Apoptosis or Programmed Cell Death is an important biological process that is required to maintain the integrity and homeostasis of multi-cellular organisms. Apoptosis is initiated by extracellular or intracellular signals in which a complex machinery is activated to start a cascade of events that ultimately leads to the degradation of nuclear DNA and a dismantling of the cell. Apoptosis is a tightly regulated and conserved mode of cell death that does not result in injury to adjacent cells. However, necrosis is a catastrophic, unregulated mode of cell death that is characterized by an invasion of inflammatory cells and injury to adjacent tissue. Abnormal or unregulated apoptosis is believed to occur in a variety of disease states, including ischemia-reperfusion injury (i.e., myocardial infarction and stroke), neuro-degeneration (Parkinson's Disease, Alzheimer's Disease, and ALS), sepsis, and diabetic cardiomyopathy. On the other hand, the inability of a cell to undergo apoptosis is believed to play a major role in the resistance of tumor cells toward radiation, chemotherapy, and immunotherapy. Therefore, the development of a non-invasive imaging procedure that can measure apoptosis is of importance to many institutes within the National Institutes of Health. The current method for imaging apoptosis uses radiolabeled analogs of Annexin V. Annexin V is a protein that binds with high affinity to phosphatidyl serine residues that are exposed as part of membrane inversion that occurs during apoptosis. However, since membrane inversion also occurs during necrosis, imaging studies using radio-labeled Annexin V measure both apoptosis and necrosis. An alternative strategy for measuring apoptosis is to determine the level of caspase-3 activity in the cell. Caspase-3 is an """"""""executioner"""""""" caspase and is released from an inactive zymogen (procaspase-3) late in the process of apoptosis. Furthermore, evidence suggests that inhibitors of caspase-3 may provide an effective method for preventing cellular death that occurs in diseases characterized by an increase in apoptosis (i.e., ischemia-reperfusion injury, neuro-degeneration). Therefore, a noninvasive imaging procedure that can quantify caspase-3 activity would be useful in the study of apoptosis in a wide variety of clinical conditions. However, one of the complications of imaging apoptosis is that the amount of cells undergoing apoptosis at any given time may represent only a small fraction of the total cells in an intact organ. The goal of this research project is to develop PET imaging agents possessing a high affinity and selectivity for caspase-3. The design of the imaging agents is based on a novel class of caspase 3 inhibitors developed in our laboratory, termed the Isatin Michael Acceptors or IMAs. These compounds are expected to bind irreversibly to activate caspase 3 in cells undergoing apoptosis. The irreversible binding property of the IMAs is expected to provide a high signal: noise ratio and provide a sensitive method for imaging cells undergoing apoptosis that would be of tremendous value to the PET research community.
