Recombinetics, Inc. | Date: 2016-09-20
Disclosed herein are genomically modified livestock animals and methods to provide them that express the SLICK phenotype. The animals disclosed herein express a truncated allele for the prolactin receptor (PRLR) gene. When expressed, the livestock animals produce a PRLR that is missing up to the terminal 148 amino acid (aa) residues of the protein all ranges and values within the explicitly stated range are contemplated: e.g., from 148 to 69. Animals expressing SLICK have superior thermoregulatory ability compared to non-slick animals and experience a less drastic depression in milk yield during the summer.
Recombinetics, Inc. | Date: 2016-08-31
Methods and kits to determine the presence of exogenous alleles within a native haplotype are provided. Introduction of foreign alleles into livestock genomes has provided the ability to introduce specific desirable traits. The present disclosure provides methods to identify the presence of exogenous alleles that foreign to a haplotype at a target locus, and identify specific markers that are native to the haplotype. Identification of exogenous genes at a target locus, flanked by native markers is indicative that the exogenous gene is present through molecular engineering. Conversely, the presence of an exogenous gene that are only partially flanked by native markers is indicative that the allele is present due to sexual breeding.
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 376.08K | Year: 2015
DESCRIPTION provided by applicant Immunodeficient rodents has revolutionized the study of regenerative medicine and cancer to date no analogous large animals with T B and NK cell deficiency are available Weandapos ve used our novel multiplex gene editing platform to knockout each allele of two genes Recombination Activating Gene RAG and Interleukin Receptor Gamma IL Rg to establish RG KO cell lines This will dramatically reducing the time and cost associated with breeding individual genetic mutations into pigs In addition we propose to restore the immune systems in RG KO pig by blastocyst complementation and cord blood transplantation The restored immune function in these RG KO swine will enable breeding animals to be maintained in standard housing with little risk of illness or death due to infection In addition breeding between chimeric immune restored RG KOandapos s has nearly a fold advantage over intercross of between heterozygous animals This innovative method for establishment and breeding of immunodeficient provides a unique sustainable supply of animals for research in regenerative medicine xenogenic organ tissue production and cancer using a large animal which is anatomically and physiologically similar to humans This project will also establish a platform for the creation of a pig with a human immune system that would have profound impacts on the development and testing of novel therapeutics transplantation rejection studies and vaccine development PUBLIC HEALTH RELEVANCE This SBIR proposes to develop an immunodeficient pig which could be propagated in standard housing with little risk of infection This novel model would allow the study of cancer regenerative medicine and xenogeneic organ tissue production Additionally this animal would serve as a platform to make a humanized pig in which the animal would have the components of a human immune system and would allow for further studies of therapeutics transplantation rejection and vaccine development
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 301.10K | Year: 2015
DESCRIPTION provided by applicant Organ transplantation is routinely used to replace entire organs in patients with a variety of health issues yet there are currently more than patients waiting to receive transplants due to lack of donors and this number is rising dramatically Exogenic organ production has the potential to alleviate the lack of available organs and help patients who are in desperate need In this proposal we aim to develop a facile platform upon which human organs can be grown in a pig and demonstrate the feasibility of this by generating pig pig chimeras in which the pancreas and its associated vasculature are entirely derived from donor pig cells We will be using blastocyst complementation which has been demonstrated to work in both mouse and pig and here we will address some major issues to make this technique clinically applicable First we will assess various pluripotent cells that coud be used for donor cells to overcome the availability cost and feasibility of using pig or human ES cells and allow us to query a number of developmental genes for exogenic organ production in the pig Second we will employ Recombineticsandapos innovative and proprietary livestock gene editing techniques to quickly and easily ablate the function of multiple genes allowing the production of chimeric animals with multiple cell lineages being donor derived Specifically two genes that are necessary for the formation of the pancreas and pancreatic vasculature will be knocked out allowing for the formation of an exogenic pancreas in pigs harboring vessels that are all solely donor derived This platform will accelerate the field of exogenic organ production and meets several needs identified in the NIH Symposium for Improving Animal Models for Regenerative Medicine In follow on Phase II studies we will evaluate organogenesis in multi lineage ablated hosts after complementation with human pluripotent cells PUBLIC HEALTH RELEVANCE This SBIR aims to develop a platform upon which human organs could be grown in a pig and used for transplantation into patients This study will allow us to develop rapid and facile techniques for investigating many different developmental genes that could be ablated for the production of many different organs and tissues Here we will develop a pig model in which the pancreas and pancreatic vasculature are completely derived from the cells of a donor pig The successful completion of this study will allow us to begin human organ production in pigs which could then be applied clinically
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 358.34K | Year: 2016
PROJECT SUMMARY Alzheimer s disease AD affects million people worldwide costing $ billion yearly One in nine Americans over the age of have AD the th leading cause of death in the US Annually $ billion is spent to care for the million Americans with AD AD is the only cause of death in the top that cannot be prevented cured or slowed and without the development of treatments for AD it is expected that by there will be million Americans with AD costing $ trillion yearly Life expectancy following AD diagnosis is only years AD patients suffer from chronic neurodegeneration gradual loss of bodily functions and a poor quality of life Studying patients with early onset AD has elucidated many genes that play a role in AD including the critical role of the Amyloid Precursor Protein gene hAPP and the Microtubule Associated Protein Tau gene hMAPT To date models of AD have been largely developed in mice and no therapies developed in these models have proven effective in humans likely due to the large anatomical and physiological variation between human and rodent brains coupled with the failure of mice to develop the neurodegenerative processes and brain lesions seen in AD patients Compared to rodents swine have much greater genetic anatomic and physiological similarity to humans offering an opportunity to model a complex neuropathological disease in a large animal The large gyrencephalic brain of the pig is similar in size and structure to humans and a model of diffuse brain injury in swine shows AD like pathology with accumulation of amyloid beta A and Tau We propose to establish a swine model of AD allowing researchers to understand the biology of AD enable doctors to establish methods of early detection and ultimately lead to the identification of new therapies to prevent halt the progression of and reverse AD We will establish this model using our powerful and proprietary genetic engineering techniques to replace swine APP and MAPT genes with human AD associated alleles of these genes This will allow us to precisely model human AD in swine and develop therapeutics that could be pre clinically tested for safety and efficacy in our swine and moved directly to human clinical trials We will assess the resulting animals at months of age by magnetic resonance imaging to determine if they have suffered neurodegeneration by whole brain and hippocampal volume measurement as well as if amyloid plaques have formed or they display a pattern of abnormal brain metabolite concentrations consistent with AD patients PROJECT NARRATIVE Alzheimer s disease AD affects million people worldwide costing $ billion yearly and one in nine Americans over the age of have AD the th leading cause of death in the US Annually $ billion is spent to care for the million Americans with AD but without the development of treatments for AD it is expected that by there will be million Americans with AD costing $ trillion yearly Because animal models of AD have failed to reproduce the biology of the disease and no treatments that have been able to stop slow or prevent the disease have been developed using these models we propose to establish a swine model of AD allowing researchers to understand the biology of AD enable doctors to establish methods of early detection and ultimately lead to the identification of new therapies to prevent halt the progression of and reverse AD
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 347.89K | Year: 2016
PROJECT SUMMARY Neurofibromatosis type NF is a genetic disorder associated with the development of nervous system tumors including vestibular schwannomas meningiomas cranial nerve tumors and spinal tumors due to germline loss of one copy of the NF gene While the prevalence of NF syndrome is only NF is commonly lost sporadically and up to people will develop a tumor with an underlying NF mutation therefore developing therapies that target NF mutant tumors is critical for NF patients and the general population The current standard of care for NF patients is surgical resection of the tumors although surgery is often not feasible and there is a high risk of hearing loss facial weakness and dysphagia Radiation therapy has been utilized in NF but can be associated with chronic neurologic dysfunction and or malignant transformation The goal of this proposal is to establish a swine model of NF that recapitulates the disease seen in NF patients to better understand disease etiology and progression and provide a reliable preclinical model for establishing safety and efficacy of new therapies prior to clinical trials A common human NF disease allele will be engineered into the genome of swine fibroblasts using site specific nucleases and those fibroblasts will undergo somatic cell nuclear transfer to generate pigs that harbor the human mutant NF allele At months of age these NF pigs will be evaluated by MRI for the presence of brain tumors and by brain auditory evoked response and compared to control animals to determine if they have NF related hearing deficits often seen in NF patients This model would allow the field to overcome two major hurdles in NF research First the mouse models of NF and NF related tumors do not fully recapitulate the disease seen in NF patients and have been poor predictors of clinical efficacy Second due to a small patient population and orphan disease status the ability to recruit enough patients for clinical trials is nearly impossible A large animal model that could serve as a preclinical platform for drug safety toxicology and efficacy would dramatically progress the development of NF therapeutics and candidate drug prioritization for a patient population that is too small to recruit enough patients for many clinical trials Further the NF mutation that was engineered in our swine model is a premature termination codon in exon of the NF gene allowing our model to serve as a platform for testing premature termination codon suppression therapies which are applicable to NF but also in the one third of genetic disorders characterized by premature termination codons including cystic fibrosis and Duchenne muscular dystrophy PROJECT NARRATIVE This SBIR aims to develop a swine model of Neurofibromatosis type NF a genetic disorder associated with a high risk of cancer and the development of nervous system tumors and a desperate need for better treatments and disease management While NF is a rare genetic disorder up to people will develop a tumor with the NF gene mutated in their lifetime and therapeutics aimed at treating NF patients may also be applicable for NF mutant tumors The goal of this application is to concentrate on research efforts that will significantly contribute to proving the scientific and technical feasibility of establishing a swine model of NF that can be applied to better understand NF etiology disease development and progression the application of novel imaging and monitoring techniques and the identification and testing of new therapies
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 394.73K | Year: 2016
DESCRIPTION provided by applicant Microvillus inclusion disease MVID is the most severe cause of Congenital Diarrheal Diseases in neonates MVID results from mutations that lead to malabsorption and life threatening intractable secretory diarrhea To date no curative therapy exists MVID patients are largely dependent on parenteral nutrition Prognosis is generally poor due to metabolic decompensation dehydration infections and liver complications associated with parenteral nutrition The only alternative therapy to parenteral nutrition is intestinal transplantation MVID accounts for of pediatric bowel transplantation worldwide However the overall five year survival after small bowel transplantation is only about Parenteral nutrition and bowel transplants are non permanent solutions for treating MVID further work in deciphering how inactivating mutations in MYO B lead to aberrant trafficking in enterocytes will provide novel insights into genotype phenotype relations and pave the way for development of improved diagnosis and viable alternative therapeutic strategies There is a limited availability of patient material and no suitable animal models for MVID hampering the thorough understanding of the diseaseandapos s molecular mechanisms While we have recently been able to develop a mouse model of germline and intestinally targeted deletion of MYO B these mice die in their first week of life so any analysis of interventions that might alter the course f disease are not possible due to the small size To that end we propose utilizing our state of the art gene editing platform to develop swine with a specific mutation P L corresponding to the human P L mutation of the MYO B gene present in Navajo MVID patients We hypothesize that introducing a Proline P to Leucine L mutation in pigs at a site analogous to the human P L allele can induce MVID in piglets Execution of the hypothesis can be accomplished with the following specific aim develop and evaluate the MYO B P L mutant pigs as models of human MVID In addition to state of the art gene editing platform with expertise of Drs Melkamu Veterinary Physiology and Carlson Animal biotechnology from Recombinetics we have engaged a world renown expert in MVID Dr James R Goldenring MD PhD a gastroenterologist and a Professor of Experimental Surgery from Vanderbilt University School of Medicine A reliable large animal model of MVID will have tremendous impact on industry and academic research to develop and test new drugs and novel therapeutic approaches to treat this awful disease PUBLIC HEALTH RELEVANCE This SBIR aims to develop a swine model of microvillus inclusion disease MVID the most severe cause of Congenital Diarrheal Diseases in neonates We use gene editing to mimic the most common and severe allele corresponding to the human P L mutation of the MYO B gene present in Navajo MVID patients There is still no cure for MVID besides parenteral nutrition and bowel transplants without which all patientsandapos progress into severe dehydration and death We propose that this unique large animal model will mimic the human condition and accelerate translation of novel therapies into the clinic
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 355.20K | Year: 2016
DESCRIPTION provided by applicant Phenylketonuria PKU is one of the most common inborn errors of metabolism PKU commonly detected in infants during newborn screening is due to recessively inherited phenylalanine hydroxylase PAH deficiency Current treatment strategies rely on restriction of dietary phenylalanine Phe intake to prevent the major manifestations of the disease however dietary therapy is complicated unpalatable and must be sustained throughout life to prevent neurocognitive disability associated with chronic hyperphenylalaninemia Here we propose to generate a large animal swine model of PKU to evaluate novel treatments strategies that are difficult to evaluate in murine PKU models due to their small size and physiological and anatomical differences These novel therapies including enzyme substitution therapy hepatocyte transplantation and gene therapy liver directed and muscle directed gene therapy strategies employing both recombinant adeno associated virus rAAV and non viral naked DNA vector systems could be applied to both PKU and many other inherited genetic diseases A large animal model to evaluate the safety and efficacy of these therapies is critical to bridge the gap between laboratory experiments and the treatment of patients in the clinic Additionally a swine model of PKU is critical to understanding the molecular mechanisms through which hyperphenylalaninemia causes harm to the brain and results in physical neurological and behavioral deficits seen in patients Further a model in which physical and molecular phenotypes can be evaluated during Phe restricted diet treatment following discontinuation of Phe restricted diet and during the implementation of novel treatments such as gene therapy will be critical for better treating patients with PKU The preclinical experiments we propose to conduct will be vital to understanding the limiting issues in treating patients and will have direct influence upon the design of future human clinical trials PUBLIC HEALTH RELEVANCE This SBIR aims to develop a swine model of Phenylketonuria PKU a commonly inherited genetic disease affecting in children born in the US Left untreated PKU is associated with growth failure acquired microcephaly hypopigmentation and severe developmental disability While dietary phenylalanine restriction prevent the major manifestations of the disease this dietary restriction is complicated unpalatable and must be sustained throughout life The physical behavioral and neurological symptoms that PKU patients often experience are poorly understood and several academic and industry researchers are seeking new treatment modalities to treat PKU Several successful gene or cell therapies have been developed in mice that have yet to enter clinical trials in patients due to the immense size differential having impact on therapeutic delivery and dosage parameters Pigs are comparable in size and anatomy to humans enabling optimization of treatment modalities prior to clinical trials Further lessons learned from a PKU pig model would be directly applicable to the development of novel treatments for related inborn errors of metabolism
Recombinetics, Inc. | Date: 2015-04-28
Materials and methods for making multiplex gene edits in cells or embryos are presented. Further methods include animals and methods of making the same. Methods of making chimeric animals are presented, as well as chimeric animals.
Recombinetics, Inc. | Date: 2016-03-30
Swine animal models comprising a genomic disruption of an endogenous gene chosen from the group consisting of a Low-Density Lipoprotein Receptor gene LDLR, Duchenes Muscular Dystrophy (DMD) gene, and hairless gene (HR). Methods of preparing transfected cells useful for making a transgenic animal comprising exposing a first group of cells to a transfection agent and reseeding the group with additional cells that have not been exposed to the agent. The transgenic animals are useful for medical and scientific animal models of human diseases and conditions, as well as sources for cells, tissues, and biomaterials.