RESEARCH
Project 1: Drug Discovery using Flow Cytometric Techniques
Examination and Interruption of Malaria Cell Cycle
There is an unfavorable outlook for treatment and control of Plasmodium falciparum because of the limited supply of new antimalarial drugs under development, widely distributed resistance to many existing antimalarial drugs, and reduced effectiveness of some artemisinin combination therapies. Development of resistance is not surprising because many antimalarial drugs are cyto-static rather than lethal to the parasites. Coupling this lack of parasite death with patient treatment non-compliance and the high numbers of parasites exposed to drug pressure (1.2 x 10^17/year globally) increases the opportunity for the parasites to develop resistance mutations. The geographic isolation of many patients with malaria allows for these mutations to become established in the parasite population. With the recent annotation of the P. falciparum genome, rational-based searches for new, more effective antimalarial drugs can nowbe performed. We have recently made progress in developing a platform to discover  and investigate compounds with activity against specific proteins that regulate parasite growth and development employing new flow cytometry strategies (Grimberg et al. 2007). Our methods enable determination of IC50 values for major stages of P. falciparum development (eg, rings, trophozoites, schizonts) within infected red blood cells. This approach provides significantly more insight compared to current methods focused solely on parasite metabolism regarding the action of drugs against P. falciparum.
Evaluation of compounds to identify life cycle stage specific antimalarial drugs.
Access to the annotated P. falciparum genome sequence (PlasmoDB) facilitates exploration of different therapeutic targets. A preliminary PlasmoDB search identified 35 unexplored targets that should be important for parasite blood stage proliferation. These targets include cell cycle control proteins, cell cycle transcription factors, S-phase kinase associated protein family and anaphase-promoting complexes, DNA and RNA polymerases, helicases, and kinases that regulate chromatin condensation and merozoite formation.
New Antimalarial Drugs Detected Using High Content Screening
Project 2: P. vivax and P. knowlesi Invasion Inhibition
Investigation of Erythrocyte Duffy Antigen Expression and Interaction with Parasite Duffy Binding Protein
It has been observed that only people who have the Duffy antigen on the surface of their erythrocytes become infected with blood-stage P. knowlesi or P. vivax. Additionally, people who are  heterozygous for the Duffy antigen are half as likely to become infected by these same malaria species. Using a Duffy Binding Protein, which normally functions as a sink for excess chemokines, invading malaria parasites exploit the Duffy receptor on the erythrocyte to facilitate invasion. Several current vaccine strategies are focused on interruption of the interaction of the Duffy antigen and the parasite’s Duffy Binding Protein (Grimberg et al. 2007). While this strategy is promising, little is known about the expression of the Duffy receptor and whether such a vaccine strategy would be effective. Previous studies have suggested that Duffy antigen expression may vary across Duffy positive populations.
In our Duffy related research, my laboratory seeks to answer the following questions: a) is there a difference in the erythrocyte Duffy receptor expression or Duffy Binding Protein binding between individual blood donors of different sexes and races, b) does the age of the donor and of their red blood cells have an affect on Duffy antigen expression, Duffy Binding Protein binding, or parasite invasion, and c) does erythrocyte surface morphology influence the binding and invasion of erythrocytes by malaria parasites? Our overarching hypothesis is that decreased levels of functional Duffy receptors on the surface of host erythrocytes results in decreased malaria parasite invasion. To test this hypothesis we are examining the Duffy surface expression and parasite invasion of blood from human volunteers of different ages, sex, and races, hemoglobinopothies such as Sickle Cell, Southeast Asian Ovalocytosis, and b-thalassemia, and red blood cells of different ages using umbilical cords and stem cell derived erythrocytes. Our collaborations with the American Red Cross, University Hospitals of Cleveland, the Cleveland Clinic, the Papua New Guinea Institute for Medical Research, and the Kenya Medical Research Institute will help us to answer these questions. The level of Duffy antigen expression from the above studies are correlated to parasite invasion frequency. Human erythrocytes with a range of Duffy antigen expression and morphologies are being cultured in the presence of P. knowlesi or P. vivax to demonstrate the influence of level and functionality of Duffy antigen surface receptors on successful parasite invasion.
Other Projects:
Confocal microscopy, flow cytometry, and cell sorting techniques we recently developed (Grimberg et al., 2009) will be used to identify the induction of an apoptosis-like mechanisms which have been indicated in malaria but little studied. With my laboratory’s new methods, the activity of cell cycle specific antibodies, and sera isolated from patients infected with malaria, we will observe the timing and induction of parasite cell death. Furthermore, a panel of transfected GFP labeled malaria parasite strains will also be utilized to further explore the interruption of parasite life cycle stages and further understand the mechanisms of action of antimalarial drugs.
Potential Student Projects
We have had a great deal of success streamlining and simplifying the process of culturing malaria parasites without contamination. Several graduate, undergraduate, and even high school students have been successful at accomplishing this previously daunting and uncertain task. We have developed a stage-specific malaria parasite assay (Grimberg et al., 2008) which student’s could utilize to successfully accomplish a variety of research projects. Potential research projects include drug discovery utilizing my collaboration with local drug companies to test the effectiveness of novel compounds against malaria. Also, students could continue my work on the creation of drug resistant strains to allow for deeper understanding of the genetic mechanisms of drug resistance. Finally, answering basic questions about the biology of malaria parasites would serve as an excellent Graduate and Undergraduate projects. Students could culture and compete strains of malaria in various blood types, medias, and drugs to learn more about how the parasites react to different environments. These simple and straightforward tests would help increase our understanding of the history of this disease and unraveling its complexity.
PROFESSIONAL MEMBERSHIPS
American Association for the Advancement of Science
Scientist’s Without Borders, New York Academy of Sciences
American Committee of Molecular, Cellular, and Immunoparasitology
Faculty Senate Committee on Graduate Studies, CWRU
American Society of Tropical Medicine & Hygiene
Sigma Xi, Scientific Research Society
Genetics Society of America
American Society for Microbiology
AWARDS
Excellence in Science Award, American Association for the Advancement of Science
Postdoctoral Research Award for Outstanding Research, CWRU
Ruth L. Kirschstein National Research Service Award, NIH/NHLBI
Young Investigator Award, Great Lakes International Imaging & Flow Cytometry Association
Young Investigator Award, Honorable Mention, American Society. of Tropical Medicine & Hygiene
Postdoctoral Research Award for Outstanding Research, CWRU
Seniors’ Salute Honoree for Exemplary Undergraduate Mentoring, CWRU
Ruth L. Kirschstein National Research Service Award, NIH/NIAID
Outstanding Graduate Student Award, Department of Biology, WFU
On the Spot Award for Critical Contributions to the Institute, DHHS, NIH
Staff Recognition Award for Service Excellence, DHHS, NIH
Superior Merit Award for Efforts Above the Call of Duty, DHHS, NIH
CONTACT INFORMATION
If you are a highly motivated student, fellow, technician, or potential collaborator and are interested in working with my laboratory or if you just have a question about malaria, flow cytometry, drug resistance, or drug discovery, please feel free to contact me.
Brian T. Grimberg, Ph. D.
Case Western Reserve University
Biomedical Research Building Suite 427
2109 Adelbert Road
Cleveland, OH 44106
Phone: 216-368-6328
Fax: 216-368-4825
Email: endmalaria@case.edu
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