自噬相关基因功能突变的筛选鉴定及其在SAPHO综合征中的作用机制研究

SAPHO, synovitis, acne, pustulosis, hyperostosis, and osteitis, syndrome is a rare chronic autoinflammatory disease presenting with dermatological, acne on the face and neck, pustulosis affecting the palms of the hands and soles of the feet and osteoarticular symptoms synovitis resulting in painful swelling, hyperostosis at sites of tendons bone attachments and osteitis affecting the sternocostal, sternoclavicular and sacroiliac joints. This rare condition has prevalence of 1 in 10 000 Caucasians and likely similar incidence in China, affecting 140 000 Chinese poeple. .SAPHO syndrome can affect any age and gender and is usually clinically worse in female patients. The clinical heterogeneity and lack of laboratory diagnostic tests makes the diagnosis challenge. No laboratory diagnostic tests are available. Nonsteroidal anti-inflammatory drugs, corticosteroids and antibiotics are usually prescribed to treat SAPHO syndrome, and sometimes therapy with infliximab, however the effectiveness of these drugs is variable as they do not target the underlying problem. The underlying pathophysiology of SAPHO syndrome is not known and there is a pressing need to identify new targeted treatments patients that will benefit from such targeted therapies based on the molecular profiling of the specific causal pathway. .A genetic predisposition to SAPHO syndrome has been described based on familial clustering and monozygotic twin concordance studies. A number of genes have been suggested to play a role in SAPHO syndrome, such as proline serine threonine phosphatase interacting protein 2, PSTPIP2, however no association between gene variants and disease development have been identified. While genetics is likely to play a significant role in the pathogenesis of SAPHO syndrome, it is evident that the search for the causative genes will need to employ genetic strategies that can overcome the hurdles of clinical heterogeneity and very low prevalence of the disease..The pace of discovery of rare gene variants causing disease, Mendelian and hidden Mendelian or sporadic due to de novo mutations, including autoinflammatory diseases, has accelerated with the use of next generation sequencing technologies. In addition, the ability to prove the causality of these rare variants was greatly aided by the recent discovery of fast DNA editing tools allowing to introduce these human alleles in mouse zygotes and study their effect..Our proposal is for a study aiming to study both Caucasian and Chinese patients with SAPHO syndrome in order to elucidate the genetics and pathophysiology of this rare auto-inflammatory disease. Our main hypothesis is that SAPHO syndrome is caused by a small number of rare and deleterious gene variants, conferring a strong genetic component to the disease. We have already identified several mutations in an autophagy pathway in a cohort of Chinese SAPHO patients. Besides analysing the already-sequenced exomes from an additional European-Australian cohort, we plan to validate the identified variants using various in vitro laboratory techniques to detect functional defects, as well as in vivo modelling by introducing these alleles in mouse models using the CRISPR/Cas9 technology. We will also aim to perform detailed immune phenotyping using PBMCs and specifically neutrophils from patients with mutations in autophagy related genes. We also propose to trial predicted effective therapies based on identified rare genetic variants in the generated personalised mouse models in order to validate newly identified targets for the personalised treatment approach. The scientific questions we aim to answer are: is the incidence of autophagy-related gene mutations high and relevant to the SAPHO syndrome, what are dysregulated the immune cell types, do and how the rare genetic variants alter protein structure and function and what are the most effective therapies for SAPHO syndrome.

SAPHO（滑膜炎、痤疮、脓疱病、骨肥厚和骨炎）综合征是一种罕见自身炎症性骨关节病，严重影响患者皮肤和骨骼系统，其发病机制不清楚，也没有治愈手段。SAPHO家族聚集性提示遗传因素可能参与了发病，罕见疾病可能由罕见突变所致。我们在6名中国SAPHO患者全外显子测序结果中发现数个有可能的致病突变，主要集中在自噬通路。本项目中将在澳洲和欧洲患者中验证这些突变，并采用多种体外实验技术研究突变的功能意义；我们还将采用CRISPR/Cas9技术构建和患者突变具有同源突变的基因工程小鼠，进行突变的体内功能研究；此外，我们将在突变小鼠进行针对受累通路的药物治疗试验。通过本项目，我们将阐明自噬通路突变是否是SAPHO致病突变，是通过何种细胞何种机制参与疾病发生，突变是如何调节蛋白结构和功能，并探索针对相关通路的药物是否能为SAPHO治疗提供新的思路。本项目的实施将为罕见病的机制研究提供创新性的研究策略和方案

SAPHO（滑膜炎、痤疮、脓疱病、骨肥厚和骨炎）综合征是一种罕见自身炎症性骨关节病，严重影响患者皮肤和骨骼系统，其病因和发病机制尚不清楚，也没有治愈手段。SAPHO综合征具有家族聚集性，提示遗传因素可能参与了发病。一般认为，SAPHO综合征的发生和发展是遗传、感染和免疫因素共同作用的结果，感染在具有遗传易感性的个体中触发疾病的发生，继而先天免疫系统引起反复发作的炎症。有研究表明SAPHO综合征存在家族聚集性，且在同卵双胞胎存在较高的共患率，说明SAPHO综合征存在遗传易感性。目前已经发现某些原因不明的自身炎症性疾病其实是由罕见遗传突变导致的，根据临床表型和孟德尔遗传模式可以明确的对其定义和诊断。因此我们通过全外显子组测序技术，研究了29名SAPHO患者中可能的致病罕见突变。发现了一系列具有潜在致病性的罕见突变，这些罕见突变主要集中在自噬通路，LIFR通路，TLR通路，NLRP通路以及OCSTAMP、PSTPIP2等可能在骨生成以及骨关节炎中发挥作用的基因上。提示上述通路和基因在SAPHO疾病发生发展中发挥作用。通过本项目的实施能加深对SAPHO综合征这类罕见疾病发病机制研究提供创新性的研究策略和方案，为SAPHO治疗提供新的思路。