- Development of a Bovine Milk Immunoglobulin (BIgG) Supplement that Prevents Traveler's Diarrhea
- Vasopressin as a First-Line Pressor Agent in the Treatment of Septic Shock
- Understanding a Role of Bone Marrow in Development of Childhood Asthma
- Future Molecular Diagnosis of Systemic Lupus Erythromatosis
- ON 01210: A Novel Radioprotectant Drug
- Insights into Acute Respiratory Distress Syndrome
Development of a Bovine Milk Immunoglobulin (BIgG) Supplement that Prevents Traveler's Diarrhea
Posted December 18, 2007
Stephen J. Savarino, M.D., M.P.H, Naval Medical Research Center, Silver Spring, Maryland, and Henry M. Jackson Foundation for the Advancement of Military Medicine, Rockville, Maryland
Diarrhea caused by enterotoxigenic Escherichia coli (ETEC) is a significant health threat for military personnel and civilians traveling to developing countries and is the leading bacterial cause of childhood diarrhea on a global scale. Incidence rates as high as 50% occur in travelers where food and water sanitation is poor. ETEC elicits diarrhea through expression of fimbrial colonization factors that promote intestinal adherence and production of enterotoxins that cause the extrusion of essential electrolytes and water into the small bowel. Rehydration and antibiotic treatment are the cornerstones of disease management. Even with early institution of appropriate therapy, however, ETEC diarrhea exacts a toll in terms of lost duty and diminished performance. Currently, there is no licensed ETEC vaccine on the U.S. market.
CAPT Stephen Savarino and his colleagues, recipients of a Fiscal Year 2003 Department of Defense Peer Reviewed Medical Research Program Award, are developing a promising, new anti-diarrheal oral supplement. Their main objective is to produce an oral immunoprophylactic that will confer protection against a broad number of types of ETEC. CAPT Savarino had previously identified a class of adhesin proteins that are found at the tips of ETEC fimbriae and that directly mediate bacterial adherence to the intestine. Because antibodies to these adhesins prevent ETEC bacteria from binding to target cells in vitro, he proposed that oral delivery of such antibodies to humans would prevent ETEC binding the gut and thereby prevent diarrhea. These antigens are highly conserved, and antibodies to a panel of as few as three adhesins could neutralize adherence to a broad number of ETEC types. This could have the benefit of reducing the eventual cost of a passive or active adhesin-based ETEC vaccine.
The scope of work in this research program included the isolation and purification of a panel of ETEC antigens that were used to immunize pregnant cows. After giving birth, the cows' earliest milkings (colostrum) were collected as a concentrated source of hyperimmune anti-adhesin antibodies. In a process previously developed by collaborators at ImmuCell Corporation, milk whey was collected as the byproduct of a cheese-making process and further processed to increase the concentration of antibodies in the final powdered product. These high-titer antibody preparations (referred to as bovine milk immunoglobulin or BIgG) were administered orally to healthy adult volunteers in a randomized, placebo-controlled, double-blinded in-patient study at the Johns Hopkins Bloomberg School of Public Health. After three days of oral treatment with the BIgG preparations, volunteers were challenged orally with a live, diarrhea-causing ETEC strain. Antibodies to the prototype adhesin conferred significant protection against diarrhea. This has provided the first evidence that such adhesins play a central role in human disease and that anti-adhesin antibodies alone can prevent ETEC diarrhea, representing a major breakthrough in the field.
In addition to evaluating the protective efficacy of anti-adhesin BIgG products, CAPT Savarino's research team has also manufactured a panel of ETEC strains being used in the development of new volunteer challenge models. For the first time, ETEC challenge strains that express CS17 and CS19 fimbrial colonization factors are available. Furthermore, a human challenge model with CS17-producing ETEC has been established. A model for challenge with a CS19-producing ETEC strain is currently under development at the Johns Hopkins Bloomberg School of Public Health. CAPT Savarino's colleagues in Lima, Peru, have also established new ETEC challenge models in nonhuman primates. These new human and monkey models will be invaluable for screening vaccine candidates in the future.
Taken together, the results of CAPT Savarino's studies hold great promise for the future development of an oral anti-diarrheal supplement that provides broad protection against ETEC diarrhea.
Savarino SJ, et al. Bovine milk anti-adhesin antibodies confer protection against enterotoxigenic Escherichia coli diarrhea in the volunteer challenge model (manuscript in preparation).
Septic shock is a high-risk disease of infection seen in all Army hospitals, and its efficient cure is a significant military-relevant disease management concern. The success rate of septic shock treatment remains poor due to the inability to consistently improve organ perfusion and prevent multi-system organ failure. Currently, vasopressin (VP) is not part of the early intervention strategy for septic shock, but it is used, instead, only after all other pressor agents have failed. This avoidance of VP is due to a clinician's concern that the drug's strong vasoconstricting action may be detrimental to perfusion of certain organs such as the gastrointestinal tract.
With support from a Fiscal Year 2002 Peer Reviewed Medical Research Program Award, Dr. Catherine Uyehara and her research team have utilized successfully established animal models to demonstrate that VP may have a great beneficial effect in treating septic shock beyond its ability to simply raise blood pressure. The group found that VP could 1) completely reverse the hypotension induced by endotoxin, in contrast to dopamine (DA) and norepinephrine (NE), 2) preserve blood flow to the brain, heart, and kidneys while decreasing flow to the skin, gut, and muscle, and 3) preserve renal function, with a dramatic increase in urine flow, in contrast to the renal shutdown seen with DA and NE. Importantly, the VP-induced decrease in blood flow to the gut did not result in pathological tissue changes different from that of untreated or NE-treated endotoxin groups. Also, DA appears to actually protect against early ischemic changes due to DA-dependent increase in blood flow to the small intestine. As a result of the differential effects that VP has on different compartments, the scientific team has hypothesized that there is a differential distribution of vascular VP receptors within different vascular beds. Their data also suggests that VP may stimulate adrenal function, resulting in greater release of cortisol. Thus, VP may help counteract adrenal insufficiency in septic shock by stimulating the adrenal gland.
Dr. Uvehara's study has provided a better understanding of the effects of VP on organ perfusion in comparison to other commonly used pressor agents, and this information may be used to design optimal pharmacologic tools and clinical guidelines in the treatment of septic shock.
Asthma is one of the most common chronic diseases of childhood. In the U.S. there are approximately nine million children under 18 years of age who have been diagnosed with this disease. Childhood asthma is a complex disease in which multiple mediators and cell types contribute to airway pathogenesis. Dr. Mary Beth Hogan, a recipient of the Fiscal Year 2001 (FY01) Department of Defense Peer Reviewed Medical Research Program (DOD PRMRP) Investigator-Initiated Research Award at West Virginia University, has adapted an animal model of asthma to study the effects of pulmonary allergen exposure on bone marrow eosinophil progenitor cells (CFU-eo). Pulmonary exposure to allergen results in damage to bronchioles by invasion of eosinophils in asthmatic children. Eosinophils are inflammatory cells that play a role in allergic response, have limited life spans, and must be continually renewed in the bone marrow. Since eosinophils are considered the main effector cells in asthma pathogenesis and are associated with disease severity, Dr. Hogan's study demonstrates the critical importance of studying other regulatory mechanisms in the bone marrow, such as stromal cells. Data generated indicated that stromal cells may contribute to an increase in eosinophil production found during an asthmatic episode. Previous research had suggested that IL-5 is the primary cytokine that regulates eosinophil production and was originally thought to be synthesized exclusively by T lymphocytes. Dr Hogan's work has revealed that both stromal cells and T lymphocytes have a contributory role in both normal and accelerated eosinophil production in asthma. In addition, inflammatory mediators released from the lung were shown to modify stromal cell support of eosinophilopoiesis, which may contribute to the chronic inflammation associated with long-term asthma. Dr. Hogan has shown that asthma has systemic effects upon bone marrow regulation of hematopoiesis, in particular eosinophilopoiesis. She and her team's interest in childhood asthma has led them to investigate events in eosinophilopoiesis during the initial phase of the development of asthma and in events contributing to the development of chronic asthma. Taken together, Dr Hogan's findings on the relative roles of T lymphocytes and stromal cells in eosinophilia may aid in the design of new asthma treatments.
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Future Molecular Diagnosis of Systemic Lupus Erythromatosis
Posted October 25, 2007
Timothy W. Behrens, M.D., Kathy Moser, Ph.D. and Emily Baechler Gillespie, Ph.D., University of Minnesota, Minneapolis, Minnesota
Systemic Lupus Erythematosus (SLE) is a chronic, inflammatory autoimmune disease that can cause irreversible damage to a broad range of organs and tissues, and it affects women more often than men. The extensive clinical heterogeneity of SLE, primarily due to the involvement of a broad range of self-antigens, presents a tremendous challenge in diagnosis and management of this disease. Consequently, there is significant interest in identifying biomarkers that will help physicians better manage these patients. More accurate diagnosis and prediction of the disease would allow physicians to: (1) recommend appropriate treatment, (2) identify a risk group for adverse effects of vaccines, and (3) distinguish between lupus and other inflammatory as well as non-inflammatory illnesses. Furthermore, a panel of lupus-specific biomarkers is likely to be relevant for diagnosing many other autoimmune disorders, which affect up to 5% of the population, since a vast array of self-antigens are targeted in SLE patients.
With the support of the Peer Reviewed Medical Research Program Fiscal Year 2005 Investigator-Initiated Research Award, Dr. Emily Gillespie and colleagues have identified numerous gene expression signatures that are differentially expressed in peripheral blood cells of patients with different states of lupus activity: current, future, and quiescent. The researchers selected unique lupus-specific gene signatures using bioinformatic modeling of existing expression profiles generated with genome-wide microarrays (143 SLE patients). Additional statistical analyses of the selected genetic data demonstrated the ability to classify the current, future, and quiescent disease states of these lupus patients with 90%, 90%, and 89% accuracy, respectively. In order to test these predictors in a larger, independent group of patients, Drs. Moser and Gillespie designed and successfully tested a multiplexed real-time PCR assay for detection of the first, most promising gene-expression signature. This group continues their research efforts to further refine and validate the most specific genetic biomarkers which will help to significantly improve clinical management for those affected by lupus.
Nuclear/radiological agents are a significant threat to the United States military. The chances of exposure or actual occurrence of exposure to ionizing radiation has the potential to significantly impact the planning and execution of military operations, making it essential that radiological therapeutics are available. With the support of the Peer Reviewed Medical Research Program (PRMRP) fiscal year 2002 (FY02) Investigator-Initiated Research Award, Drs. K. Sree Kumar, Thomas Seed, and colleagues are conducting preclinical studies with ON 01210, a drug that appears to be highly promising for radioprotection. Due to the nature of the threat posed by the radioactive hazards and the current global circumstances, Dr. Kumar's study is aimed at rapid development of the ON 01210 drug as a radioprotectant therapy.
ON 01210 is a synthetic, small molecular weight, sulfur-containing organic compound designed and developed by Onconova Therapeutics Inc., New Jersey, for the specific purpose of selectively modifying cell cycle distribution patterns of critical cells of the body in order to shield those sensitive cells from genotoxic damage. ON 01210 is uniquely specific to target mammalian cells and has the ability to arrest normal cells at the G1 and G2 checkpoints of the cell cycle. Alkaline comet assays conducted by Drs. Perkin and Kumar have shown that exposure of cells to ON 01210 before irradiation results in significant protection from DNA damage, whereas untreated cells experience significant DNA damage due to irradiation exposure. Toxicology studies have shown the drug to be pharmacologically safe, and no major differences have been observed in the behavior and physiological state of ON 01210-treated animals. Dr. Kumar's research of ON 01210 is showing promising results and may be used as a preventive medical countermeasure for high-risk military personnel as protection from lethal radiation exposures in the future.
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Acute respiratory distress syndrome (ARDS) is a complication arising from both direct and indirect lung injury, such as blast injury and hemorrhagic shock/resuscitation, respectively. It is characterized by alveolar damage, disruption of alveolar epithelium, and infiltration by immune cells into the parenchyma. This inflammatory process, as indicated by the activation and entry of immune cells into the parenchyma, can either be localized in the lung, or it can involve other organs, which is the case for multi-organ failure. The mortality rate for ARDS is high, 45% to 92%, depending on the etiology, with the majority of deaths occurring in those patients that have multi-organ failure. Labile iron released from hemoglobin and extravasated blood is often associated with traumatic injuries, including those leading to ARDS, and it is known to catalyze production of free radicals. Dr. James Atkins of the Walter Reed Army Institute of Research, with funding from the Fiscal Year 2003 Peer Reviewed Medical Research Program Investigator-Initiated Award, is examining the relationship between the iron and the inflammatory response seen in ARDS. Dr. Atkins found that there is a robust production of nitric oxide (NO) following traumatic injury, such as hemorrhagic lung trauma. Nitric oxide bioactivity can alter proteins and release protein-bound iron. The radicals produced through iron initiate inflammatory signals within the cells. Redox-sensitive transcription factors move to the nucleus and lead to increased expression of molecules that bind the immune cells (ICAM). At the same time, the junctions between endothelial cells are opened (VE-cadherin). These two changes help to "pull" the immune cells across the endothelial lining of the blood vessels and into the lung parenchyma. Dr. Atkins and co-workers also found that hemorrhage results in a large increase in the production of NO. Resuscitation from hemorrhage increases iron in the blood stream and increases the amount of labile iron in the lung. The results suggest that even though the sources of iron in the lung are quite different in direct lung trauma and hemorrhage/resuscitation, the increase in labile iron may be a common mechanism helping to initiate the inflammatory cascade. The source of increased blood iron in hemorrhage/resuscitation is a focus of ongoing investigation. The link between ARDS and multi-organ failure is controversial and it may involve mediators released from the inflamed lung or a common mechanism of injury such as increased iron.
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Gorbunov NV, Asher LV, Ayyagari V, et al. 2006. Inflammatory leukocytes and iron turnover in experimental hemorrhagic lung trauma. Experimental and Molecular Pathology 80(1):11-25.
Atkins JL, Day BW, Handrigan MT, et al. 2006. Brisk production of nitric oxide and associated formation of S-nitrosothiols in early hemorrhage. Journal of Applied Physiology 100(4):1267-1277.
Gorbunov NV, Das DK, Goswami SK, et al. 2006. Spatial Coordination of Cell Adhesion Molecules and Redox Cycling of Iron in the Microvascular Inflammatory Response to Pulmonary Injury. Antioxid Redox Signal. In press.