Imaging Survivin mRNA for Cancer Detection

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DESCRIPTION (provided by applicant): The dichotomous expression of survivin mRNA in normal versus cancer cells is among the most tumor-specific of all human gene products thus garnering great interest as a potential diagnostic marker and a therapeutic target. The goal of this investigation is to develop a novel radiolabeled antisense probe against survivin mRNA as a next-generation radiopharmaceutical for in vivo mapping of this transcript. The dichotomous expression of survivin mRNA may lead to a selective retention of radioactivity on tumors mediated by the in vivo hybridization. Capturing the radioactive signal with a gamma camera will produce a "clean-cut" tumor imaging-antisense imaging. Using technetium-99m (99mTc) radiolabeled antisense DNA probe to RIalpha mRNA, we have observed what is almost certainly an antisense effect. Our results, therefore, show that antisense imaging is feasible and could already be achievable if the tumor uptake and tumor/nontumor ratios can be improved. We, therefore, propose a series of novelties designed to accomplish this goal: 1). Select a superior antsiense sequence targeting a common stretch shared by all human survivin mRNA splice variants. 2). Use a just-emerging synthetic oligomer morpholino (MORF) bearing superior properties as antisense chemical form. 3). Design a multifunctional Tat vector with cell transduction potency to enhance cell uptake and accelerate clearance. MORF will be conjugated to the overhung cysteine on the Tat vector to form chimeric antisense construct (Tat-MORF), which can easily be radiolabeled with 99mTc via a built-in N2S2 chelator by a one-step procedure. The 99mTc-Tat-MORF will be evaluated first in cell culture with tumor cells overexpressing the survivin mRNA (Survivin+) and tumor cells without expression of human survivin mRNA (Survivin-). The kinetics of cellular accumulation and efflux will be measured with antisense vs. scrambled control in Survivin+ and Survivin- tumor cells. Inhibition assay and the correlation of antisense accumulation to the level of target mRNA expression will be performed to reinforce the antisense mechanism, ie. the selective antisense accumulation in target cells is mediated by hybridization of antisense probes. For in vivo study, a dual-xenograft tumor model in nude mice will be established bearing Survivin+ and Survivin- tumors. This animal model will allow us to address almost all concerns for antisense tumor targeting. Longitudinal scintigraphies will be complemented with periodical biodistribution of tissue dissection and whole body autoradiographies to supplement relative tissue radioactivity measurements with absolute measurements. We believe that using this novel chimeric antisense probe as a next-generation radiopharmaceutical to target the dichotomous expression of survivin mRNA would allow us to obtain a "clean-cut" tumor imaging by antisense mechanism. This "proof-of-concept" of antisense imaging will be helpful to provide a paradigm of the merger of modern biology and conventional nuclear medicine.