Drug Discovery and Development: The Senna Plant and Phosphomevalonate Kinase Inhibition Valders High School SMART Team: Corrine Brandl, Andrea Herrmann, Katarena Hubbartt, Nicole Maala, Alexandria Meidl, Hallie Reznichek Valders High School Instructor: Mr. Joseph Kinscher Mentor: Dr. Daniel S. Sem, Marquette University

Abstract. Heart disease is one of the most prominent causes of death in the United States. Hypercholesterolemia is an underlying factor leading to heart disease. By inhibiting the cholesterol synthesis pathway, scientists can develop ways to reduce the number of heart disease‐related deaths. One way could be by inhibiting the actions of phosphomevalonate kinase (PMK), a cytoplasmic enzyme found in the liver. The Valders SMART Team (Students Modeling A Research Topic), in collaboration with MSOE, built a 3D physical model of PMK. Both mevalonate 5‐phosphate (M5P) and ATP bind to the enzyme. Using ATP, PMK converts M5P to mevalonate 5‐diphosphate, a precursor to cholesterol. The inhibition of this process may be possible through a drug derived from the Indian senna plant, which is predicted to bind in place of M5P.

C. Brandl I. Introduction IV. A Closer Look at Cholesterol Synthesis V. Bringing It Together Mevalonate 5‐Phosphate + ATP

Heart disease is the leading cause of death in It now becomes necessary to consider the specifics of the Using nuclear magnetic resonance (NMR) the United States and as such, naturally attracts the process that cholesterol‐lowering drugs attempt to inhibit (figure 1). technology, Dr. Daniel Sem and his lab attention of persons looking to improve the There is only one pathway by which cholesterol is synthesized in determined that during the synthesis of lives of U.S. citizens. Developing drugs to humans, the HMG‐CoA reductase pathway. Statins inhibit an early mevalonate‐5 diphosphate, the ligand binding region of combat heart disease is a step in this process. The phenolic senna compound would inhibit phosphomevalonate kinase called the “Walker A” loop oscillated at A. Herrmann major focus of proteomic a later step, the synthesis of mevalonate‐5 diphosphate a rate of one hundred to six hundred times per second. (These data researchers. One success (a precursor to cholesterol) from mevalonate‐5 phosphate. can be seen in figure 3.) The phenolic senna compound was was the creation of statins, Phosphomevalonate kinase is the enzyme that makes the predicted to cause the same oscillation rate, suggesting that it was but the treatment is not transition possible, as it positions M5P and ATP in a way that likely to bind in the same manner and location in the kinase as M5P perfect; it may result in allows for thetransfer of the γ‐phosphate on ATP to the M5P. would. By binding in the same place, it would act as an effective side effects such as kidney inhibitor. damage. For this reason, research continues into K. Hubbartt Figure 1 different treatment http://www.teaflavin.com/images/new_site_images/deaths.jpg •The chemical shift perturbation options with the intention data obtained when M5P binds of eliminating these concerns. to PMK

II. Old Idea, New Application Figure 3 N. Maala Researchers are exploring whether compounds extracted from the Asian senna plant, Cassia VII. Conclusion angustifolia, may be used as a In 2008, the leading cause of death in the United States was heart substitute to statin drugs. The senna disease. Familial hypercholesterolemia (genetically high cholesterol) plant has long been used in folk VI. Mechanism for Phosphorylation of M5P is a major factor responsible for this unfortunate statistic. Inhibiting this remedies in Myanmar and Thailand, pathway is the purpose of cholesterol lowering drugs, such as statins. but has only recently attracted Mevalonate‐5 phosphate becomes mevalonate‐5 diphosphate by Proteomic researchers are looking into new potential drugs, such as A. Meidl the attention of researchers means of phosphorylation. This process requires the binding of phenolic compounds found in the senna plant, to be used instead of who are looking for new drug options. two ligands, M5P and ATP, to PMK. ATP binds first, resulting in a statins. Phenolic compounds http://product‐image.tradeindia.com/00239324/s/Senna‐Plant.jpg widening of the PMK binding site. When M5P binds later, it will hopefully inhibit the actions of contracts the binding site to allow γ ‐phosphate transfer to occur. phosphomevalonate kinase, (PMK), The “Walker A” loop counters these opposing charges with its which phosphorylates a precursor to high positive charge density (figure 2). A number of positively III. Exploring the Options Senna Phenol cholesterol called mevalonate 5‐ charged amino acids (such as Arginine 18, 19 and 110 phosphate. If proven successful, the and Lysine 17, 19 and 22), work to stabilize this negative charge to H. Reznichek Initially, the senna plant was analyzed next logical step is to develop a way to permit the phosphate transfer reaction to occur . There are also and nearly seven hundred compounds extract the necessary compound from hinge residues that permit the two domains to move together so were identified as potential therapies the senna plant in an efficient manner, the reaction may occur. (i.e. PMK inhibitors). Via computer that would allow the drug to be readily simulation, the compounds were available to those suffering from hypercholesterolemia. tested for their ability to inhibit the synthesis of Figure 2 cholesterol and the top five were selected for VIII. References •The chemical shift •NMR in drug discovery. more thorough testing. According to computer •Functional evaluation of conserved basic J. Kinscher of R18 and G21 showing Pellecchia M, Sem DS, Wüthrich K. predictions, the most likely candidate is a residues in human phosphomevalonate kinase. Nat Rev Drug Discov. 2002 Mar;1(3):211‐9. the Loop Motion of the Herdendorf TJ, Miziorko HM. phenolic compound. (As shown in the photo to the right.) Biochemistry. 2007 Oct 23;46(42):11780‐8 •Substrate induced structural and dynamic “Walker A” binding Site changes in human phosphomevalonate kinase when M5P binds •Phosphomevalonate kinase: functional and implications for mechanism. to PMK investigation of the recombinant human enzyme. Olson AL, Yao H, Herdendorf TJ, Miziorko HM, Herdendorf TJ, Miziorko HM. Hannongbua S, Saparpakorn P, Cai S, Sem DS. Biochemistry. 2006 Mar 14;45(10):3235‐42. Proteins. 2008 Sep 17;75(1):127‐138.

Dr. Daniel Sem A SMART Team project supported by the National Institutes of Health (NIH) –National Center for Research Resources Science Education Partnership Award (NCRR‐SEPA)