Thiroredoxin Targeted Nanoparticles for Cancer Research

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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.