Davidson E.,Integral Molecular |
Doranz B.J.,Integral Molecular
Immunology | Year: 2014
Characterizing the binding sites of monoclonal antibodies (mAbs) on protein targets, their 'epitopes', can aid in the discovery and development of new therapeutics, diagnostics and vaccines. However, the speed of epitope mapping techniques has not kept pace with the increasingly large numbers of mAbs being isolated. Obtaining detailed epitope maps for functionally relevant antibodies can be challenging, particularly for conformational epitopes on structurally complex proteins. To enable rapid epitope mapping, we developed a high-throughput strategy, shotgun mutagenesis, that enables the identification of both linear and conformational epitopes in a fraction of the time required by conventional approaches. Shotgun mutagenesis epitope mapping is based on large-scale mutagenesis and rapid cellular testing of natively folded proteins. Hundreds of mutant plasmids are individually cloned, arrayed in 384-well microplates, expressed within human cells, and tested for mAb reactivity. Residues are identified as a component of a mAb epitope if their mutation (e.g. to alanine) does not support candidate mAb binding but does support that of other conformational mAbs or allows full protein function. Shotgun mutagenesis is particularly suited for studying structurally complex proteins because targets are expressed in their native form directly within human cells. Shotgun mutagenesis has been used to delineate hundreds of epitopes on a variety of proteins, including G protein-coupled receptor and viral envelope proteins. The epitopes mapped on dengue virus prM/E represent one of the largest collections of epitope information for any viral protein, and results are being used to design better vaccines and drugs. © 2014 John Wiley & Sons Ltd.
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase I | Award Amount: 224.75K | Year: 2015
DESCRIPTION provided by applicant Membrane proteins control the flow of information nutrients and signals through the cell membrane and are the targets for more than of FDA approved drugs Monoclonal antibodies MAbs that target membrane proteins can be exceptionally useful in research diagnostic and therapeutic applications but for most membrane proteins there are no MAbs that recognize the native protein on the cell surface The need for such MAbs has been recognized by industry and the NIH but efforts to identify such MAbs are limited by the difficulty in expressing and purifying membrane proteins in exogenous systems and by conventional MAb isolation strategies that typically focus on one target at a time A novel approach to identify membrane protein MAbs in a high throughput manner is needed to derive MAbs against the entire human membrane proteome Here we propose a platform technology that can be used to rapidly isolate MAbs against structurally intact membrane proteins for therapeutic development diagnostics and biomedical research PUBLIC HEALTH RELEVANCE This proposal will contribute to public health and the cure of human disease by identifying MAbs against native membrane proteins enabling their development for therapeutics diagnostics and biomedical research
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 660.81K | Year: 2011
DESCRIPTION (provided by applicant): Because of their importance in cell signaling and human disease, integral membrane proteins such as G protein-coupled receptors (GPCRs) and ion channels comprise over 40% of existing drug targets. Monoclonal antibodies(MAbs) that recognize conformation-dependent epitopes on membrane proteins are usually the most valuable type of antibody because they often bind to critical structures of the receptor that can be exploited for its detection or inhibition. However, the development of conformation-dependent, inhibitory MAbs against membrane proteins is especially difficult because, unlike soluble proteins, most membrane proteins are dependent on a lipid environment to maintain their native tertiary and quaternary (oligomeric) structures. Membrane proteins are often difficult to purify, often express at low concentrations on the cell surface, and are usually poorly represented by linear peptides. New approaches are needed to develop such MAbs for therapeutic, diagnostic, and research applications. Here we propose to use a novel technology, the Lipoparticle, to capture and concentrate structurally intact membrane proteins in a format amenable to immunization. The concept of using Lipoparticles to develop antibodies against cellular membrane proteins builds on the historic use of viral particles as successful vaccines. PUBLIC HEALTH RELEVANCE: This proposal will result in monoclonal antibodies against important membrane protein targets for therapeutic development, diagnostics, and biomedical research. Lipoparticles optimized as immunogens will be developed as commercial products.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 242.38K | Year: 2011
DESCRIPTION (provided by applicant): The identification of highly potent and broadly neutralizing antibodies isolated from HIV-1- infected donors suggests that when presented with a suitable antigenic structure, the human immune system can produce protective antibodies that HIV-1 is unable to evade. Understanding these antigenic structures is a primary objective in developing a vaccine capable of generating broadly protective humoral immunity. Most of these broadly neutralizing MAbs bind conformationally complex epitopes on Envelope that have been difficult to epitope map, and an increasing number of such MAbs are being identified using more efficient approaches to MAb isolation. However, the ability to characterize MAbs has not kept up, leaving a major gapbetween the growing ability to isolate relevant MAbs and the ability to molecularly define the immunogenic structures that gave rise to them. The goal of this proposal is to develop tools to rapidly and comprehensively map MAb epitopes on structurally complex viral Envelope proteins. Identifying the epitopes of potent and broadly neutralizing MAbs will enable Env variants exhibiting improved antigenic characteristics to be more rapidly designed and tested as vaccine candidates. PUBLIC HEALTH RELEVANCE: This project will contribute to human health by identifying the epitopes of potent and broadly neutralizing MAbs so that Env variants exhibiting improved antigenic characteristics can be more rapidly designed and tested as vaccine candidates.
Integral Molecular | Date: 2013-11-01
The present invention relates to lipoparticles. The invention also relates to producing lipoparticles. The invention further relates to lipoparticles comprising a viral structural protein. The invention further relates to a lipoparticle comprising a membrane protein, and the lipoparticle can be attached to a sensor surface. The invention further relates to methods of producing and using the lipoparticle to, inter alia, assess protein binding interactions.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 546.30K | Year: 2012
DESCRIPTION (provided by applicant): When presented with a suitable antigenic Envelope (Env) structure, the human immune system can produce protective antibodies that HIV is unable to evade. Many of the most potently neutralizing HIV antibodies identifiedto date preferentially bind the trimeric structure of Env. Thus, recapitulating the native trimeric structure of Env as it exists on the virus and cell surface is a primary objective in developing an immunogen capable of generating broadly protective humoral immunity. The development of a soluble, trimeric Env structure that recapitulates the native, pre-fusion structure of Env has become a major goal for HIV vaccine advancement. However, the inherent instability of the gp120 and gp41subunit complex has posed a major obstacle in designing a soluble Env trimer (gp140) capable of presenting an effective antigenic structure to the immune system. The goal of this project is to engineer a gp140 variant with disulfide bonds that stabilize the trimer to yield conformationally intact, trimeric gp140. To accomplish the Aims of this project, we will engineer and test up to 10,000 gp140 variants that have been systematically mutated to contain paired disulfide bonds designed to covalently stabilize the protein. Each mutant will be individually expressed in human cells to maintain native post-translational processing, and each mutant will be tested for retention of its pre-fusion conformation, antigenic integrity, and trimeric structure. PUBLIC HEALTH RELEVANCE:This project will contribute to human health by identifying improved gp140 immunogens as vaccine candidates. An improved gp140 immunogen that is stable as a pure, covalently associated trimer and that represents the native gp160 structure as it exists onvirions and cells in its pre-receptor binding, pre-fusion conformation is likely to elcit a potent and broadly neutralizing antibody response against HIV.
Agency: Department of Health and Human Services | Branch: National Institutes of Health | Program: SBIR | Phase: Phase II | Award Amount: 1.43M | Year: 2014
DESCRIPTION provided by applicant Acute and chronic pain resulting from injury surgery or disease afflicts andgt million Americans each year having a severe impact on mood mental health and quality of life and costing the U S approximately $ billion in economic costs annually For many patients treatment options provide inadequate relief because of the shortcomings of available therapeutics To date most treatments for pain have been small molecule compounds that block the activity of select ion channels or other pain receptors but these therapeutics often result in side effects caused by off target binding or suffer from poor bioavailability These limitations have prompted renewed searches for novel targets for the treatment of pain and novel types of inhibitors capable of achieving the specificity and bioavailability needed for a successful therapeutic The P X ion channel is a primary mediator of pain triggered by ATP release and drugs that target P X could be efficacious in treating chronic pain Here we propose to develop MAbs targeting the ion channel P X for the treatment of neuropathic and inflammatory pain PUBLIC HEALTH RELEVANCE This proposal will contribute to public health and the cure of human disease by resulting in the development of therapeutic MAbs against P X to treat neuropathic and inflammatory pain Lead MAbs with confirmed specificity and efficacy will be developed with the goal of entering pre clinical studies and the filing of an IND for studies in humans
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 214.21K | Year: 2012
DESCRIPTION (provided by applicant): We propose to use a novel technology, the Lipoparticle, to capture and concentrate structurally intact membrane proteins in a format amenable to phage panning for MAb isolation. The use of Lipoparticles to pan phage display libraries represents a valuable approach for obtaining antibodies against conserved membrane protein antigens because 1) Lipoparticles contain high concentrations of conformationally-intact target receptors, 2) target receptors within Lipoparticles are not exposed to adjuvants or biologically destructive environments so remain structurally intact, and 3) target receptors with high sequence conservation across species can be used as targets. PUBLIC HEALTH RELEVANCE: This proposal will result in monoclonal antibodies against important membrane protein targets for therapeutic development, diagnostics, and biomedical research. Lipoparticles optimized as panning reagents will be developed as a commercial product, and Lipoparticle-derived MAbs against biomedically important membrane protein targets will be licensed for therapeutic development or diagnostic use, or enter the commercial market as research reagents.
Integral Molecular | Date: 2013-01-15
The present invention relates to the use of lipoparticles, virus-like particles, and viruses. The present invention also relates to testing ion channel function and modulators of ion channels.
Integral Molecular and Blood Systems Research Institute | Date: 2014-07-21
Embodiments disclosed herein provide for antibodies, including neutralizing antibodies, against Chikungunya virus, uses thereof, and methods of identifying antibodies, including neutralizing antibodies, against Chikungunya virus. In some embodiments, antibodies that binds to a CHIKV antigen, wherein the antigen is the CHIKV E1, E2, E3 protein, or any heterocomplex thereof are provided. In some embodiments, the antibody is an isolated antibody, a neutralizing antibody, a recombinant antibody, or any combination thereof. In some embodiments, the antibodies described herein bind to an epitope of E2 Domain A, E2 Domain B, or E1 Domain II of the CHIKV antigen. In some embodiments, the antigen is E2 protein.