Thiroredoxin Targeted Nanoparticles for Cancer Research

用于癌症研究的硫氧还蛋白靶向纳米颗粒

• 批准号: 7546497
• 负责人: STEVEN Sidney SMITH
• 金额: $ 8.31万
• 依托单位: BECKMAN RESEARCH INSTITUTE/CITY OF HOPE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7546497
• 关键词: Aftercare; Antibodies; Back; Binding; Biological Assay; CWR22Rv1; Cells; Chimeric Proteins; Clinical; Color; Custom; DNA; Diagnostic; Diagnostic Procedure; Epithelial Cells; Fluorescein; Fluoresceins; Freezing; Gleason Grade for Prostate Cancer; Glutamate Carboxypeptidase II; Goals; Histology; Home environment; Image; Imagery; Immunohistochemistry; LNCaP; Label; MCF7 cell; Malignant Neoplasms; Malignant neoplasm of prostate; Methyltransferase; Microscope; Nomograms; PC3 cell line; PSA level; Particle Size; Patients; Prostate; Prostatic Neoplasms; Recurrence; Sampling; Staining method; Stains; Standards of Weights and Measures; Stromal Cells; Technology; Testing; Therapeutic; Thioredoxin; Work; abstracting; anticancer research; cancer cell; design; follow-up; improved; nanobiotechnology; nanoparticle; nanoscale; neoplastic cell; research study; scaffold; tool; tumor

DESCRIPTION (provided by applicant): Abstract Currently an important clinical problem in prostate cancer research is predicting the likelihood of recurrence. Our hypothesis is that histology using nanoparticles displaying thioredoxin on tumor sections will improve on the current standard diagnostic methods (e.g. PSA level and Gleason's score) in predicting the likelihood of recurrence. Using this bionanotechnology we have constructed a nanoparticle consisting of a DNA scaffold displaying three copies of the methyltransferase-thioredoxin fusion protein. The DNA scaffold has been modified so as to permit the visualization of the cancer cells using fluorescein. Our preliminary work shows that the nanoparticle displaying thioredoxin binds selectively to LNCaP and MCF-7, but not PC-3, COS-7 and Primary Prostate Epithelial Cells, suggesting that the nanoparticle is selectively targeting certain types of cancer cells. We propose to follow up on these preliminary findings with the following Specific Aims: Specific Aim 1: (i) Generate nanoparticles comprising a Y-junction DNA scaffold displaying three identical methyltransferase thioredoxin fusion proteins (YRII-Trx-F) in quantities necessary for the experiments described in Specific Aims 2 and 3. (ii) Generate and purify thioredoxin labeled directly with fluorescein (Trx-F) as a control. Specific Aim 2: Test the nanoparticle displaying the thioredoxin (YRII-Trx-F) and thioredoxin labeled directly with fluorescein (Trx-F) for their ability to discriminate between various available prostate cancer cell lines: (e.g. LNCaP, PC-3, DU-145, MDA PCa 2b and 22Rv1) while not detecting primary prostate stromal cells, and primary prostate epithelial cells. Specific Aim 3: Test the nanoparticle displaying the thioredoxin (YRII-Trx-F) and thioredoxin labeled directly with fluorescein (Trx-F) determine whether or not the binding to frozen prostate tumor sections can be correlated with Gleason's score and/or recurrence. For comparison, immunohistochemistry will be performed on sections of each tumor sample to evaluate the level of expression of PSMA and AMACR using anti-PSMA antibodies and and-AMACR antibodies. Each diagnostic assay will be analyzed to see if there is a correlation between the Gleason's Sum, antibody binding intensity and fluorescent intensity of the nanoparticles. Our long-term goals are two-fold: First, we hope that our findings will provide a new parameter that can contribute to a nomogram for the prediction of recurrence. Second, the modular design of the nanoparticle allows us to custom design it to target almost any cancer for potential delivery into cells. If the results of this study show that this nanoparticle can specifically target aggressive prostate tumor cells, we could then modify the design for potential delivery of a therapeutic into the cells. Both goals are beyond the scope of the present proposal. Project Narrative Currently our tools for determining who is in danger of having prostate cancer return are not as helpful as they should be, so we cannot follow up with additional treatment until the cancer reappears. We have developed a technology for creating nanoscale (molecule size) particles that can home in on the cancer cells and make them glow. In this proposal we will attempt to determine whether or not the color we see under the microscope will permit us to pick out those cancers that are likely to come back after treatment and help the patients who are likely to come down with the cancer again to take additional action.

描述（由申请人提供）：摘要目前前列腺癌研究中的一个重要临床问题是预测复发的可能性。我们的假设是，在肿瘤切片上使用展示硫氧还蛋白的纳米颗粒的组织学将在预测复发可能性方面改善当前的标准诊断方法（例如PSA水平和Gleason评分）。使用这种生物纳米技术，我们构建了一种纳米颗粒，由展示三个拷贝的甲基转移酶-硫氧还蛋白融合蛋白的DNA支架组成。DNA支架已被修改，以便允许使用荧光素可视化癌细胞。我们的初步工作表明，展示硫氧还蛋白的纳米颗粒选择性地结合LNCaP和MCF-7，但不结合PC-3、COS-7和原代前列腺上皮细胞，这表明纳米颗粒选择性地靶向某些类型的癌细胞。我们建议以以下具体目的对这些初步发现进行后续研究：具体目的1：（i）产生包含Y-连接DNA支架的纳米颗粒，所述Y-连接DNA支架以具体目的2和3中所述实验所需的量展示三种相同的甲基转移酶硫氧还蛋白融合蛋白（YRII-Trx-F）。(ii)产生并纯化直接用荧光素（Trx-F）标记的硫氧还蛋白作为对照。具体目标二：测试展示硫氧还蛋白（YRII-Trx-F）和直接用荧光素标记的硫氧还蛋白（Trx-F）的纳米颗粒区分各种可用前列腺癌细胞系（例如LNCaP、PC-3、DU-145、MDA PCa 2b和22 Rv 1）而不检测原代前列腺基质细胞和原代前列腺上皮细胞的能力。具体目标3：测试展示硫氧还蛋白（YRII-Trx-F）和直接用荧光素标记的硫氧还蛋白（Trx-F）的纳米颗粒，确定与冷冻前列腺肿瘤切片的结合是否可以与格里森评分和/或复发相关。为了进行比较，将对每个肿瘤样本的切片进行免疫组织化学，以使用抗PSMA抗体和抗AMACR抗体评价PSMA和AMACR的表达水平。将分析每个诊断测定以查看格里森总和、抗体结合强度和纳米颗粒的荧光强度之间是否存在相关性。我们的长期目标是双重的：首先，我们希望我们的发现将提供一个新的参数，可以有助于预测复发的诺模图。第二，纳米颗粒的模块化设计使我们能够定制设计它，以靶向几乎任何癌症，并将其输送到细胞中。如果这项研究的结果表明，这种纳米颗粒可以特异性地靶向侵袭性前列腺肿瘤细胞，那么我们可以修改设计，将治疗药物输送到细胞中。这两个目标都超出了本提案的范围。目前，我们用于确定谁有前列腺癌复发的危险的工具并不像它们应该的那样有用，所以我们不能随访额外的治疗，直到癌症复发。我们已经开发出一种制造纳米级（分子大小）颗粒的技术，这种颗粒可以定位在癌细胞上并使它们发光。在这个建议中，我们将试图确定我们在显微镜下看到的颜色是否允许我们挑选出那些可能在治疗后复发的癌症，并帮助那些可能再次患上癌症的患者采取额外的行动。