Impact of Heart Failure and Omecamtiv Mecarbil on Beta-Cardiac Myosin Function
Open Access
- Author:
- Fetrow, Christopher Michael
- Graduate Program:
- Anatomy
- Degree:
- Master of Science
- Document Type:
- Master Thesis
- Date of Defense:
- March 17, 2016
- Committee Members:
- Christopher Martin Yengo, Thesis Advisor/Co-Advisor
Patricia Mc Laughlin, Thesis Advisor/Co-Advisor
Charles H Lang, Thesis Advisor/Co-Advisor - Keywords:
- Cardiac Myosin
Heart Failure
Omecamtiv Mecarbil - Abstract:
- Abstract Purpose: Heart failure (HF) affects approximately 5.1 million people in the United States and is treated with a variety of drugs, many of which have long-term side effects such as cardiac arrhythmias. Due to a high mortality rate, it is important to find a drug that is more efficient and safe for long-term use. The small molecule cardiac myosin activator drug, omecamtiv mecarbil (OM), has been proposed as an effective treatment of heart failure. In this study we investigate the effects OM has on the interaction of actin and myosin. Methods: Recombinant myosin was expressed in a C2C12 expression system which was compared to myosin purified from human heart tissue provided by The University of Kentucky Biobank. We performed, in vitro motility and ATPase assays with and without OM to investigate the mechanisms of action of this drug. In vitro motility was obtained using fluorescent microscopy after fixing myosin to the surface of a microscope slide and inserting fluorescently labeled actin to create interaction with the surface attached myosin. The velocity, myosin density dependence, and actin filament length dependence were examined. ATPase assays were performed by examining actin-activated ATPase in the presence and absence of OM. The maximum ATPase rate (Kcat) and actin concentration at which the ATPase activity is ½ the maximum (KATPase) were obtained by plotting the ATPase activity as a function of actin concentration. Results: ATPase assays showed there was a 6-fold decrease in the actin-activated ATPase rate. In vitro motility showed a 100-fold decrease in filament velocity and a small difference in length dependence with and without OM. Density dependence demonstrated that in the absence of OM, velocity increased as the density of myosin was increased while the trend was the opposite in the presence of OM. Lastly, purified myosin from human heart tissue showed little movement in the absence of OM (3% filaments moving). However, the presence of OM dramatically increased the percentage of filaments moving (93%). Conclusion: Based on our results, OM increased myosin’s affinity for actin; however it remains unclear what the mechanism is behind the decrease in velocity in the in vitro motility assay. The decrease in sliding filament velocity may be detrimental to heart function and thus a therapeutic dose may need to be carefully chosen. Although the changes in the specific steps of the myosin ATP cycle are unclear, the enhanced interaction with actin in the presence of OM may increase force production in cardiac muscle.