Immunotoxins. Protein toxins such as diphtheria toxin have three distinct domains that function together to kill cells. Chimeric toxins can be made by replacing the native toxin receptor domain with monoclonal antibodies or ligands that bind alternate receptors. These engineered toxins, or immunotoxins, can selectively kill various cell types based upon their surface receptor phenotype and selectively eliminate cancer cells. We have designed immunotoxins for brain tumor therapy and promising results have been obtained in phase 1 and phase 2 clinical trials. Study of improved immunotoxin designs, improved delivery and minimization of nonspecific toxicity is currently underway. Use of human cytotoxic proteins for cancer therapy, including apoptosis promoting proteins, is being explored.

Programmed cell death. Neurons die in great numbers during normal human development. Our early work showed that developmental neuron death occurs by apoptosis and studies with knockout mice revealed that p53 can play a role in neurodegenerative processes but not developmental neuron death. Bax, Bcl-XL and caspases, however, regulate developmental apoptosis of neurons. Understanding the molecular basis of cell death may lead to strategies to block neuron loss during acute and possibly chronic neurodegenerative diseases. We have found that Bcl-XL and Bax move from the cytosol compartment to the mitochondria during apoptosis. Two major aspects of this process are under investigation. The molecular trigger for Bax migration into mitochondria and the consequences of Bax insertion into mitochondria are not known and appear to be a key for the control of cell death. We are studying the effect of Bax on mitochondria morphology (see figure) and the molecular mechanism of mitochondrial fission that occurs during apoptosis. Use of green fluorescent protein fused to Bcl-2 family members analyzed by confocal microscopy is instrumental in these studies. We also have been able to engineer Bcl-2 family members into potentially therapeutic fusion proteins that prevent apoptosis of neurons in vitro and in vivo and may have potential for treatment of spinal cord injury and stroke.

Figure above: Mitochondria in a single cell upon inhibition of the large GTPase, Drp1, labeled with green fluorescent protein (left) and Mitotracker and overlayed (right) show an unusual fused morphology and different mitochondrial membrane potentials.

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