Biomolecular Markers for Safe Minimization of Immunosuppression

我们寻求更多资金用于应对SARS-CoV-2/新冠肺炎爆发的研究，范围包括 我们正在进行的资助R37AI051652“安全最小化免疫抑制的生物分子标记”。从… 2020年3月13日至2020年4月20日，我们收治了39例SARS-CoV-2阳性的肾移植患者和 有新冠肺炎症状。在这些患者中，20例(51%)发展为急性肾损伤(AKI)。重要的是，贪污 截至2020年5月15日，因AKI导致的功能障碍只有9名患者恢复，11名患者没有恢复。无 接受了诊断性同种异体移植活检，因为活检相关的并发症，如出血 在这个队列中可能会更加严重，并限制医护人员接触SARS的潜在风险- 在侵入性活检过程中发现CoV-2病毒。在没有诊断性活检的情况下，没有一例接受抗排斥治疗。 心理治疗。移植物功能障碍是可逆的还是不可逆的，在移植时是无法预测的。 功能障碍诊断。抗同种异体移植物免疫抑制反应的动态变化 治疗不能通过可用的临床分析来捕捉。为了应对这些现有的挑战，我们 提出以下建议：具体目标1.进行尿液细胞的RNA测序 在移植物功能障碍时确定的尿细胞转录组是否可以预测预后 移植物功能障碍。将在移植物功能障碍诊断时收集尿液，并从 尿路细胞。30例可逆性移植物功能障碍患者的RNA；30例不可逆性移植物功能障碍患者的RNA 新冠肺炎以来3个月内无移植物功能障碍的30例患者的核糖核酸 诊断将通过RNA测序和生物信息学进行。目标是确定尿液是否 在移植物功能障碍时确定的细胞转录组图谱是移植物功能障碍和 区分可逆性移植物功能障碍和不可逆性移植物功能障碍。特定的 目的2.测定新冠肺炎肾序贯尿标本中尿细胞3基因特征记分 同种异体移植受者。每两周收集一次尿液，为期三个月。 新冠肺炎诊断。我们将从30名新冠肺炎肾移植受者中检索210份序贯样本 (每个患者的基线和6个连续样本)发生可逆性移植物功能障碍的患者；210 来自30例发生不可逆性移植功能障碍的新冠肺炎肾移植受者的序贯样本； 以及30名新冠肺炎肾移植受者，他们在此后的3个月内没有出现移植肾功能障碍 新冠肺炎诊断。从尿液细胞中提取的RNA将被逆转录成cDNA和绝对拷贝 CD3E mRNA、CXCL10 mRNA和18S rRNA的数量将使用定制的PCR方法和 尿液细胞3基因签名分数将使用验证的回归方程来计算。我们的目标是 为了确定序贯样本中的尿细胞3-基因评分是否预测了那些发生 不可逆性移植物功能障碍来自发生可逆性移植物功能障碍的患者。

We seek additional funds for research responsive to the SARS-CoV-2/COVID-19 outbreak that is in scope of our ongoing grant R37AI051652 “Biomolecular Markers for Safe Minimization of Immuno-suppression.” From March 13, 2020 to April 20, 2020, we hospitalized 39 kidney allograft recipients positive for SARS-CoV-2 and with Covid-19 symptoms. Among these patients, 20 (51%) developed acute kidney injury (AKI). Importantly, graft dysfunction due to AKI recovered in 9 patients only and did not recover in 11 patients as of May 15, 2020. None underwent a diagnostic allograft biopsy because of biopsy-associated complications such as bleeding are potentially much more serious in this cohort and also to limit potential exposure of healthcare workers to SARS- CoV-2 during the invasive biopsy procedure. In the absence of a diagnostic biopsy, none received anti-rejection therapy. Whether the graft dysfunction was reversible or nonreversible could not be predicted at the time of graft dysfunction diagnosis. The dynamics of anti-allograft response from the reductions in their immunosuppressive therapy could not be captured with the available clinical analytes. To address these existing challenges, we propose the following: Specific Aim 1. To perform RNA sequencing of urinary cells and investigate whether the urinary cell transcriptome, ascertained at the time of graft dysfunction, is prognostic of allograft dysfunction. Urine will be collected at the time of graft dysfunction diagnosis and RNA isolated from urinary cells. RNA from 30 patients with reversible graft dysfunction; RNA from 30 patients with nonreversible graft dysfunction; and RNA from 30 patients with no graft dysfunction during the 3 months since Covid-19 diagnosis will be RNA sequenced and bioinformatics performed. The goal is to determine whether the urinary cell transcriptome profile, ascertained at the time of graft dysfunction, is prognostic of graft dysfunction and distinguishes those with reversible graft dysfunction from those with nonreversible graft dysfunction. Specific Aim 2. To measure urinary cell 3-gene signature score in sequential urine samples from Covid-19 kidney allograft recipients. Urine will be collected at baseline and sequentially every two weeks for 3 months since Covid-19 diagnosis. We will retrieve 210 sequential samples from 30 Covid-19 kidney allograft recipients (Baseline and 6 sequential samples from each patient) who developed reversible graft dysfunction; 210 sequential samples from 30 Covid-19 kidney allograft recipients who developed nonreversible graft dysfunction; and 30 Covid-19 kidney allograft recipients who did not develop graft dysfunction during the 3-months since Covid-19 diagnosis. RNA isolated from urinary cells will be reverse transcribed to cDNA and absolute copy numbers of CD3E mRNA, CXCL10 mRNA and 18S rRNA will be measured using customized PCR assays and urinary cell 3-gene signature score will be computed using a validated regression equation. The objective is to determine whether the urinary cell 3-gene scores in sequential samples anticipate those who develop nonreversible graft dysfunction rom those who develop reversible graft dysfunction.