Bionanoconjugates for Detection of Circulating Tumor Cells in Lung Cancer

用于检测肺癌循环肿瘤细胞的生物纳米缀合物

• 批准号: 8203793
• 负责人: Edward A. Hirschowitz
• 金额: $ 7.43万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-18 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8203793
• 关键词: Antibodies; Antigens; Binding; Biological; CTAG1 gene; Cancer Patient; Cell Adhesion Molecules; Cell surface; Cells; Clinical; Clinical Management; Complex; Complex Mixtures; Coupling; Cultured Tumor Cells; DNA; DNA Sequence; Detection; Development; Diagnostic Neoplasm Staging; Disease; Epithelial Cells; Fluorescence; Foundations; Frequencies; Future; Gold; Heating; Heterogeneity; Histocytochemistry; Imagery; Literature; Logic; Lung Neoplasms; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of prostate; Measurement; Measures; Mediating; Messenger RNA; Methodology; Methods; Microscopy; Monoclonal Antibodies; Nanoconjugate; Non-Small-Cell Lung Carcinoma; Peripheral Blood Mononuclear Cell; Phase; Population; Prognostic Marker; Proteins; Recurrence; Reverse Transcriptase Polymerase Chain Reaction; Sampling; Signal Transduction; Single-Stranded DNA; Solid Neoplasm; Solutions; Specificity; Staining method; Stains; Surface; Suspension substance; Suspensions; System; TACSTD2 gene; Techniques; Technology; Testing; Transcript; Tumor Antigens; Validation; Visual; Whole Blood; antigen antibody binding; antigen binding; base; clinical application; clinical care; comparative; cost; cost effective; design; ds-DNA; improved; malignant breast neoplasm; nano; nanoparticle; neoplastic cell; novel; particle; peripheral blood; protein expression; research study; surface coating; survivin; tumor

DESCRIPTION (provided by applicant): Measurement of circulating tumor cells (CTC) has potential to fill major gaps in the clinical care and management of lung cancer patients. Validation of CTC as a prognostic marker in breast cancer patients set an important precedent for CTC measurement; however, histochemical CTC enumeration now used as a clinical correlate in breast and prostate cancer has proven to be neither robust nor reliable enough for application to lung cancer. This is due in large part to the phenotypic heterogeneity and high antigen variability associated with lung tumors as compared to other solid tumors. PCR amplification of cancer protein transcripts is a conceptually appealing alternative approach to CTC measurement that offers sensitivity and unlimited capabilities for multiple antigen measurements. Although literature indicates that PCR detection of CTC correlates with lung cancer stage, disease recurrence and survival, this approach remains investigational. Since mRNA transcript is used as a surrogate for tumor protein expression, PCR is criticized for lack of specificity. We have devised an approach that combines the specificity of immuno-histochemistry with the sensitivity of PCR. This application details testing and optimization of a novel nanoparticle-based system to overcome technical and biological hurdles encountered in lung cancer CTC measurement. Specifically, bio-nanoconjugates are constructed to carry one antigen specific monoclonal antibody and a conjugate-specific double stranded DNA tag sequence for nano-specific identification. When conjugates bind antigen on a cell surface, a single strand of the DNA tag can be released into suspension by briefly heating the complex mixture; liberated DNA strands amplified by PCR provide a highly sensitive and quantifiable measure of cellular binding and corresponding antigen expression. Signal amplification of the antigen-bound bio-nanoconjugate allows detection of low frequency proteins on low concentration CTC in peripheral blood, and multiplexing capabilities inherent to the technology offer increasing sensitivity without reduced specificity, making this particularly well suited for the measurement of lung cancer CTC. Sensitivity of the approach will be determined with peripheral blood spiked at various concentrations with cultured tumor cells with known antigen expression. CTC enrichment using antibody-mediated separation of CTC from peripheral blood mononuclear cells is expected to further increase the limits of detection, while also reducing the potential for false positive results. We anticipate being able to achieve unrivaled detection limits with high degrees of specificity for lung cancer CTC. This application details the comparative and systematic phases of initial development and testing of this promising initiative. PUBLIC HEALTH RELEVANCE: Measurement of circulating tumor cells (CTC) has potential to fill major gaps in the clinical care and management of lung cancer patients. There is not a currently reliable method for clinical application in lung cancer. This application details testing and optimization of a novel nanoparticle-based system to overcome technical and biological hurdles encountered in lung cancer CTC measurement.

描述（由申请方提供）：循环肿瘤细胞（CTC）的测量有可能填补肺癌患者临床护理和管理的主要空白。CTC作为乳腺癌患者预后标志物的验证为CTC测量树立了重要的先例;然而，组织化学CTC计数现在用作乳腺癌和前列腺癌的临床相关性，已被证明既不稳健也不可靠，不足以应用于肺癌。这在很大程度上是由于与其他实体瘤相比，与肺肿瘤相关的表型异质性和高抗原变异性。癌症蛋白转录物的PCR扩增是CTC测量的概念上有吸引力的替代方法，其提供用于多种抗原测量的灵敏度和无限能力。尽管文献表明CTC的PCR检测与肺癌分期、疾病复发和生存相关，但该方法仍处于研究阶段。由于mRNA转录被用作肿瘤蛋白表达的替代物，PCR因缺乏特异性而受到批评。我们设计了一种结合免疫组化的特异性和PCR的灵敏度的方法。本申请详细介绍了一种新型纳米颗粒系统的测试和优化，以克服肺癌CTC测量中遇到的技术和生物学障碍。具体地，构建生物纳米缀合物以携带一种抗原特异性单克隆抗体和缀合物特异性双链DNA标签序列用于纳米特异性鉴定。当缀合物结合细胞表面上的抗原时，DNA标签的单链可以通过短暂加热复杂混合物而释放到悬浮液中;通过PCR扩增的释放的DNA链提供了细胞结合和相应抗原表达的高度灵敏和可定量的测量。抗原结合的生物纳米缀合物的信号放大允许检测外周血中低浓度CTC上的低频蛋白质，并且该技术固有的多路复用能力提供增加的灵敏度而不降低特异性，使得这特别适合于测量肺癌CTC。该方法的灵敏度将用以不同浓度掺入具有已知抗原表达的培养肿瘤细胞的外周血来确定。使用抗体介导的从外周血单核细胞中分离CTC的CTC富集预计将进一步提高检测限，同时还降低假阳性结果的可能性。我们期望能够实现无与伦比的检测限，对肺癌CTC具有高度的特异性。这份申请书详细介绍了这一有希望的倡议的初步开发和测试的比较和系统阶段。 公共卫生关系：循环肿瘤细胞（CTC）的测量有可能填补肺癌患者临床护理和管理的主要空白。目前还没有一种可靠的方法用于肺癌的临床应用。本申请详细介绍了一种新型纳米颗粒系统的测试和优化，以克服肺癌CTC测量中遇到的技术和生物学障碍。