addressing the diagnostic accuracy of skin biopsy over time comparing the different normative values for intraepidermal nerve fiber density (IENFD) adopted from 1999 to 2019. This study, comparing skin biopsy results in 439 patients according to different cut-off values, showed a significant improvement of skin biopsy diagnostic specificity after the introduction of the age-and-sex-adjusted normative reference values in the 2010, reporting a reduction of false positive of more than 50% when compared with the cut-off values previously adopted. The second study investigated the circadian dynamics of neuropathic pain intensity scored using the numeric rating scale (PI-NRS) in a cohort of 253 patients with suspected painful SFN. This study revealed a circadian pattern of pain features, showing an increase of NRS scores towards the evening, suggesting a possible role for the intra-day PI-NRS variations as adjunctive outcome measure in clinical trials for analgesic drug in SFN-related neuropathic pain. The second section of the thesis provided a genetic characterization of SFN patients. A candidate-gene analysis has been conducted, looking for rare and low frequency genetic variants in VGSCs genes expected to have a large effect on clinical phenotype and describing their frequency in phenotypically well-defined cohorts of SFN patients. The analysis conducted on 1,015 patients grouped according to etiology and painful phenotype showed a slightly higher frequency of VGSCs variants in painful compared to painless phenotype (13.5% and 9.7%, respectively) but no significant differences between diabetes and idiopathic SFN patients (11.5%). Looking at the variants distribution in VGSCs genes, idiopathic and painful patients showed a significant higher frequency of SCN9A variants whereas diabetes and painless patients had more variants in SCN10A gene. However, concerns have been raised about the pathogenicity of single rare gain-of-function variants, since most of them were classified as VUS (variants of unknown significance). Based on these findings, we adopted a new research approach to investigate the risk of neuropathic pain in our diabetic cohort of 513 patients. The work hypothesis relied on a polygenic architecture of painful neuropathy in which all variants, whether rare or common, might contribute with a small effect size to compose the clinical phenotype. Therefore, we computed a polygenic risk score (PRS) combining the weight of each variant identified in a panel of 107 pain-related genes in diabetic neuropathy patients. The PRS was able to discriminate with sufficient accuracy painful from painless patients with an AUC of 60.3%. This study represented the first application of PRS for addressing the risk of neuropathic pain in diabetic neuropathy, pioneering the use of this tool in this clinical context.

Il dolore neuropatico è una caratteristica frequente delle neuropatie periferiche, in particolare quando sono coinvolte le piccole fibre nervose che veicolano la sensibilità termo-dolorifica. La neuropatia delle piccole fibre (SFN) tipicamente si presenta con dolore urente distale agli arti e rappresenta un buon modello per lo studio del dolore neuropatico. Mutazioni nei geni dei canali del sodio voltaggio dipendenti (VGSC) sono state descritte in sindromi algiche familiari e nell’insensibilità congenita al dolore. Più recentemente, varianti in questi geni sono state identificate in associazione a condizioni più comuni quali le neuropatie dolorose. Obiettivo di questa tesi è di indagare il rischio di dolore neuropatico in una coorte ben caratterizzata di pazienti affetti da SFN e neuropatia diabetica. Nella prima sezione sono stati indagati gli aspetti clinici mediante una dettagliata caratterizzazione fenotipica dei pazienti con sospetto di SFN e dolore neuropatico, attraverso la progettazione di un database per la raccolta sistematica dei dati, l'integrazione e la condivisione tra medici e ricercatori. Tali dati sono stati utilizzati per condurre due studi retrospettivi. Il primo ha valutato l'accuratezza diagnostica della biopsia cutanea nel tempo, confrontando i diversi valori normativi per la densità di innervazione intraepidermica adottati dal 1999 al 2019. I risultati su 439 pazienti hanno evidenziato un significativo miglioramento della specificità diagnostica della biopsia cutanea dopo l'introduzione dei valori normativi corretti per età e sesso nel 2010, con una riduzione dei falsi positivi di oltre il 50%. Il secondo studio ha indagato le variazioni circadiane dell'intensità del dolore neuropatico in una coorte di 253 pazienti con sospetta SFN dolorosa rivelando un significativo incremento dell’intensità del dolore dal mattino/pomeriggio alla sera. Lo studio ha mostrato un pattern circadiano del dolore neuropatico che potrebbe fornire una misura di outcome aggiuntiva negli studi clinici per il trattamento del dolore neuropatico nella SFN. La seconda sezione include due studi finalizzati ad identificare determinanti genetiche associate a diversi fenotipi clinici in relazione all’eziologia e alla presenza o assenza di dolore neuropatico. Una prima analisi è stata effettuata su geni candidati, ricercando varianti rare e a bassa frequenza nei geni VGSC potenzialmente esercitanti maggiore effetto sul fenotipo clinico. L'analisi condotta su 1.015 pazienti ha mostrato una maggiore frequenza di varianti rare nei geni VGSC in pazienti con dolore rispetto al fenotipo senza dolore (13,5% e 9,7%) ma nessuna differenza significativa dividendo il campione sulla base dell’eziologia diabetica o idiopatica (11,5%). Osservando la distribuzione delle varianti nei geni VGSCs, i pazienti idiopatici e quelli con dolore presentavano una frequenza significativamente più alta di varianti in SCN9A, mentre nei pazienti con diabete e in quelli senza dolore prevalevano varianti nel gene SCN10A. Tuttavia, la maggior parte di queste varianti sono state classificate come VUS (varianti di significato sconosciuto), pertanto esistono dubbi circa il loro reale significato patogenetico. Sulla base dell'ipotesi di un'architettura poligenica della neuropatia dolorosa in cui tutte le varianti, sia rare che comuni, possono contribuire al fenotipo clinico, è stato adottato un nuovo approccio per indagare il rischio di dolore neuropatico nei pazienti con diabete. E’ stato calcolato un punteggio di rischio poligenico (PRS) combinando il peso di ogni variante identificata in un pannello di 107 geni correlati al dolore, in pazienti affetti da neuropatia diabetica con e senza dolore. Il PRS è stato in grado di discriminare con una sufficiente accuratezza pazienti con dolore da quelli senza dolore. Questo studio rappresenta la prima applicazione del PRS nello studio del dolore neuropatico associato a neuropatia diabetica.

Clinical and genetic characterization of neuropathic pain through the model of small fiber neuropathy: implication for diabetic neuropathy

CAZZATO, Daniele
2020

Abstract

Neuropathic pain is a frequent feature in peripheral neuropathy in particular when small nerve fibers, which convey thermal and nociceptive sensations, are involved. Excruciating burning pain at feet and hand is the most common feature of small fiber neuropathy (SFN) which represents a good model for studying neuropathic pain. Voltage gated sodium channel (VGSCs) genes mutations have been found in rare familial painful disorders and more recently, variants in the same genes have been identified in idiopathic and diabetic painful neuropathies, thus widening the spectrum of genetic pain disorders. This PhD thesis aimed at investigating the risk for neuropathic pain in a well-phenotyped cohort of SFN and diabetic neuropathy patients in order to provide a clinical and genetic characterization of patients. The first section focused on the deep-phenotyping of patients with suspected SFN or neuropathic pain through the development of a database for systematic data collection, integration and sharing among clinicians and researchers. Collected data have been used to conduct two retrospective studies. The first study aimed at addressing the diagnostic accuracy of skin biopsy over time comparing the different normative values for intraepidermal nerve fiber density (IENFD) adopted from 1999 to 2019. This study, comparing skin biopsy results in 439 patients according to different cut-off values, showed a significant improvement of skin biopsy diagnostic specificity after the introduction of the age-and-sex-adjusted normative reference values in the 2010, reporting a reduction of false positive of more than 50% when compared with the cut-off values previously adopted. The second study investigated the circadian dynamics of neuropathic pain intensity scored using the numeric rating scale (PI-NRS) in a cohort of 253 patients with suspected painful SFN. This study revealed a circadian pattern of pain features, showing an increase of NRS scores towards the evening, suggesting a possible role for the intra-day PI-NRS variations as adjunctive outcome measure in clinical trials for analgesic drug in SFN-related neuropathic pain. The second section of the thesis provided a genetic characterization of SFN patients. A candidate-gene analysis has been conducted, looking for rare and low frequency genetic variants in VGSCs genes expected to have a large effect on clinical phenotype and describing their frequency in phenotypically well-defined cohorts of SFN patients. The analysis conducted on 1,015 patients grouped according to etiology and painful phenotype showed a slightly higher frequency of VGSCs variants in painful compared to painless phenotype (13.5% and 9.7%, respectively) but no significant differences between diabetes and idiopathic SFN patients (11.5%). Looking at the variants distribution in VGSCs genes, idiopathic and painful patients showed a significant higher frequency of SCN9A variants whereas diabetes and painless patients had more variants in SCN10A gene. However, concerns have been raised about the pathogenicity of single rare gain-of-function variants, since most of them were classified as VUS (variants of unknown significance). Based on these findings, we adopted a new research approach to investigate the risk of neuropathic pain in our diabetic cohort of 513 patients. The work hypothesis relied on a polygenic architecture of painful neuropathy in which all variants, whether rare or common, might contribute with a small effect size to compose the clinical phenotype. Therefore, we computed a polygenic risk score (PRS) combining the weight of each variant identified in a panel of 107 pain-related genes in diabetic neuropathy patients. The PRS was able to discriminate with sufficient accuracy painful from painless patients with an AUC of 60.3%. This study represented the first application of PRS for addressing the risk of neuropathic pain in diabetic neuropathy, pioneering the use of this tool in this clinical context.
LAURIA PINTER, GIUSEPPE
CASETTA, Ilaria
FADIGA, Luciano
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11392/2478814
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