Lexington, KY, United States
Lexington, KY, United States
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Grant
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 220.20K | Year: 2016

DESCRIPTION provided by applicant ParaTechs Corporation aims to provide researchers with advanced technology and methods to simplify procedures for laboratory animal models While actively supporting efforts to `reduce replace and refineandapos animal use ParaTechs provides safe and efficient alternatives for surgical procedures We have already brought to market a nonsurgical embryo transfer device NSETTM which replaces surgical embryo transfers with a nonsurgical method that takes only seconds and does not require anesthesia We have developed a new protocol for nonsurgical artificial insemination AI of mice which replaces existing protocols for surgical AI and is a technical advancement for existing nonsurgical AI protocols We plan to use this newly developed protocol to revolutionize the process of archiving and global distribution of mouse models through a novel sperm cryopreservation and strain recovery system The use of mouse strains in research is one of the most important tools used to study mammalian gene function While hundreds of thousands of mouse strains have been catalogued and cryopreserved sharing of strains is becoming increasingly difficult as regulatory constraints and the cost of shipping live animals increase We can solve this problem and provide a technically superior alternative with the cryopreservation and insemination Candamp I device This system will integrate storage and shipment of cryopreserved sperm with nonsurgical artificial insemination The device will be suitable for storage of cryopreserved sperm will be shippable will allow viable sperm recovery upon thawing and then will be used directly for nonsurgical artificial insemination This device will eliminate the need to perform in vitro fertilization IVF upon thawing of cryopreserved sperm greatly reducing the number of mice required for the procedure and replacing a surgical method of artificial insemination both of which promote animal health and welfare Therefore the goals of this project are to design manufacture and test a Candamp I device for mice The goals will be accomplished in two aims First a Candamp I device prototype will be designed and produced Second the feasibility of using the Candamp I device prototype for sperm cryopreservation and mouse strain recovery will be tested This new technological breakthrough will provide an alternative to cryorecovery of mouse strains by in vitro fertilization methods the currently preferred but technically challenging and labor intensiv strain recovery method In doing so the Candamp I device will provide a substantial cost savings to researchers and a convenient method for shipping and strain recovery PUBLIC HEALTH RELEVANCE The ability to genetically modify the laboratory mouse is now producing an ever rising number of increasingly valuable and unique strains to investigate human disease While maintaining these mouse strains has become a significant and recurring effort and budgetary commitment cryopreservation of strains in sperm repositories has proven successful as a means to permanently and economically preserve them ParaTechs proposes to design manufacture and test a cryopreservation and insemination Candamp I device for the purpose of integrating cryopreservation storage shipping and strain recovery of mice through a newly developed nonsurgical artificial insemination method


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 216.25K | Year: 2013

DESCRIPTION (provided by applicant): The baculovirus expression vector system (BEVS) has been successfully utilized to produce thousands of proteins for use as vaccines, therapeutics, and for structure-function studies. One limitation of BEVS is the propensity of baculoviruses to accumulate transposon insertions into the fp25k gene leading to the few polyhedra (FP) phenotype. This mutation shifts the balance of virus production from occlusion-derived viruses, which are not infectious in tissue culture, tobudded viruses, which are the form of virus that is used in baculovirus expression. These higher levels of budded virus would be advantageous for BEVS users, but FP mutants are also deficient in transcription from the polyhedrin promoter, which drives expression of target genes. Baculoviruses also rapidly accumulate defective interfering particles (DIP), which are linked to a sharp decrease in target gene expression due to deletion of the target gene and/or viral genes needed for its expression. One factorthat promotes DIP formation is transposition into fp25k. The goal of this project is to develop baculovirus expression vectors that allow for manipulation of the fp25k gene. By reducing or eliminating expression of FP25K during virus amplification, we expect to obtain 5- to10- fold higher levels of budded virus titers because most of the replicative potential of the cel would go to producing budded virus instead of occlusion-derived virus. Then FP25K expression would be restored when target protein expression is desired. This would significantly lower costs for large-scale production of baculovirus-expressed proteins because high titer BV stocks would be easier to produce and the occurrence of deleterious mutations would be reduced. ParaTechs will pursue two complementary approaches to achieve this goal. One involves the production of a virus with an inducible fp25k gene that can be turned off during amplification and activated during target protein expression. The other approach utilizes a virus with a deletion in fp25k coupled with a cell line that expresses FP25K. Each of these approaches has advantages and disadvantages, both in the design phase and for the end user. Experiments described in this proposal will determine which provides higher levels of BVproduction, polyhedrin-linked expression, and stable genome maintenance. Ultimately, the system developed here would be combined with ParaTechs vankyrin expression technology, which increases polyhedrin-driven expression in BEVS from 2- to 20-fold.PUBLIC HEALTH RELEVANCE PUBLIC HEALTH RELEVANCE: The baculovirus expression vector system (BEVS) is commonly utilized to produce thousands of proteins for use as vaccines, therapeutics, and for structure-function studies. Unfortunately many researchersare not aware that certain mutations rapidly accumulate leading to lower levels of target gene expression. ParaTechs has developed a strategy to prevent these mutations, while simultaneously providing higher levels of virus production, stable genome maintenance, and target gene expression.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 200.98K | Year: 2011

DESCRIPTION (provided by applicant): Most cell culture lines are anchorage-dependent and require surface attachment for proliferation. For industrial production, increased surface area can be provided by microcarrier beads, but the associated increases incost and the resulting complexity of manipulation may preclude their use. Due to these limitations, anchorage-independent cells are preferred for the production of biopharmaceuticals, but appropriate anchorage-independent cells are not available for all applications; e.g., production of vaccines and cell-type specific proteins. This proposal explores the use of a novel insect virus protein to transform adherent cells to cells that can thrive in suspension culture. ParaTechs has identified a cell line fromAgrotis ipsilon (black cutworm), which provides levels of recombinant protein expression that are 3-to-10 times higher than the standard Sf9 cell line. Unfortunately, the cells are strongly adherent, which makes them unsuitable for large-scale protein production. We intend to stably transform the A. ipsilon cells with an insect virus gene that causes a loss of adhesion in hemocytes. We predict that expression of this protein in transformed cells will enable them to grow in suspension culture. In combinationwith our Vankyrin-Enhanced Baculovirus Expression Vector System (VE-BEVS), which has the ability to increase protein production per cell by a factor of 4 to 22, we anticipate achieving a level of protein expression per cell that is at least 12-to-220 times higher than currently possible. This dramatic increase in yield will be significant for all BEVS users, from individual researchers to large biopharmaceutical companies. We also will apply this technology to adherent mammalian tissue cultures. A simple method for converting mammalian cells from anchorage-independent to suspension culture would make a significant contribution to the production of vaccines and bio-therapeutics, and would result in improved human health. ParaTechs personnel are experiencedbaculovirologists and cell biologists. The protocols use standard technologies that are routinely adopted in our company. We do not anticipate difficulty with the experimentation. Current literature strongly supports our hypothesis and we are confident ofour success. PUBLIC HEALTH RELEVANCE: The future of human medicine will involve a dramatic increase in development of protein-based drugs and subunit vaccines, whose production will require large-scale propagation of tissue culture cells in suspension culture. Most continuous cell lines, however, are anchorage-dependent, and no method exists to routinely and easily transform adherent cells to suspension culture. ParaTechs will test an insect virus gene that causes hemocytes to lose adhesion for the ability to adapt anchorage- dependent insect and mammalian cells to suspension culture. Achievement of this goal would make a significant contribution to biopharmacology and human health.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 217.54K | Year: 2013

DESCRIPTION (provided by applicant): Chronic wounds and ulcers are increasing with the growing incidence of diabetes and associated illnesses and have annual costs exceeding 20 billion. These non-healing wounds result in chronic disease, limb amputationsand death and are complicated by antibiotic resistant bacteria infection. This proposal seeks to improve and develop new capabilities of a biological treatment of such wounds, maggot debridement therapy (MDT) that is effective but underutilized. Some maggot species have evolved to digest fluids, dead tissues and bacteria in wounds and have been recognized for over 200 years for this improved wound healing behavior. For approximately a century, Lucilia sericata maggots have been produced for MDT. This proposal seeks to create transgenic flies with properties that will support increased use of MDT. The goals of the proposal are 1) to produce flightless flies with limited dispersal capability, 2) to engineer fluorescent flies that are more readily observed in wounds, 3) to engineer flies that facilitate analgesic and therapeutic peptide expression to improve patient comfort and wound healing. These mutants will facilitate fly production and also impede genetically engineered insect escape. The objectives of thisproposal will support greater use of MDT thereby reducing treatment costs, improving wound healing and limiting the persistent use of antibiotics for treating chronic wounds. PUBLIC HEALTH RELEVANCE PUBLIC HEALTH RELEVANCE: Chronic wounds and ulcers are increasing with the growing incidence of diabetes and associated illnesses and have annual costs exceeding 20 billion. Concurrently, treatment of bacterial infections that are common complications of chronic wounds and ulcers is becoming more difficult with the rapid emergence of antibiotic resistant bacteria. This proposal seeks to expand the use and improve the efficacy of a biological method for treatment of chronic wounds and their associated diseases. )


Grant
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase II | Award Amount: 1.04M | Year: 2015

DESCRIPTION provided by applicant The baculovirus expression vector system BEVS has been successfully utilized to produce thousands of proteins for use as vaccines and therapeutics as well as for studies of protein structure and function One limitation of BEVS is the propensity of baculoviruses to accumulate transposon insertions into the fp k gene leading to the andquot few polyhedra FP andquot phenotype This mutation shifts the balance of virus production from occlusion derived viruses which are not infectious in tissue culture to budded viruses BV which are the form of virus that is used in baculovirus expression Higher levels of BV would be advantageous for BEVS users but FP mutants are also deficient in transcription from the polyhedrin promoter that drives expression of target genes Baculoviruses also rapidly accumulate defective interfering particles DIP which are often linked to a sharp decrease in target gene expression due to deletion of the target gene and or the viral genes needed for its expression One factor promoting DIP formation is transposition into fp k The goal of this proposal is to limit deleterious effects of transposition into fp k while taking advantage of the fact that elimination of the FP K activity significantly increases BV production During Phase I two strategies an inducible construct for controlling fp k and an fp k deletion mutant coupled with a cell line constitutively expressing FP K were explored for regulating FP K expression Both strategies sought to produce high titer virus during the amplification stage of baculovirus infection and enable a switch to high level transcription from the polyhedrin promoter during the recombinant protein expression phase Results document achievement of Phase I objectives with deletion of fp k from the virus and complementary expression from an engineered cell line enabling the predicted control of budded virus and recombinant protein production However the inducible construct did not provide sufficient FP K control due to the toxicity of the heavy metal inducer to insect cells Therefore in Phase II we will develop an improved inducible construct with tighter transcriptional regulation using an ecdysone receptor based inducible promoter validate the beneficial effects of FP K regulation using viruses that express intracellular and secreted yellow fluorescent protein YFP for BEVS and BacMam mammalian expression technology develop user friendly fp k mutant backbones for simplified BEVS and BacMam cloning and demonstrate their utility with several medically relevant transgenes and test the fp k expression system in the context of vankyrin enhanced BEVS VE BEVSTM technology ParaTechsandapos VE BEVS products delay death and lysis of baculovirus infected cells thereby boosting target protein expression up to fold Many BEVS users would want to incorporate both vankyrin and FP K technologies therefore it is important to determine whether they are compatible Taken together completion of Phase II objectives will enable users of the BEVS to control and optimize production of BV and recombinant protein expression PUBLIC HEALTH RELEVANCE The baculovirus expression vector system BEVS is commonly used for production of proteins for structure function studies vaccines and therapeutics Unfortunately many researchers are not aware that certain mutations rapidly accumulate leading to lower levels of target gene expression ParaTechs has developed a strategy to prevent these mutations while simultaneously providing higher levels of virus production stable genome maintenance and target gene expression


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase II | Award Amount: 1.07M | Year: 2012

DESCRIPTION (provided by applicant): The baculovirus expression vector system (BEVS) is a proven, powerful and versatile method of eukaryotic protein expression. It is used to produce vaccines, diagnostics, and biologically active proteins for a multitudeof research projects. Like all expression systems, however, BEVS has its disadvantages. One is the fact that expression is short-lived due to virus-induced cell death and lysis. ParaTechs has already commercialized a product that addresses this shortcoming. Cell lines that express a viral ankyrin gene show delayed death and lysis of baculovirus-infected cells, thereby significantly enhancing recombinant protein production. This activity, referred to as vankyrin-enhanced BEVS (VE-BEVSTM), boosts target protein expression up to 22-fold. A second limitation of baculovirus expression is that insect cells lack the ability to produce terminally sialylated, complex N-glycans, which limits the usefulness of BEVS for the expression of human therapeutic proteins. GlycoBac LLC has developed a transgenic insect cell line (SfSWT4) that expresses six mammalian glycosylation enzymes, allowing synthesis of terminally sialyated proteins. This Phase II proposal combines ParaTechs' VE technology with GlycoBac's cell line to optimize expression of humanized N-glycans. In Phase I, the transgenic cell line SfSWT4 was transformed with several vankyrin genes under the control of different promoters. Polyclonal cells were screened for enhanced glycoprotein expression. Data demonstrating that infected cells lived longer and produced more authentically sialylated protein confirmed our hypothesis. Phase II will extend these studies by (1) cloning and characterization of VE-SWTTM cells according to FDA guidelines for cells used to produce vaccines and biological; (2) examining synergistic effects between VE virus vectors and VE-SWT cell lines to provide greater levels of enhancement; and (3) demonstrating enhanced expression of medically relevant glycoproteins in VE-SWTTM cells. These Phase II studies will significantly expand the applications of ParaTechs' VE-BEVS technology and GlycoBac's glycoengineered cell lines. Personnel at both companies have experience with the techniques to be used. Preliminary studies indicate that this technology has a significant chance of performing as envisioned. Furthermore, prior marketing experience with transformed cell lines previously released from the two companies suggests a significant demand for the expanded technology. This new technology shouldbe relatively easy to commercialize based on the established reputations of ParaTechs, Inc. and GlycoBac, LLC and the growing demand for improved cell lines to express recombinant humanized glycoproteins. PUBLIC HEALTH RELEVANCE: The inability ofinsect cells to produce terminally sialylated, complex N-glycans limits the usefulness of the baculovirus expression system for the production of human therapeutic proteins. This proposal addresses that deficiency and aims to develop new cell lines that produce authentic humanized N-glycans in the context of ParaTechs' vankyrin-enhanced baculovirus technology. Successful completion of these objectives will produce cells that provide the highest levels of accurately processed secreted and transmembrane proteins and will be marketed to individual researchers and pharmaceutical companies engaged in structure-function studies of the human secretome and development of protein therapeutics.


Grant
Agency: National Science Foundation | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 150.00K | Year: 2014

The broader impact/commercial potential of this Small Business Innovation Research (SBIR) Phase I project is the development of a technology to sustain the efficacy of transgenic plants expressing the Bacillus thuringiensis (Bt) toxin. Transgenic expression of Bt toxins is a major tool for the control of insect pests in agriculturally important crops. Bt transgenic technology has been deployed on over one billion acres because it improves crop yields and reduces the environmental problem of using toxic chemical pesticides. Insects, however, are becoming increasingly resistant to the Bt toxin, which is threatening its sustainability. This project proposes to develop alternative means to suppress populations of important lepidopteran pest(s) targeted by Bt. There is no greater threat to sustainability of the Bt technology than the development of resistance, with current evidence indicating that resistance is increasingly impacting producers globally. Novel strategies to suppress or eliminate insect populations resistant to Bt represent a significant commercial opportunity. This SBIR Phase I project proposes to develop a novel biological control agent for use in specifically suppressing insect populations that are developing resistance to the Bt toxin. This project will produce a biological control agent, and test it for the ability to replicate in the target insect and determine if it has the predicted pathology. Once the agent is developed and its properties evaluated in the laboratory, experiments will be performed to establish methods for efficient infection of target pests. Phase II will assess impacts on pest populations in microcosm experiments with data used to develop population models of effects in the field. Successful experimental results will develop the technology to the point of field testing and commercialization planned for Phase III.


Grant
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase II | Award Amount: 920.70K | Year: 2015

DESCRIPTION provided by applicant ParaTechs Corporation develops innovative technologies for biomedical research while seeking to improve animal welfare for laboratory animals One of ParaTechsandapos major goals is to further the guiding principles of ethical animal use by andapos reducing replacing and refiningandapos the use of animals in research according to the guidelines of Russell and Burchandapos s Rs of animal research ParaTechs already has brought to market a non surgical embryo transfer device NSETTM for mice which replaces surgical embryo transfers with a non surgical method that has been proven to cause less stress in research animals The goal of this project is to develop a device and a research protocol for nonsurgical embryo transfer in rats The device and protocol is to be rapid easy to use cost effective and efficien for embryo transfer Poof of principle for the project was established during Phase I where multiple prototypes of a rat NSET device were designed manufactured and tested Rat embryos were transferred and viable rat pups were obtained demonstrating the first successful nonsurgical embryo transfer in rats The goal of Phase II is to further establish the parameters of rat NSET technology and to develop the device for commercialization to the biomedical research community Four objectives are proposed to achieve this goal First we will determine the efficiency of non surgical embryo transfer relative to surgical embryo transfer in Sprague Dawley and Fischer rats for both morula and blastocyst stage embryos This study will have broad applications for transfer of cryopreserved embryos and for transfer of genetically modified embryos after introduction of embryonic stem cells Second we will transfer in vitro cultured embryos using the rNSET and compared efficiency to surgical embryo transfer techniques This aim will address successful transfer of rat embryos that may be modified very early in development for example after pronuclear injection for the purpose of genetic modification Third we will test the use of the rat NSET for artificial insemination as an alternative to surgical techniques This aim will support the use of the device as an alternative t surgical artificial insemination and as a possible alternative to in vitro fertilization Fourth w will assess the level of stress reduction of the NSET procedure compared to surgical embryo transfer These aims are consistent with ParaTechsandapos overall goal to improve animal welfare and support more effective and humane use of animal models in biomedical research PUBLIC HEALTH RELEVANCE The ability to genetically modify the laboratory rat is producing increasingly valuable and unique strains to investigate human disease Currently this technology is dependent on implanting embryos in female rats by surgical means ParaTechs proposes to develop and commercialize a unique device and technology for fast simple and painless nonsurgical embryo transfer NSET in rats which will support the Office of Laboratory Animal Welfare OLAW objectives of reducing pain and distress for laboratory animals because it will provide an alternative to surgical methods for assisted reproductive techniques


Grant
Agency: NSF | Branch: Standard Grant | Program: | Phase: | Award Amount: 150.00K | Year: 2014

The broader impact/commercial potential of this Small Business Innovation Research (SBIR) Phase I project is the development of a technology to sustain the efficacy of transgenic plants expressing the Bacillus thuringiensis (Bt) toxin. Transgenic expression of Bt toxins is a major tool for the control of insect pests in agriculturally important crops. Bt transgenic technology has been deployed on over one billion acres because it improves crop yields and reduces the environmental problem of using toxic chemical pesticides. Insects, however, are becoming increasingly resistant to the Bt toxin, which is threatening its sustainability. This project proposes to develop alternative means to suppress populations of important lepidopteran pest(s) targeted by Bt. There is no greater threat to sustainability of the Bt technology than the development of resistance, with current evidence indicating that resistance is increasingly impacting producers globally. Novel strategies to suppress or eliminate insect populations resistant to Bt represent a significant commercial opportunity.

This SBIR Phase I project proposes to develop a novel biological control agent for use in specifically suppressing insect populations that are developing resistance to the Bt toxin. This project will produce a biological control agent, and test it for the ability to replicate in the target insect and determine if it has the predicted pathology. Once the agent is developed and its properties evaluated in the laboratory, experiments will be performed to establish methods for efficient infection of target pests. Phase II will assess impacts on pest populations in microcosm experiments with data used to develop population models of effects in the field. Successful experimental results will develop the technology to the point of field testing and commercialization planned for Phase III.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase II | Award Amount: 1.03M | Year: 2011

DESCRIPTION (provided by applicant): The use of research animals in research has been essential to development of vaccines and study of most human diseases. Modern rodent research is increasingly powerful with the ability to manipulate the genomes of miceand rats such that they closely mimic complex human diseases such as Alzheimer's and artherosclerosis. Generating transgenic mice and rats, requires that embryos that are manipulated by researchers be transferred into recipient female mice where they can complete their development. Up to this point, these transfers have required surgical procedures in which the embryo is implanted into the uterus. Recently, a device has been developed that enables these embryo transfers to be performed without surgery whicheliminates the post-operative recovery period thereby reducing pain of the animals. This proposal investigates and expands upon the utility of this embryo transfer device by establishing the optimal ages and mouse strains for embryo transfer (Aim 1), determining whether embryonic stem cells can be effectively and efficiently transferred with this device (Aim 2), determining whether the device is useful for artificial inseminations (Aim 3), directly visualizing the device in the uterus which may enable design improvements (Aim 4) and quantifying measures of stress relative to surgical procedures to support widespread adoption of this procedure in the research community (Aim 5). The primary project objective is to produce data that will prove the efficacy ofthis embryo transfer device and thereby enable researchers to replace surgical procedures with a non-surgical method that is equally effective. Proving that this non-surgical transfer method is effective under a range of conditions will expand its use overthis range of applications and thereby maximize its impact in enabling researchers to refine and reduce the number of surgical procedures performed in mice. PUBLIC HEALTH RELEVANCE: Animal use in research is essential for biomedical research and with the developing abilities to genetically engineer mice for research; these animals are becoming even more important as they can be modified to mimic most human diseases. Genetic engineering of mice requires, as one of many other things, the ability to transfer embryos into mice where they complete their development. This project supports further development of a device that enables these embryo transfers to be performed without surgery and therefore greatly reduces the pain category that these research animals experience and thereby supports progress in biomedicine.

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