Degenerative Disease Research

Developing Therapies to Prevent Neuronal Apoptosis

STUART LIPTON, MD., PH.D. Program Director
BIOGRAPHY
Stuart Lipton went to Cornell University, entered an M.D./Ph.D. Program at the University of Pennsylvania (UPENN), and completed his M.D. at UPENN and his Ph.D. with John Dowling at Harvard in 1977. He was then a medical intern and neurology resident at Harvard and a postdoctoral fellow of Torsten Wiesel when Wiesel won the Nobel prize in 1981. In 1997, after spending 15 years on the staff of Children's Hospital in Boston, Dr. Lipton moved to Brigham and Women's Hospital to become Chief of the CNS Research Institute at Harvard Medical School. Dr. Lipton was recruited to the Burnham Institute in September 1999 as Professor and Director of the Del E.Webb Center for Neuroscience and Aging Research. At that time he initiated the Center's program on Neurodegenerative Disease Research.

The Lipton laboratory studies molecular mechanisms of neurodegenerative diseases and stroke, including the role of excessive stimulation of ion channels and intracellular signaling pathways in nerve cells.Among the laboratorys accomplishments and ongoing activities are (i) the development of the first glutamate receptor/channel antagonist drug (Memantine) to be clinically approved for dementia (approved by the European Union in May, 2002), (ii) characterization of signaling events leading to neuronal injury and apoptosis in AIDS, and (iii) cloning of a gene that programs Embryonic Stem Cells to become nerve cells in the brain. These studies have led to the development of the first neuroprotective drugs to be administered successfully to humans to combat various neurodegenerative and vascular diseases of the brain.

ur laboratory uses basic molecular signaling pathways to prevent neuronal apoptosis and to promote neuronal survival and outgrowth during normal aging and various neurodegenerative diseases,

including cerebrovascular disease (stroke) and AIDS dementia. Neuronal damage is curtailed by preventing excessive activity of the NMDA subtype of glutamate receptor and its downstream effectors (see figure). Cultures of cerebrocortical neurons as well as transgenic and knock-out animal models are used to show the involvement of calcium, free radicals, caspases, and transcription factors in NMDA receptor-mediated neuronal apoptosis. Two NMDA antagonists that we have developed are clinically tolerated because they have been designed using biophysical principles to decrease

Dr. Lipton has been interested in the role of ion channels in neuronal outgrowth, plasticity and survival. His group has developed several clinically-tolerated drugs to prevent neuronal damage and apoptosis due to excessive stimulation of ion channels by excitatory neurotransmitters acting at the NMDA subtype of glutamate receptor in the brain. These drugs may be useful for several neurological disorders, including stroke, spinal cord and head injury, glaucoma, Huntington's disease, and AIDS dementia. Drugs developed in his laboratory are currently in advanced clinical trials. Recently, Dr. Lipton was asked to deliver a Nobel Foundation Lecture at the Karolinska Institute and a plenary lecture at the National Academy of Sciences on the topic of NMDA open-channel blockers and nitric oxide-related drugs for the treatment of AIDS dementia, a field in which he continues to be among the leaders.

only excessive NMDA receptor activity while leaving physiological levels of activity relatively spared - these drugs are now in clinical trials. Techniques used in the laboratory include patch-clamp recording, site-directed mutagenesis of recombinant NMDA receptor subunits and GABAC subunits, multi-photon confocal imaging of mitochondrial activities, deconvolution microscopy, gene reporter assays, and various fluorogenic methods for apoptosis assessment. Additionally, during the past few years we cloned and are currently characterizing two novel NMDA receptor subunits (one was recently published in Nature), and cloned a transcription factor, MEF2C, that controls the expression of NMDA receptor subunit genes and determines whether neurons undergo apoptosis after glutamaterelated insults (recently published in PNAS and JBC). MEF2C is activated by the p38 stress kinase pathway, an active area of research in the laboratory that mediates both neuronal cell apoptosis and ischemic tolerance in the brain. Recently, we also discovered a new action of nitric oxide-related species on cysteine residues of the NMDA receptor.This reaction, termed S-nitrosylation (transfer of the NO group to critical cysteine sulfhydryls), down-regulates NMDA receptor activity as well as caspase activity and may be useful clinically. Several other protein targets of nitrosylation are being examined in the laboratory (recently published by us in Neuron and in Nature). We have also found a possible cause of neuronal apoptosis in AIDS brains (about one-third of AIDS patients eventually develop dementia). We discovered that the coat protein gp120 of HIV-1 produces a dramatic rise in neuronal calcium. This destructive process is primarily mediated by stimulation/activation of macrophage

S TA F F
Mehrdad Alirezaei, Ph.D. Ella Bossy-Wetzel, Ph.D. Eleobardo Castro-Luque, M.D. Jiankun Cui, M.D. Murat Digicaylioglu, M.D., Ph.D. Thomas Goetz, M.D., Ph.D. Zezong Gu, M.D., Ph.D. Marcus Kaul, Ph.D. Shinichi Manabe, M.D., Ph.D. Bernd Moosmann, Ph.D. Shu-ichi Okamoto, M.D., Ph.D. Hiroto Takahashi, Ph.D. Maria Talantova, M.D., Ph.D. Gang Tong, M.D., Ph.D. Dongdong Yao, M.D., Ph.D. Mark Barsoum Chris Brechtel, B.S. Ruth Chen Vaninder Chhabra Brendan Eckelman Lauren Fletcher, B.S. Anne Harrop, B.S. Wilson Lee Zhen Li Katie Medders, B.S. Lutz Paessens, B.S. Dhakshin Ramanathan, B.S. Yeonsook Shin Lida Sionit, B.S. Jin Yang, B.S.

chemokine receptors by gp120 to release toxins that in turn trigger NMDA receptor-mediated neuronal destruction. Therefore, in some ways, this pathway resembles neuronal damage observed after stroke and other neurodegenerative diseases. (recently published in Nature, and Neuron, and JAMA). The involvement of apoptotic pathways in this type of cell death, involving reactive oxygen species, nitric oxide, mitochondrial toxins and caspases, is currently being explored.

Figure Legend Schematic illustration of the signaling pathways discovered or characterized in the Neurodegenerative Disease Program that can be targeted to prevent neuronal apoptosis and thus treat various neurologic diseases. Drug or molecular therapies are being developed to (1) antagonize NMDA receptors (NMDA-Rc), (2) modulate activation of the p38 mitogen activated kinase (MAPK) MEF2C (transcription factor) pathway, (3) prevent toxic reactions of free radicals such as nitric oxide (NO) and reactive oxygen species (ROS), and (4) inhibit apoptosis-inducing enzymes including caspases.

Publications Kaul, M. Garden, G.A., and Lipton, S.A. Pathways to neuronal injury and apoptosis in HIV-associated dementia. Nature 410:988-994, 2001. Digicaylioglu M. and Lipton, S.A. Erythropoietin mediated neuroprotection involves cross-talk between Jak2 and NF-kB signalling cascades. Nature 412:641-647, 2001. Chatterton, J.E., Awobuluyi, M., Premkumar, L.S.,Takahashi, H.,Talantova, M., Shin, Y., Cui, J., Sevarino, K.A.,Tu, S., Nakanishi, N.,Tong, G., Lipton, S.A., and Zhang, D. Excitatory glycine receptor containing the NR3 family of NMDA receptor subunits. Nature 415:793-798, 2002.