Integrating human functional genomics and transcriptomics in highly phenotyped severe asthmatics

哮喘是最常见的慢性肺部疾病，每年影响全球3.5亿人，造成40万人死亡。多达10%的哮喘患者患有严重的难治性疾病，由于病情恶化、医疗费用和死亡率，构成了大量未得到满足的临床需求。哮喘在病理生物学和临床上是异质性的。识别可治疗的特征，包括2型细胞因子介导的嗜酸性气道炎(T2-High)，使高效的生物标记物导向方法成为可能，根据血液和痰中嗜酸性粒细胞增多和部分呼出的一氧化氮(FeNO)来靶向新的生物药物。然而，其他表型和途径仍有待定义，并被广泛认为是一个紧迫的研究重点。在过去的几十年里，由于可以从呼吸道样本中获得的生物材料数量很少，研究也受到了限制。最近在单细胞测序、多参数流式细胞术和详细的临床和生理表型分析方面的进展使得对高度特征性哮喘亚型中的人类呼吸道组织进行详细的转录和功能分析成为可能。我们开始对60名也接受过WGS的选定参与者的呼吸道样本进行详细的转录分析，使用支气管镜下获得的支气管镜下获得的内膜刷子和胶原酶弥散的支气管活检组织上的10X平台进行配对的单细胞RNAseq和单细胞ATACseq。主要的挑战将是将全基因组测序数据与通过支气管镜从这些患者那里获得的呼吸道样本的转录基因表达数据以及牛津航空研究数据库和NHS电子患者记录的表型数据相结合。在过去10个月里，样本收集工作取得了进展。我承担了验证和优化步骤，并开发了一条样品处理管道。现在有6个人的初步RNAseq数据，我开始分析这些数据。多组文库已经准备好，但尚未测序。我参加了相关的培训，以发展我的生物信息学技能和统计学知识，包括Wellcome Sanger单细胞技术和分析课程和华盛顿大学统计遗传学夏季研究所。我们的目标是在未来12个月内收集至少20个确定的支气管镜检查样本。受试者招募是速率限制因素，但这取决于在严重哮喘诊所内找到合适的参与者，他们有动力同意这项研究。临床医生每年审查528名服务中的新患者，并预计我们的应计费用将继续以目前的速度增长。

Asthma is the most common chronic lung disease, affecting 350 million people worldwide and causing 400,000 deaths annually. Up to 10% of asthmatics have severe, treatment-refractory disease, constituting a significant unmet clinical need due to exacerbations, healthcare costs and mortality. Asthma is pathobiologically and clinically heterogeneous. Identifying 'treatable traits', including type-2 cytokine-mediated ('T2-high') eosinophilic airway inflammation, has enabled highly-effective biomarker-directed approaches, targeting novel biologics according to blood and sputum eosinophilia, and fractional exhaled nitric oxide (FeNO). However other phenotypes and pathways remain to be defined and is widely recognised as an urgent research priority. In previous decades research has also been limited by the very small quantity of biological material which can be obtained from respiratory samples. Recent advances in single cell sequencing, multi-parameter flow cytometry and detailed clinical-and physiological phenotyping now make possible detailed transcriptional and functional analyses of human airway tissue in highly-characterised asthmatic subgroups.We set out to perform detailed transcriptomic analysis of samples from the airways of 60 selected participants who have also undergone WGS, performing paired single-cell RNAseq and single cell ATACseq using the 10X platform on bronchoscopically-obtained endobronchial brushes and scRNAseq on collagenase-dispersed bronchial biopsies. The main challenge will be to integrate the whole genome sequencing data with the transcriptomic gene expression data from airway samples obtained bronchoscopically from these patients and phenotypic data from the Oxford Airways Research Database and the NHS electronic patient record. Sample collection has progressed over the past 10 months. I undertook validation and optimisation steps and developed a sample processing pipeline. Preliminary RNAseq data are now available from 6 individuals which I begun analysing. Mulitiome libraries have been prepared but are yet to be sequenced. I undertook relevant training to develop my bioinformatics skills and knowledge of statistics, including Wellcome Sanger Single Cell Technologies and Analysis course and University of Washington Summer Institute in Statistical Genetics.We are aiming to collect at least 20 definitive bronchoscopy samples within the next 12 months. Subject recruitment is the rate limiting factor, but this depends on finding appropriate participants within the severe asthma clinic who are motivated to consent to this study. The clinicians review 528 new patients per year in the service and expect our accrual to continue at the current rate.