Hepcidin Lateral Flow Device for Prediction of Acute Kidney Injury

描述（由申请人提供）：急性肾损伤（阿基）是心肺转流（CPB）辅助手术的常见严重并发症，包括冠状动脉旁路移植术、瓣膜置换术、主动脉瓣狭窄和器官移植术。老年患者、糖尿病患者和慢性肾病（CKD）患者术后发生阿基的风险非常高。每年有100万例CPB手术导致数千例涉及肾脏替代治疗（RRT）的AKI相关死亡和残疾，累计花费数十亿美元。确认阿基功能丧失的现有肾脏标志物（例如血清肌酸酐）是诊断阿基的晚期标志物（1-3天）。很少有生物标志物（例如NGAL、KIM-1）可用于早期预测阿基。预测CPB后阿基严重程度的早期生物标志物和快速检测将使患者分层、早期治疗干预和阿基药物开发成为可能。最近，我们已经证明铁调素是一种在肝脏中产生并在尿液中排泄的小肽（2.78kD），可能是CPB辅助手术后阿基的良好生物标志物。在与我们的合作者在澳大利亚（NCT 00910221; Prowle et al. 2012）和德国（NCT 00672334; Haase-Felietz et al. 2011）进行的两项100例患者临床试验中，在基线、基线后6小时和24小时采集血浆和尿液样本。与阿基的既定生物标志物血浆肌酐（pCr）和中性粒细胞明胶酶相关脂质运载蛋白（NGAL）相比，在这两项研究中，当通过步枪标准（基于pCr）评分时，尿铁调素（uHep; ng/ml）和尿肌酐校正的uHep（uHep/uCr; ng/mg）与阿基的严重程度呈负相关。无AKI的患者在6小时时具有高度升高的uHep和uHep/uCr水平，该水平在24小时内升高。在德国研究中，与阿基患者相比，6小时时uHep和uHep/uCr增加3- 7倍（P = 0.004，P = 0.002），预测无AKI恢复的AUC-ROC分别为0.80; 0.88，预测无RRT恢复的AUC-ROC分别为0.81; 0.88。这些数据支持uHep和uHep/uCr作为6小时无AKI恢复的有希望的生物标志物，比现有的生物标志物如血浆肌酐、尿排出量和GFR早得多。然而，对于可用于诊断阿基的生物标志物，必须对其进行测量并将数据快速返回给外科医生和ICU工作人员，因为几乎没有针对阿基的既定临床干预措施。我们建议开发一种便携式定量侧流装置（LFD），可以快速测定尿液和血浆hepcidin浓度迅速和定量在ICU。我们最近在正在进行的SBIR研究中发现并表征了两种新的小鼠抗铁调素单克隆抗体（MAb）（PA 08- 114; 2-R44-DK 083843 -02）。对于我们的最后一次提交，我们使用我们最终放弃的金缀合的铁调素示踪剂制备了原型铁调素H1 MAb LFD。我们修订后的第一阶段研究和开发工作的具体目标是：1）。在来自前瞻性临床研究的1800个去识别的时间匹配样本中评估尿和血浆铁调素、NGAL和KIM-1，并进行广泛的统计分析，以及2）开发适合于血浆和尿铁调素的快速定量侧向流装置（LFD），并在来自前瞻性研究的临床样本中测试新装置，以评估铁调素LFD用于CPB后阿基的效用。

DESCRIPTION (provided by applicant): Acute kidney injury (AKI) is a common, severe complication of cardiopulmonary bypass (CPB) assisted surgeries, and include coronary artery bypass grafts, valve replacements, aortic aneurisms, and organ transplants. Elderly patients, diabetics, and chronic kidney disease (CKD) patients, have very high risk of developing AKI postoperatively. One million CPB surgeries yearly result in thousands of AKI-related deaths and disabilities involving renal replacement therapy (RRT), cumulatively costing billions. Existing renal markers confirming loss of function (e.g. serum creatinine) in AKI are very late markers (1-3 days) for diagnosis of AKI. Few biomarkers (e.g. NGAL, KIM-1) are commercially available for early prediction of AKI. Early biomarkers and rapid tests predicting the severity of post-CPB AKI will enable patient stratification, earlier therapeutic interventions, and drug development for AKI Recently, we have shown that hepcidin is a small peptide (2.78kD) produced in the liver and excreted in urine and may be a good biomarker for AKI post CPB-assisted surgery. In two 100 patient clinical trials with our collaborators in Australia (NCT00910221; Prowle et al. 2012) and Germany (NCT00672334; Haase-Felietz et al. 2011), plasma and urine samples were taken at baseline, 6hr and 24hr from baseline. In contrast to established biomarkers of AKI, plasma creatinine (pCr) and neutrophil gelatinase-associated lipocalin (NGAL), in both these studies urinary hepcidin (uHep; ng/ml) and uHep adjusted for urine creatinine (uHep/uCr; ng/mg) was inversely correlated with severity of AKI when scored by RIFLE criteria (based on pCr). AKI-free patients had highly elevated levels of uHep and uHep/uCr at 6hr that increased through 24hr. uHep and uHep/uCr increased 3- to 7-fold compared to patients with AKI (P = 0.004, P = 0.002) at 6hr in the German study with a AUC-ROC 0.80; 0.88, respectively, for prediction of AKI-free recovery and an AUC-ROC 0.81; 0.88, respectively, for prediction of RRT-free recovery. These data support uHep and uHep/uCr as a promising biomarker of AKI-free recovery at 6 hours, much earlier than existing biomarkers such as plasma creatinine, urine output volume, and GFR. However, for a biomarker to be useful for diagnosis of AKI, it must be measured and the data returned to the surgeon and ICU staff quickly as there are few established clinical interventions for AKI. We propose development of a portable, quantitative lateral flow device (LFD) that can rapidly determine urine and plasma hepcidin concentrations rapidly and quantitatively in the ICU. We have recently discovered and characterized two new mouse anti-hepcidin monoclonal antibodies (MAb) in ongoing SBIR research (PA 08- 114; 2-R44-DK083843-02). For our last submission we made a prototype hepcidin H1 MAb LFD using a gold- conjugated hepcidin tracer that we ultimately abandoned. The specific aims for our revised Phase I research and development efforts are, 1). Assess urinary and plasma hepcidin, NGAL, and KIM-1 in 1800 de-identified time-matched samples from a prospective clinical study and perform extensive statistical analysis, and 2) Develop a rapid, quantitative lateral flow device (LFD) suitable for plasma and urine hepcidin and test the new device in the clinical samples from the prospective study to evaluate utility of a hepcidin LFD for post-CPB AKI.