Development and Validation of a Novel Biomarker Assay for Cancer Diagnostics

Period of Performance: 06/15/2015 - 04/30/2016

$375K

Phase 1 SBIR

Recipient Firm

Blondin Bioscience, LLC
Birmingham, AL 35203
Principal Investigator

Abstract

DESCRIPTION (provided by applicant): In this Phase I project, Blondin Bioscience LLC, a start-up company in Birmingham, AL, proposes to develop a rapid, accurate and precise fluorometric assay for quantifying cell-free telomeric DNA fragments (cf-tDNA) within serum in order to monitor the effectiveness of chemotherapy treatments. In our preliminary work, conducted at the University of Alabama at Birmingham (UAB), we have detected a significant increase in cf- tDNA released from cancer cells into extracellular milieu following chemotherapy-induced apoptosis. We also found a reproducible serum peak of cf-tDNA release following initiation of chemotherapy in leukemia patients. We attribute this release of cf-tDNA to in vivo cancer cell apoptosis. We propose that the measurement of cf-tDNA would decrease the time, cost, and side effects of a potentially inadequate cancer treatment, allowing early switching to a better treatment. Blondin is negotiating an exclusive license for the use of this intellectual property from UAB. Currently cf-tDNA are measured with quantitative PCR (qPCR), which is cumbersome and imprecise, due to the difficulty in amplifying repetitive telomere sequences, and the very low (ng/ml) amounts of cell-free DNA obtained from clinical samples. This new assay will utilize novel, gamma modified peptide nucleic acid (γPNA) miniprobes that are complimentary to the telomeric sequence. γPNA hybridize to complementary DNA with 100,000 fold higher affinity than unmodified PNA. This increase in binding affinity should allow detection of the very dilute target cf-tDNA found in clinical samples. The γPNA miniprobes will be designed to exhibit a FRET signal only in the presence of a telomeric DNA target, thus further improving the signal to noise ratio. The proposed research will have three specific aims. In the first aim we will characterize and adapt γPNA miniprobes for use in a purified model system to detect and quantify cf-tDNA. Test conditions with the best precision and accuracy at the lowest target concentrations will be determined. The second aim is directed toward optimizing and validating the γPNA cf-tDNA assay for use with complex biological samples. The assay will be validated by assessing the accuracy, precision, specificity, limit of detection and quantification, linearity and range, ruggedness, and robustness as per current FDA method validation guidelines. The final aim will assess the γPNA cf-tDNA assay in a clinical setting. We will test the ability of the γPNA cf-tDNA assay to prospectively predict survival in a trial enrolling 30 metastatic prostate cancer patients starting chemotherapy treatment. The main goal of this aim is to test the performance of the γPNA cf-tDNA assay in a clinical situation. The study is statistically powered to determine if the γPNA cf-tDNA assay is able to outperform the current response biomarkers prostate specific antigen and circulating tumor cells in this group of patients.