Bartlett B.J.,San Diego State University |
Bartlett B.J.,Expression Drug Designs, Llc |
Isakson P.,University of Oslo |
Lewerenz J.,Salk Institute for Biological Studies |
And 11 more authors.
Autophagy | Year: 2011
Suppression of macroautophagy, due to mutations or through processes linked to aging, results in the accumulation of cytoplasmic substrates that are normally eliminated by the pathway. This is a significant problem in long-lived cells like neurons, where pathway defects can result in the accumulation of aggregates containing ubiquitinated proteins. The p62/Ref(2)P family of proteins is involved in the autophagic clearance of cytoplasmic protein bodies or sequestosomes. These unique structures are closely associated with protein inclusions containing ubiquitin as well as key components of the autophagy pathway. In this study we show that detergent fractionation followed by western blot analysis of insoluble ubiquitinated proteins (IUP), mammalian p62 and its Drosophila homologue, Ref(2)P can be used to quantitatively assess the activity level of aggregate clearance (aggrephagy) in complex tissues. Using this technique we show that genetic or age-dependent changes that modify the long-term enhancement or suppression of aggrephagy can be identified. Moreover, using the Drosophila model system this method can be used to establish autophagy-dependent protein clearance profiles that are occurring under a wide range of physiological conditions including developmental, fasting and altered metabolic pathways. This technique can also be used to examine proteopathies that are associated with human disorders such as frontotemporal dementia, Huntington and Alzheimer disease. Our findings indicate that measuring IUP profiles together with an assessment of p62/Ref(2)P proteins can be used as a screening or diagnostic tool to characterize genetic and age-dependent factors that alter the long-term function of autophagy and the clearance of protein aggregates occurring within complex tissues and cells. © 2011 Landes Bioscience.
Herr D.R.,Expression Drug Designs, Llc |
Herr D.R.,San Diego State University
International Review of Cell and Molecular Biology | Year: 2012
The therapeutic use of monoclonal antibodies (mAbs) is the fastest growing area of pharmaceutical development and has enjoyed significant clinical success since approval of the first mAb drug in1984. However, despite significant effort, there are still no approved therapeutic mAbs directed against the largest and most attractive family of drug targets: G protein-coupled receptors (GPCRs). GPCRs regulate essentially all cellular processes, including those that are fundamental to cancer pathology, such as proliferation, survival/drug resistance, migration, differentiation, tissue invasion, and angiogenesis. Many different GPCR isoforms are enhanced or dysregulated in multiple tumor types, and several GPCRs have known oncogenic activity. With approximately 350 distinct GPCRs in the genome, these receptors provide a rich landscape for the design of effective, targeted therapies for cancer, a uniquely heterogeneous disease family. While the generation of selective, efficacious mAbs has been problematic for these structurally complex integral membrane proteins, progress in the development of immunotherapeutics has been made by several independent groups. This chapter provides an overview of the roles of GPCRs in cancer and describes the current state of the art of GPCR-targeted mAb drugs. © 2012 Elsevier Inc.
Harris G.L.,Expression Drug Designs, Llc |
Harris G.L.,San Diego State University |
Creason M.B.,Expression Drug Designs, Llc |
Brulte G.B.,Expression Drug Designs, Llc |
And 2 more authors.
PLoS ONE | Year: 2012
Background: S1P 3 is a lipid-activated G protein-couple receptor (GPCR) that has been implicated in the pathological processes of a number of diseases, including sepsis and cancer. Currently, there are no available high-affinity, subtypeselective drug compounds that can block activation of S1P 3. We have developed a monoclonal antibody (7H9) that specifically recognizes S1P 3 and acts as a functional antagonist. Methodology/Principal Findings: Specific binding of 7H9 was demonstrated by immunocytochemistry using cells that over-express individual members of the S1P receptor family. We show, in vitro, that 7H9 can inhibit the activation of S1P 3-mediated cellular processes, including arrestin translocation, receptor internalization, adenylate cyclase inhibiton, and calcium mobilization. We also demonstrate that 7H9 blocks activation of S1P 3 in vivo, 1) by preventing lethality due to systemic inflammation, and 2) by altering the progression of breast tumor xenografts. Conclusions/Significance: We have developed the first-reported monoclonal antibody that selectively recognizes a lipidactivated GPCR and blocks functional activity. In addition to serving as a lead drug compound for the treatment of sepsis and breast cancer, it also provides proof of concept for the generation of novel GPCR-specific therapeutic antibodies. © 2012 Harris et al.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 167.05K | Year: 2011
DESCRIPTION (provided by applicant): G protein-coupled receptors (GPCRs) represent a diverse family of cell surface receptors that mediate important biological responses in nearly all cells. These responses include proliferation, migration, tissue invasion, and cell survival. As such, this large, ~700 member family represents the most attractive single family of drug targets for a variety of diseases including cancer. A great deal of work has been performed in an effort to generate specific pharmacologic antagonists for individual family members, but the success of this approach has been limited by their structural similarity. This has made it difficult to produce suitably selective compounds that are not complicated by off-target effects. One approach thatoffers the potential for unparalleled specificity is the development of monoclonal antibodies. However, GPCRs have historically been considered intractable to antibody antagonism due to poor antigenicity of critical, exposed, extracellular motifs that must be targeted to block receptor activation. Breast cancer is a physically and emotionally devastating diagnosis affecting over 2.3 million Americans living with the disease and killing over 100 women each day. Although the prognosis for this disease is gradually improving with the continued development of antineoplastic drugs, hormonal therapies, and targeted therapies, many aggressive forms of breast cancer are resistant to chemotherapy and result in a 10% mortality rate within 5 years of diagnosis. A compound known as sphingosine 1-phosphate (S1P) may be a major determinant of the aggressiveness and drug resistance of breast cancer. S1P is a small molecule normally present in high concentrations in the blood that accelerates the progression of breast cancer. It does this by promoting the growth and spreading of cancer cells and by stimulating the formation of new blood vessels, thereby increasing the supply of oxygen and nutrients to the tumor. Evidence suggests that these actions are largely the result of the stimulation of a cognate GPCR for S1P called S1P3. Since S1P has been shown to promote growth of breast cancer cells, and since it causes blood vessels to grow uncontrollably in tumors, it is likely that blocking S1P3 will inhibit the growth of mostforms of breast cancer. Animal studies suggest that loss of this receptor is not associated with undesirable effects, providing evidence for the safety of this approach. Until recently, however, there were no reports of any specific antagonists for S1P3. Our previous work (1R43CA132400) resulted in the development of a monoclonal antibody that specifically recognizes S1P3 and blocks its activation. Since this is the first-reported antibody to block a non-cytokine GPCR, it represents a breakthrough in antibody drug development. The goals of this project are to 1) quantitatively validate the functional efficacy of this antibody, and 2) demonstrate its activity and bioavailability in vivo. PUBLIC HEALTH RELEVANCE: Breast cancer is the second most common form of cancer in women causing the death of over 35,000 Americans each year. The proposed research will characterize a new drug developed at Expression Drug Designs that interrupts cellular processes known to promote growth of breast tumors, thus limiting cancer growth. Completion of the proposed project will determine if this new drug is likely to be effective in treating breast tumors.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase II | Award Amount: 1.26M | Year: 2011
DESCRIPTION (provided by applicant): Defining the molecular mechanism that leads to cellular aging and neural degeneration has proven to be difficult. Many types of damage are thought to contribute to senescence and neural degeneration and include mitochondrial and nuclear DNA mutations, protein misfolding, aggregate formation, reactive oxygen species (ROS), and stem cell senescence. However, a consensus has not yet developed as to which mechanism plays a causal role in aging. Indeed, each tissue may have aselect set of processes, or Achilles' heel that further exacerbated cellular decline. In the nervous system there is an agreement that mitochondrial senescence, the accumulation of ROS-dependent damage and the formation of protein aggregates or inclusion containing ubiquitin are involved with the most human neurological disorders, such as Alzheimer's and Parkinson's disease. There is a growing understanding that the autophagy pathway is involved with maintaining the mature nervous system by facilitatingthe removal of cellular damage and protein aggregates. The pathway is highly conserved and we found that expression profiles of autophagy genes show a significant decrease in the aging Drosophila CNS. At the same time, markers of cellular damage and aggregates, such as insoluble ubiquitinated aggregates (IUP), show a dramatic increase. Genetic analysis also shows that mutations in key genes significantly shorten adult lifespans (35 to 60%) and cause progressive neural defects that share striking similarities to those seen with Alzheimer's and other neurodegenerative disorders. Of greater significance is our observation that enhancing autophagy in the aging nervous system suppresses the accumulation of cellular damage (IUP) and significantly extends adult life spans. This work shows that examining factors that promote healthy neuronal aging can be done using Drosophila as a model system. In this proposal we take advantage of the conserved regulation of autophagy to identify neural protective compounds that enhance the pathway, promote longevity and neural function. This project involves several validated and optimized assays proposed in our original Phase-I application that were designed to identify compounds that enhanced autophagy, suppressed aggregate formation and extend life spans. In Specific Aim 1 an additional assay, which assesses the ability of different treatments to suppress oxidative stress was included. Specific Aim 2 represents an expansion of our drug-testing platform to assess the effectivenessof different compounds to promote neuronal health and function by examining their effect on several adult behaviors that show an age-dependent decline. Specific Aim 3 takes advantage of the conserved regulation of autophgy and other key protective pathwaysto identify those neural protective compounds that alter gene expression profiles in the aging nervous system. The goal of this proposal is to better understand the role of clearance pathways on aging and to develop in vivo assays that identify drugs thatcould be used for the treatment of human neurological disorders. PUBLIC HEALTH RELEVANCE: Presently there is no effective treatment for Alzheimer's disease and other age-related disorders that affect millions worldwide. Developing an effective method for validating neural protective compounds would streamline the development of life saving therapies. The research outlined in this proposal used Drosophila to develop several medium-throughput in vivo screening techniques that can identify novel therapeutic compounds for the treatment of aging disorders.
Expression Drug Designs, Llc | Date: 2012-05-14
Materials and Method for treating cancer and screening for anti-neoplastic agents are provided. These materials and methods can include sphingosine 1-phosphate antagonists that bind to sphingosine-1 phosphate receptor subtype 3. Antibodies and aptamers that selectively bind to an epitope in the extracellular loop between transmembrane domains two and three of sphingosine-1-phosphate receptor subtype 3 are provided.
Expression Drug Designs, Llc | Entity website
Expression Drug Designs, Llc | Entity website
Expression Drug Designs was founded in San Diego in 2004 by scientists from The Scripps Research Institute and San Diego State University. We are a biologics-based drug discovery company pursuing the development of therapeutic monoclonal antibodies and aptamers ...
PubMed | Expression Drug Designs, Llc
Type: | Journal: International review of cell and molecular biology | Year: 2012
The therapeutic use of monoclonal antibodies (mAbs) is the fastest growing area of pharmaceutical development and has enjoyed significant clinical success since approval of the first mAb drug in1984. However, despite significant effort, there are still no approved therapeutic mAbs directed against the largest and most attractive family of drug targets: G protein-coupled receptors (GPCRs). GPCRs regulate essentially all cellular processes, including those that are fundamental to cancer pathology, such as proliferation, survival/drug resistance, migration, differentiation, tissue invasion, and angiogenesis. Many different GPCR isoforms are enhanced or dysregulated in multiple tumor types, and several GPCRs have known oncogenic activity. With approximately 350 distinct GPCRs in the genome, these receptors provide a rich landscape for the design of effective, targeted therapies for cancer, a uniquely heterogeneous disease family. While the generation of selective, efficacious mAbs has been problematic for these structurally complex integral membrane proteins, progress in the development of immunotherapeutics has been made by several independent groups. This chapter provides an overview of the roles of GPCRs in cancer and describes the current state of the art of GPCR-targeted mAb drugs.