Our Technologies
We have developed an extensive set of integrated technologies to study mitochondria and their connections to diseases. Our technology platform facilitates the identification of gene and protein targets and markers associated with disease. This comprehensive and integrated platform provides the basis for us to discover and develop mitochondrial drugs, research tools and diagnostic products.
Mitochondrial Proteomics
We use a combination of proprietary and other advanced technologies to access and profile the mitochondrial proteome and to identify proteins with potential commercial importance as novel drug targets or diagnostic markers. Using our mitochondrial expertise and proprietary sample preparation techniques, we isolate highly purified mitochondria from cells and tissues and separate the mitochondria into sub-components. We then identify mitochondrial proteins in each fraction using a combination of customized gel electrophoresis, solid-phase affinity separation and mass spectrometry. We use these methodologies to analyze low abundance and difficult-to-obtain membrane proteins. Using engineered cell lines and other disease models, we perform differential protein expression experiments to study mitochondrial involvement in disease and to identify additional drug targets and diagnostic markers.
We have developed proprietary bioinformatics software that enables us to search public and private databases for genes and proteins related to mitochondrial function. This software leverages our knowledge of amino acid sequences that the cell uses for the trafficking of proteins to mitochondria and of sequences associated with particular protein functions. We also search our proprietary mitochondrial genome databases to identify changes that may be associated with disease. We analyze these genes and proteins to identify new drug targets or diagnostic markers. Once we define a gene or protein sequence of potential interest, we clone, express, purify and determine the function of the resulting protein and its relevance to disease.
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Mitochondrial Genomics
We use a combination of proprietary and other technologies to identify alterations in mitochondrial and nuclear genes that may contribute to mitochondrial dysfunction and disease. We collect blood and tissue samples from individuals with neurodegenerative and metabolic diseases and from healthy individuals of the same age. We then sequence and catalogue the entire mitochondrial genome from each individual. We also compare the sequences of individuals from the same geographical location or from the same ethnic group to identify SNPs that are specific to that population, or haplogroups. After common patterns of SNPs are identified, we are able to focus on the remaining unique polymorphisms or mutations that we believe are associated with disease. We have sequenced the complete mitochondrial genomes of over 900 individuals, which we believe represents the largest human database of this kind in the world. We have identified more than 550 haplogroup-related SNPs and more than 2,000 unique SNPs that we are examining for association with disease. We have identified significant associations between mitochondrial SNPs and major diseases, including Alzheimer's disease.
We also use a broad array of advanced gene expression technologies to identify genes that may be associated with disease. We accomplish this by comparing RNA levels in diseased and normal cells or tissues to identify genes that are over-expressed or under-expressed. From this work, we have filed patents directed to more than 50 distinct genes and proteins.
High-Throughput Mitochondrial Screening
To identify novel compounds that interact with our drug targets, we have designed a number of sophisticated high-throughput screens. These screens utilize a series of proprietary reagents to evaluate mitochondrial binding, displacement or inhibitory characteristics when a test compound is introduced. In addition to binding protocols, we currently employ a large number of assays that model parameters of mitochondrial function, such as membrane potential, calcium uptake, ATP levels and mitochondrial free radical generation. We have developed a high-throughput screen capable of detecting mitochondrial stability and function in intact cells.
High-Content Mitochondrial Assays
We have developed high-content assays using an advanced bioenergetics system for real-time, simultaneous measurement of mitochondrial respiration, swelling, membrane potential and ion flux. This can be done in the presence or absence of trigger molecules that activate or inhibit pathways of interest. This technology provides detailed data about how a potential drug affects the "health" and metabolic state of mitochondria, and about the function of novel genes and proteins. We use this assay system to study and optimize compounds identified using high-throughput screens, and we iteratively optimize the activity of lead compounds through structure-activity studies. Drug candidates with the desired level of activity are advanced to studies in engineered cell lines and animal disease models.
Disease-Relevant Mitochondrial Models
We have developed patented technologies that allow us to produce disease-relevant mitochondrial models. We produce human models of mitochondrial dysfunction in cybrid and other engineered cell lines. We create cybrid cells by fusing platelets with cells that have been treated to remove their mitochondrial DNA. Since platelets contain mitochondria but have no nucleus, the fusion process should transfer only mitochondrial DNA to the recipient cells. The
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phenotype, or biochemistry, of a cybrid is then assessed using a variety of proprietary and other technologies to determine if the engineered cell line exhibits characteristics of disease. When a cybrid displays the characteristics of a patient's disease, this suggests that there is a mitochondrial genetic link to the disease. Cybrids model some of the features of major disease phenotypes. Published data on Alzheimer's cybrids show various defects consistent with Alzheimer's disease, including metabolic changes and elevated levels of beta-amyloid, a protein associated with the disease.
Through our network of scientific collaborations, we have access to animal disease models that have been treated with mitochondrial toxins and that are useful in identifying drug candidates. Parkinson's disease-like abnormalities in animals have been induced with agents that block electron transport chain complex I. We also study naturally occurring animal disease models, such as a strain of guinea pigs that spontaneously develops osteoarthritis of the joints with aging and a strain of diabetic mice.
Combinatorial and Medicinal Chemistry
We have substantial chemistry resources for lead discovery and drug candidate optimization. We use our proprietary MultiPin, solid-phase parallel synthesis and other advanced technologies to rapidly develop combinatorial libraries of drug-like molecules and peptides for primary screening. In many instances, we have based our initial chemistry on natural products or marketed drugs that are known to bind to proteins that are similar to our targets. This is an effective approach to drug development because the efficacy and safety of these classes of molecules are often already well established in animal or human studies. In addition, we have medicinal chemistry capabilities that allow us to synthesize variations of leads to optimize their desirable properties. In-house chemistry resources give us greater confidentiality, better availability and quality control and tighter focus on potential mitochondrial drug leads, thereby improving our chances for successful and competitive lead development.
Mimotopes
Our wholly-owned subsidiary, Mimotopes, is a pioneer in combinatorial chemistry and a supplier of custom peptides. Mimotopes has developed a considerable range of proprietary technologies for peptide and small molecule non-peptide drug discovery research. Mimotopes' broad combinatorial and medicinal chemistry capabilities are a key part of MitoKor's integrated drug discovery platform. Mimotopes' chemists contribute to the design and synthesis of potential drug candidates and libraries for MitoKor's internal research programs. Mimotopes' peptide expertise has also contributed to MitoKor's assay design and target validation.
Mimotopes develops, markets and sells chemistry products and services internationally to biotechnology and pharmaceutical companies and academic and government laboratories. Mimotopes works with its customers to pursue drug discovery, genomics, proteomics, antibody, vaccine and other research. Mimotopes' expertise in polymer science and grafted plastics enables the design and manufacture of various solid-phase and affinity separation research tools. Mimotopes' products include SynPhase products for solid-phase combinatorial synthesis, PepSets peptide libraries for the mapping of epitopes, the parts of proteins that induce immune responses, and the custom synthesis of peptidomimetics and other compounds. |
