Recombinetics, Inc. | Date: 2017-03-08
Materials and methods for making multiplex gene edits in cells and are presented. Further methods include animals and methods of making the same.
Recombinetics, Inc. | Date: 2016-10-27
Human or humanized tissues and organs suitable for transplant are disclosed herein. Gene editing of a host animal provides a niche for complementation of the missing genetic information by donor stem cells. Editing of a host genome to knock out or disrupt genes responsible for the growth and/or differentiation of a target organ and injecting that animal at an embryo stage with donor stem cells to complement the missing genetic information for the growth and development of the organ. The result is a chimeric animal in which the complemented tissue (human/humanized organ) matches the genotype and phenotype of the donor. Such organs may be made in a single generation and the stem cell may be taken or generated from the patients own body. As disclosed herein, it is possible to do so by simultaneously editing multiple genes in a cell or embryo creating a niche for the complemented tissue. Multiple genes can be targeted for editing using targeted nucleases and homology directed repair (HDR) templates in vertebrate cells or embryos.
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.
Recombinetics, Inc. | Date: 2016-10-07
Disclosed herein are animals that are modified genetically to express one or more introduced sry alleles or to have a knockout of an existing sry allele. In various embodiments, the sry allele is inserted in at least one X chromosome of an XX resulting in an animal having two X chromosomes and appearing phenotypically male and sterile. In some embodiments the animal is modified to express multiple copies of sry inserted throughout its genome including the X chromosomes and the autosomes. In still other embodiments, the sry allele is inserted in multiple allosome sites. In these embodiments, XX individuals will be sterile and phenotypically male. XY individuals will be male and fertile but offspring will have an opportunity to inherit multiple sry alleles. In this embodiment, progeny of a male carrier will be sterile and phenotypically male if they are genotypically XX and express sry. Male progeny will be fertile and normal but will carry a hereditable copy of sry in their allosomes.
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: 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.