Development of an Analytical Detection Platform Based on Smart Molecular Probes for Cancer Diagnostics Applications (Phase 2)

The project described in this proposal is an extension of an earlier startup project that was completed in 2017. In the completed start-up project (Phase 1), a new smart probe (SP) was designed and fully characterized for in vitro detection of miR-21, a cancer biomarker. The final report was submitted to Deanship of Scientific Research (DSR) and was approved. The work has been published (S. A. Oladepo, Appl. Spectrosc. 2018, 72: 79-88). In the new project being proposed here (Phase 2), the second and final phase of the startup proposals work will be carried out. Reviewers of the original startup proposal had recommended that a portion of that original proposal be reserved for a second project due to time constraints. Hence, this proposal is addressing the second part of that project, as recommended by the reviewers. In this new project, we will further optimize and refine the original SP to increase its sensitivity and reduce its melting temperature (Tm). Increasing the sensitivity will lower the detection limit, while reducing the Tm will allow the SP to be used with simple fluorescence spectrometers that do not have temperature programming. This new SP will be fully characterized and tested. This will involve determining its thermal profiles (under various conditions of ionic strength and salt concentrations) and its Tm to ensure excellent performance characteristics. We will also determine its limit of detection (LOD), limit of quantitation (LOQ) and sensitivity, using a model miR-21 sequence. In addition, we will make and characterize a second probe containing 2-aminopurine (2AP) in a similar fashion. Having established standard optimum conditions for these new probes, we will use both probes for the direct detection of miR-21 cancer biomarker in MCF-7 cancer cell line. This detection platform will constitute a simple, mix-and-read homogeneous assay that is sensitive and fast; a notable improvement over current detection methods. We will then patent this simple, mix-and-read homogeneous detection method that is developed from this work. It must be mentioned that this proposal was previously submitted to DSR in the last cycle but it was rejected because one out of four independent reviewers did not recommend it for funding. The main argument of this reviewer is that the comments and recommendations of the reviewers of the final report of the completed Phase 1 project should have been included in the proposal. The Research Committee of DSR decided to uphold this recommendation despite the fact that DSR has that information (final report) and the outcome of the Phase 1 project has already been published. This reviewer should have just checked out the publication to see the outcome of the Phase 1 project. Regardless, we have now incorporated the comments and recommendations of the reviewers of the final report of the Phase 1 project to this proposal (Appendix A). We have also addressed other minor points raised by the reviewers of the previous submission in this revised proposal. The reviewers comments of the previous submission of this proposal is also appended for reference (Appendix B).