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Protein Advances, Inc.

SEATTLE, WA, United States
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Fox C.B.,Infectious Disease Research Institute | Baldwin S.L.,Infectious Disease Research Institute | Duthie M.S.,Infectious Disease Research Institute | Duthie M.S.,Protein Advances, Inc. | And 3 more authors.
AAPS PharmSciTech | Year: 2012

Egg phosphatidylcholine is commonly used as an emulsifier in formulations administered parenterally. However, synthetic phosphatidylcholine (PC) emulsifiers are now widely available and may be desirable substitutes for egg-derived phospholipids due to stability, purity, and material source considerations. In earlier work, we demonstrated that a squalene-1-palmitoyl-2- oleoyl-sn-glycero-3-phosphocholine (POPC) emulsion provided equivalent physical stability compared to a squalene-egg PC emulsion. In the present manuscript, we evaluate the physical stability of vaccine adjuvant emulsions containing a range of other synthetic phosphatidylcholine emulsifiers. Besides the POPC emulsion, the 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) emulsion showed good particle size and visual stability compared to emulsions made with other synthetic phospholipids. Moreover, comparable immune responses were elicited by squalene emulsions employing various synthetic PC or egg PC emulsifiers in combination with an inactivated influenza vaccine or a recombinant malaria antigen, and these responses were generally enhanced compared to antigen without adjuvant. Therefore, we show that (1) some synthetic PCs (DMPC, POPC, and to a lesser extent 1,2-dioleoyl-sn-glycero-3-phosphocholine) are effective stabilizers of squalene emulsion over a range of storage temperatures while others are not (1,2-distearoyl-sn-glycero-3-phosphocholine, 1,2-dipalmitoyl-sn- glycero-3-phosphocholine, and 1,2-dilauroyl-sn-glycero-3-phosphocholine) and (2) the immunogenicity of stable squalene emulsions is similar regardless of PC source. © 2012 American Association of Pharmaceutical Scientists.

Duthie M.S.,Infectious Disease Research Institute | Duthie M.S.,Protein Advances, Inc. | Hay M.N.,Protein Advances, Inc. | Rada E.M.,Central University of Venezuela | And 9 more authors.
European Journal of Clinical Microbiology and Infectious Diseases | Year: 2011

Although curable, leprosy requires better diagnostic and prognostic tools to accompany therapeutic strategies. We evaluated the serum samples of leprosy patients from Venezuela and Brazil for reactivity against the specific recombinant proteins, ML0405 and ML2331, and the LID-1 fusion protein that incorporates both of these antigens. Antigen-specific IgG was highest in lepromatous leprosy patients (LL) and decreased across the disease spectrum, such that only a small subset of true tuberculoid patients (TT) tested positive. The impact of multidrug therapy (MDT) on these antibody responses was also examined. Several years after treatment, the vast majority of Venezuelan patients did not possess circulating anti-LID-1, anti-ML0405, and anti-ML2331 IgG, and the seropositivity of the remaining cases could be attributed to irregular treatment. At discharge, the magnitude and proportion of positive responses of Brazilian patients against the proteins and phenolic glycolipid (PGL)-I were lower for most of the clinical forms. The monthly examination of IgG levels in LL patient sera after MDT initiation indicated that these responses are significantly reduced during treatment. Thus, responses against these antigens positively correlate with bacillary load, clinical forms, and operational classification at diagnosis. Our data indicate that these responses could be employed as an auxiliary tool for the assessment of treatment efficacy and disease relapse. © 2011 Springer-Verlag.

Fox C.B.,Infectious Disease Research Institute | Baldwin S.L.,Infectious Disease Research Institute | Duthie M.S.,Infectious Disease Research Institute | Duthie M.S.,Protein Advances, Inc. | And 3 more authors.
Vaccine | Year: 2011

Squalene-based oil-in-water emulsions have been used for years in some seasonal and pandemic influenza vaccines. However, concerns have been expressed regarding squalene source and potential biological activities. Little information is available regarding the immunomodulatory activity of squalene in comparison with other metabolizable oils in the context of oil-in-water emulsions formulated with vaccines. The present work describes the manufacture and physical characterization of emulsions composed of different classes of oils, including squalene, long chain triglycerides, a medium chain triglyceride, and a perfluorocarbon, all emulsified with egg phosphatidylcholine. Some differences were apparent among the non-squalene oils in terms of emulsion stability, including higher size polydispersity in the perfluorocarbon emulsion, more rapid visual instability at 60 °C for the long-chain triglyceride and perfluorocarbon emulsions, and an increased creaming rate in the medium-chain triglyceride emulsion at 60 °C as detected by laser scattering optical profiling. The biological activity of each of these emulsions was compared when formulated with either a recombinant malaria antigen or a split-virus inactivated influenza vaccine. Overall, vaccines containing the squalene emulsion elicited higher antibody titers and more abundant long-lived plasma cells than vaccines containing emulsions based on other oils. Since squalene-based emulsions show higher adjuvant potency compared to the other oils tested, non-squalene oils may be more suitable as carriers of amphiphilic or hydrophobic immunostimulatory molecules (such as TLR agonists) rather than as stand-alone adjuvants. © 2011 Elsevier Ltd.

Goto Y.,Obihiro University of Agriculture and Veterinary Medicine | Goto Y.,University of Tokyo | Duthie M.S.,Protein Advances, Inc. | Duthie M.S.,Infectious Disease Research Institute | And 6 more authors.
Parasitology International | Year: 2011

Serological diagnosis is a useful method to detect African trypanosome infection in livestock animals. Currently available serological tests utilize whole parasites or crude antigens, and recombinant antigens may improve reproducibility/standardization and reduce production costs. With a goal of identifying such recombinant proteins, we computationally identified proteins with tandem repeat (TR) domain from the parasite proteomes and evaluated their potential for serological diagnosis of African trypanosome infections in cattle. Among those tested, Tbg4 demonstrated the best performance with 92% sensitivity, followed by TbbGM6 (85%), TcoGM6 (85%), Tbg2 (65%) and Tbg5 (65%). Although further evaluations such as investigating cross-reactivity to other infections are needed, our data indicate the potential of these antigens for detection of African trypanosome infection in cattle. © 2011 Elsevier Ireland Ltd.

Goto Y.,University of Tokyo | Goto Y.,Obihiro University of Agriculture and Veterinary Medicine | Goto Y.,Protein Advances, Inc. | Duthie M.S.,Protein Advances, Inc. | And 5 more authors.
Biochemical and Biophysical Research Communications | Year: 2011

Trypanosoma brucei subspecies cause African trypanosomiasis in humans and animals. These parasites possess genes encoding proteins with large tandem repeat (TR) domains as do the other trypanosomatid parasites. We have previously demonstrated that TR protein of Leishmania infantum and Trypanosoma cruzi are often targets of B-cell responses. However, African trypanosomes are susceptible to antibody-mediated immunity, and it may be detrimental for the parasites to have such B-cell antigens on the cell surface. Here we show TR proteins of T. brucei subspecies are also antigenic: recombinant TR proteins of these parasites detect antibodies in sera from mice infected with the parasites by ELISA. Analysis of amino acid sequences revealed that, different from TR proteins of Leishmania species or T. cruzi, the presence of predicted signal peptides, trans-membrane domains and GPI anchor signals in T. brucei TR proteins are significantly lower than those of the whole proteome. Many of the T. brucei TR proteins are specific in the species or conserved only in the closely related species, as is the same case for Leishmania major and T. cruzi. These results suggest that, despite their sharing some common characteristics, such abundance in large TR domains and immunological dominance, TR genes have evolved independently among the trypanosomatid parasites. © 2011 Elsevier Inc.

Windish H.P.,Infectious Disease Research Institute | Duthie M.S.,Infectious Disease Research Institute | Duthie M.S.,Protein Advances, Inc. | Ireton G.,Infectious Disease Research Institute | And 5 more authors.
Vaccine | Year: 2011

Tuberculosis is a major health concern. Non-living tuberculosis (TB) vaccine candidates may not only be safer than the current vaccine (BCG) but could also be used to boost BCG to enhance or elongate protection. No subunit vaccines, however, are currently available for TB. To address this gap and to improve the global TB situation, we have generated a defined subunit vaccine by genetically fusing the genes of 3 potent protein Mtb antigens, Rv2875, Rv3478 and Rv1886, into a single product: ID87. When delivered with a TLR4 agonist-based adjuvant, GLA-SE, ID87 immunization reduced Mtb burden in the lungs of experimentally infected mice. The reduction in bacterial burden of ID87/GLA-SE immunized mice was accompanied by an early and significant leukocyte infiltration into the lungs during the infectious process. ID87/GLA-SE appears to be a promising new vaccine candidate that warrants further development. © 2011 Elsevier Ltd.

Goto Y.,Obihiro University of Agriculture and Veterinary Medicine | Goto Y.,Infectious Disease Research Institute | Goto Y.,Protein Advances, Inc. | Carter D.,Infectious Disease Research Institute | And 5 more authors.
Infection and Immunity | Year: 2010

Proteins with tandem repeat (TR) domains have been found in various protozoan parasites, and they are often targets of B-cell responses. Through systematic analyses of whole proteomes, we recently demonstrated that two trypanosomatid parasites, Leishmania infantum and Trypanosoma cruzi, are rich in antigenic proteins with large TR domains. However, the reason that these proteins are antigenic was unclear. Here, by performing molecular, immunological, and bioinformatic characterizations of Leishmania TR proteins, we found two possible factors affecting the antigenicity of these proteins; one factor is their fundamental composition as TR proteins, and the other is regulation of their expression by parasites. Enzyme-linked immunosorbent assays (ELISAs) using recombinant proteins revealed that the copy number of the repeat affects the affinity of binding between antigens and antibodies, as expected based on thermodynamic binding kinetics. Other than containing TR domains, the TR proteins do not share characteristics, such as sequence similarity or biased cellular location predicted by the presence of a signal sequence(s) and/or a transmembrane domain(s). However, the TR proteome contained a higher percentage of proteins upregulated in amastigotes than the whole proteome, and upregulated expression of a TR protein seemed to affect its antigenicity. These results indicate that Leishmania parasites actively utilize the TR protein family for parasitism in mammalian hosts. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 210.51K | Year: 2013

Develop an inexpensive, easy to use, aerosolized delivery system of a combination of anti-tubercular drugs that could be used for the treatment of multi-drug resistant tuberculosis (MDR TB ). PUBLIC HEALTH RELEVANCE

Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 600.00K | Year: 2013

DESCRIPTION (provided by applicant): Schistosomiasis is a major neglected tropical disease of public health concern to a billion people with 200 million currently infected and 779 million at risk to acquire the infection, the majority of these in Africa.The disease has a high impact on affected people's lives with disability adjusted life years at 70 million years which rank this malady ahead of malaria. Current control strategies have been geared toward repeated with a drug discovered in the 1970s and standards for monitoring administration and progress have been inconsistent and inadequate. Reliance on the drug therapy approach alone is a poor strategy since this approach has had little impact on the reduction of disease transmission and there is alwaysthe inherent threat of drug resistance being developed by the parasite. A prophylactic schistosomiasis vaccine that provides at least 50% protection would play an important role in dramatically reducing the impact of this disease. Vaccine-generated immuneresponses could lead to reduced worm burdens and lower egg production and ultimately result in lower transmission. This application is an extension of our systematic and methodical approach towards developing a vaccine for schistosomiasis. Over the last twenty years, we have applied this strategy towards developing Sm-p80 into a viable vaccine candidate. At present, to our knowledge, Sm- p80 is the sole schistosome vaccine candidate that has been tested for prevention, interruption of transmission and in therapy. Our candidate vaccine has three protective effects: worm reduction, egg reduction, and protection against acute disease. Funding this application will help our continuing efforts to develop Sm-p80 towards manufacture for future human clinical trials, ultimately resulting in an approved vaccine. PUBLIC HEALTH RELEVANCE PUBLIC HEALTH RELEVANCE: A low-cost, effective vaccine for schistosomiasis would greatly aid in the fight against this debilitating disease. By discovering and validating a new candidate vaccine we intend to progress toward a commercial product leading to an improvement in global health.

Protein Advances, Inc. | Date: 2012-07-18


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