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San Antonio, TX, United States

Bruno J.G.,Operational Technologies Corporation
In Vitro Cellular and Developmental Biology - Animal

Nucleic acid aptamers are regarded as rivals for antibodies and as such are being investigated for their therapeutic potential. In the present work, it is shown that two different high-affinity DNA aptamers developed previously by Ferreira et al. against MUC1 antigen (designated MUC1-5TR-1 and MUC1-S1.3/S2.2) on MCF7 breast cancer cells can be linked to the first component of complement (C1q) via a biotin-streptavidin system and induce significant killing of MCF7 cells in vitro. Cell viability was assessed by Trypan blue uptake and absorbance at 590 nm of stained cells following buffer washes and lysis in 1% SDS. While the killing effect is demonstrable versus various controls, dependent on aptamer dose, and reproducible, it appears to kill maximally about half of treated MCF7 cells. Possible reasons for the marginal killing effect include antigenic shedding in vitro and membrane-bound complement regulatory proteins (mCRPs) on the cell surface such as CD46, CD55, and CD59 which act to inhibit complement-mediated lysis of cells. Future in vitro research could benefit from application of mCRP-specific aptamers in combination with anti-MUC1 aptamers to overcome surface protective mechanisms while attacking the plasma membrane of MCF7 cells or other MUC1-expressing cancer cells. However, in vivo such a combination could have deleterious effects on normal MUC1-expressing cells as well. © 2009 The Society for In Vitro Biology. Source

Bruno J.G.,Operational Technologies Corporation

Despite the great promise of nucleic acid aptamers in the areas of diagnostics and therapeutics for their facile in vitro development, lack of immunogenicity and other desirable properties, few truly successful aptamer-based products exist in the clinical or other markets. Core reasons for these commercial deficiencies probably stem from industrial commitment to antibodies including a huge financial investment in humanized monoclonal antibodies and a general ignorance about aptamers and their performance among the research and development community. Given the early failures of some strong commercial efforts to gain government approval and bring aptamer-based products to market, it may seem that aptamers are doomed to take a backseat to antibodies forever. However, the key advantages of aptamers over antibodies coupled with niche market needs that only aptamers can fill and more recent published data still point to a bright commercial future for aptamers in areas such as infectious disease and cancer diagnostics and therapeutics. As more researchers and entrepreneurs become familiar with aptamers, it seems inevitable that aptamers will at least be considered for expanded roles in diagnostics and therapeutics. This review also examines new aptamer modifications and attempts to predict new aptamer applications that could revolutionize biomedical technology in the future and lead to marketed products. © 2015 by the authors; licensee MDPI. Source

Bruno J.G.,Operational Technologies Corporation

The potential to emulate or enhance antibodies with nucleic acid aptamers while lowering costs has prompted development of new aptamer-protein, siRNA, drug, and nanoparticle conjugates. Specific focal points of this review discuss DNA aptamers covalently bound at their 3′ ends to various proteins for enhanced stability and greater pharmacokinetic lifetimes in vivo. The proteins can include Fc tails of IgG for opsonization, and the first component of complement (C1q) to trigger complement-mediated lysis of antibiotic-resistant Gram negative bacteria, cancer cells and possibly some parasites during vulnerable stages. In addition, the 3′ protein adduct may be a biotoxin, enzyme, or may simply be human serum albumin (HSA) or a drug known to bind HSA, thereby retarding kidney and other organ clearance and inhibiting serum exonucleases. In this review, the author summarizes existing therapeutic aptamer conjugate categories and describes his patented concept for PCR-based amplification of double-stranded aptamers followed by covalent attachment of proteins or other agents to the chemically vulnerable overhanging 3′ adenine added by Taq polymerase. PCR amplification of aptamers could dramatically lower the current $2,000/gram cost of parallel chemical oligonucleotide synthesis, thereby enabling mass production of aptamer-3′-protein or drug conjugates to better compete against expensive humanized monoclonal antibodies. © 2013 by the authors; licensee MDPI, Basel, Switzerland. Source

Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 150.00K | Year: 2014

Development of novel antifungal drugs is spurring interest in the detection of pathogenic fungi in archival formalin-fixed tissues. While in situ hybridization and PCR-based methods for fungal detection in tissues exist. formaldehyde adducts on fungal DNA complicate or invalidate such detection methods. Operational Technologies Corporation (OpTech) has been developing DNA aptamers against a broad array of small molecule, protein and whole cell {bacterial and parasite) targets for the last decade. Much of OpTech's aptamer R & D work is published, patented, and the best (highest affinity and most specific) resultant DNA aptamers are sold on its website: www.OTCBiotech.com which has been profitable since inception in late 2012. In 2010-2011, OpTech developed and published DNA aptamers capable of specifically detecting and binding the N-acetylglucosamine (NAG) monomer of the chitin polymer in fungalcell walls for NASA under an SBIR contract to aid in protecting the recycled water supply aboard the InternationalSpace Station. OpTech now proposes to develop and characterize aptamers capable of specifically binding and detecting formalin-fixed Blastomyces, Histoplasma, Fusarium, and Scedosporium as well as the Mucormycetes (Mucor, Rhizopus, Absidia, and Cunninghamella) fungi at the genus level and species level in parffin-embedded tissue sections by fluorescence and/or peroxidase staining.

Agency: Environmental Protection Agency | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 79.98K | Year: 2011

"Operational Technology Corporation (OpTech) proposes to couple the ultrasensitivity of fluorescence-based detection with the high selectivity of synthetic DNA aptamers as replacements for antibodies to detect chemical and biological (CB) terrorism threats in 10 minutes or less from a variety of swabbed surfaces and liquid matrices. OpTech has previously provided strong proof-of-concept for a rapid one-step (“bind and detect”) Fluorescence Resonance Energy Transfer (FRET) aptamer assay for foot and mouth disease virus (J. Biomolec. Techniques, 2008), a nerve gas core molecule (J. Fluorescence 2008) as well as collagen breakdown peptides and calcidiol to monitor bone metabolism in astronauts (J. Clin. Ligand Assay, 2007). This patent-pending FRET-aptamer assay technology can now be applied in lyophilized form in plastic cuvettes to detect a variety of CB analytes on-site with low ng/ml sensitivity using a commercial off-the-shelf (COTS) handheld and battery-operated fluorometer. In Phase 1, OpTech will screen its existing methylphosphonic acid (MPA; nerve agent core), soman derivative, and Botulinum toxin A (BoNT A) candidate DNA aptamer sequences for FRET assay potential. OpTech will also develop aptmers to “anthrose” surface glycoprotein terminal sugar for detection of Bacillus anthracis spores. In Phase 2, OpTech will fully develop and lyophilize the selected FRET-aptamer assays in plastic cuvettes for use with the sensitive COTS handheld fluorometer for facile, rapid, and cost-effective detection of MPA, sarin or soman, BoNTs and anthrax spores. OpTech will also expand its repertoire of assays in Phase 2 to include more CB treat agents (viral and bacterial agents of interest). The primary market for OpTech’s initial CB FRET-aptamer assays will be federal and state first responders as well as military personnel. Clearly, however, the one-step FRET-aptamer assay approach coupled to a handheld COTS fluorometer constitutes a platform technology with broad applications in clinical diagnostics, veterinary diagnostics, food safety testing, environmental and agricultural monitoring. "

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