Lung Cancer Early Detection and Immunotherapy Response Prediction and Monitoring with an Exo-PROS Liquid Biopsy Assay

项目摘要 肺癌占全世界癌症死亡人数的25%。早期发现和有效治疗是 对降低死亡率至关重要。低剂量计算机断层扫描（CT）是推荐的筛查试验， 然而，肺癌的高假阳性率导致不必要的活检和重复放射治疗 exposure.免疫检查点抑制剂（ICI）已经彻底改变了肺癌的治疗，然而，它们 仅对少数患者有效。通过组织活检预测和监测对ICI的反应， 影像学检查受到侵入性操作、缺乏有效的肿瘤生物标志物、假性进展和 延迟响应因此，迫切需要无创、准确、经济实惠的检测方法 肺癌早期，并有效地预测和监测对ICI的反应，帮助改善患者的结果。 液体活检检测体液（如血液）中的肿瘤衍生生物标志物，补充成像和风险 因子数据，允许连续监测癌症发展，因此代表了一种新的非侵入性的方法。 以满足这些未满足的需求。外泌体是由细胞分泌的纳米尺寸的细胞外囊泡。 它们积极参与癌症发展的每一步，并已成为有希望的癌症 生物标志物。我们假设肿瘤来源的外泌体（TEXs）对肺肿瘤的发生具有高度的敏感性和特异性。 癌症的早期检测和治疗反应的预测和监测，以及TEX标志物的组合 提供比单独的单一标志物更好的诊断和预测性能。 开发基于外泌体的癌症生物标志物的主要挑战是将TEX从肿瘤细胞中分离。 由正常细胞释放的外泌体，后者可以使测试不太敏感或不能 检测TEX生物标志物。为了克服这一挑战，我们开发了一种外泌体蛋白RNA一站式 （Exo-PROS）生物传感器。Exo-PROS检测首先使用癌细胞-癌细胞技术从非TEX中选择性捕获TEX。 过表达的标记，然后，第一次，提供一站式和原位定量的三种类型的 TEX生物标志物包括蛋白质、微小RNA和碳水化合物抗原。在我们的试点研究中，我们证明了 结合TEX生物标志物（EGFR，miR-21，LG3BP，miR-210，TF-Ag-α）区分肺癌和 正常对照的敏感性为1.00，特异性为1.00。我们还发现TEX PD-L1、miR-21和 TF-Ag-α成功预测了肺癌患者对抗PD-1治疗的反应。本课题 将（1）进一步开发Exo-PROS检测试剂盒并表征其分析性能，包括灵敏度， 特异性、线性范围和重复性;（2）确定Exo-PROS检测在肺组织中的诊断价值 癌症早期检测，并证明Exo-PROS检测补充了低剂量CT，并提高了 诊断准确性;（3）评价Exo-PROS检测在预测和监测抗PD-1应答中的作用 Exo-PROS检测是一种有效的检测方法，可补充组织活检和成像 临床决策和患者护理的模式。

PROJECT SUMMARY Lung cancer causes about 25% of all cancer deaths worldwide. Early detection and effective treatment are critical in reducing the mortality. Low-dose computed tomography (CT) is the recommended screening test for lung cancer, however, its high false-positive rate leads to unnecessary biopsy and repeated radiation exposure. Immune checkpoint inhibitors (ICIs) have revolutionized treatment for lung cancer, however, they are only effective for a minority of patients. Predicting and monitoring responses to ICIs by tissue biopsy and imaging are limited by invasive procedure, the lack of effective tumor biomarkers, pseudo progression, and delayed response. Therefore, non-invasive, accurate and affordable methods are urgently needed to detect lung cancer early and to effectively predict and monitor response to ICIs, aiding in improving patient outcomes. Liquid biopsy detects tumor-derived biomarkers in body fluids such as blood, complements imaging and risk factor data, allows sequential monitoring of cancer development, and thus represents a new and non-invasive strategy to address these unmet needs. Exosomes are nano-sized extracellular vesicles secreted by cells. They are actively involved in every step of cancer development and have emerged as promising cancer biomarkers. We hypothesize that tumor-derived exosomes (TEXs) are highly sensitive and specific for lung cancer early detection and treatment response prediction and monitoring, and the combination of TEX markers provides superior diagnostic and predictive performances than single markers alone. A major challenge in developing exosomes-based cancer biomarkers is the separation of TEXs from exosomes released by normal cells, the latter of which can render the test less sensitive or incapable of detecting TEX biomarkers. To overcome this challenge, we have developed an exosome protein RNA one stop (Exo-PROS) biosensor. The Exo-PROS assay first selectively captures TEXs from non-TEXs using cancer- overexpressed markers, and then, for the first time, provides one-stop and in-situ quantitation of three types of TEX biomarkers including proteins, microRNAs and carbohydrate antigens. In our pilot study, we demonstrated that combined TEX biomarkers (EGFR, miR-21, LG3BP, miR-210, TF-Ag-α) distinguished lung cancer from normal controls with sensitivity of 1.00 and specificity of 1.00. We also showed that TEX PD-L1, miR-21 and TF-Ag-α successfully predicted the response to anti-PD-1 therapy in lung cancer patients. In this project, we will (1) further develop the Exo-PROS assay and characterize its analytical performances including sensitivity, specificity, linear range and repeatability; (2) determine the diagnostic values of Exo-PROS assay in lung cancer early detection and demonstrate that Exo-PROS assay complements low dose CT and improves the diagnostic accuracy; (3) evaluate Exo-PROS assay in predicting and monitoring response to anti-PD-1 therapy, and demonstrate that Exo-PROS assay is an effective test to complement tissue biopsy and imaging modalities in clinical decision making and patient care.