2002 Young Clinician Investigator

Nicholas Boulis, MD
The Cleveland Clinic

Phage display and In Vivo biopanning for small neurotropic peptides

The development of surgical therapies for Amyotrophic Lateral Sclerosis will depend on the production of advanced generation vector systems and therapeutic proteins targeted to spinal cord and brainstem. We intend to utilize the technique of phage display in order to define small peptides capable of triggering the uptake and retrograde transport of these therapeutic proteins and vectors into the spinal cord. By linking oligonucleotides to the peptides that they encode, phage display allows for libraries of up to 10⁹ sequences to be screened for axon terminal binding. Once defined, these oligonucleotides will be used to clone novel motor neuron targeted fusion proteins.

About Nicholas Boulis

Neuroscience Background: Dr. Boulis has conducted basic neuroscience research in six laboratories over a 13 year period. His research has evolved from a primary interest in neuroplasticity applied to human and animal emotion, learning, and memory into a focus on the recovery of the nervous system in the face of traumatic and degenerative disease. His first research effort in neuroscience addressed the issue of hemispheric asymmetry in emotional processing. This work was carried out utilizing dichotic listening and psychophysiology to address changes in hemispheric asymmetry occurring during the human menstrual cycle, and was done with Bruce Wexler, MD at the Yale School of Medicine. His second series of projects were conducted on invertebrate electrophysiology and learning with Christie Sahley, PhD in the Yale Department of Biology. In this laboratory, Dr. Boulis utilized a variety of behavioral and electrophysiological techniques to evaluate the cellular and synaptic plasticity underlying changes in leech behavior. In his third laboratory, he conducted experiments on rat learning, memory, and anxiety using the acoustic startle pathway as a focus. This work entailing rat behavior, pharmacology, HPLC, EMG, and stereotactic surgery was conducted with Michael Davis, PhD. Dr. Boulis was awarded the Angier Prize for the best senior essay on a psychological topic in his graduating class of over 100 students in psychology related disciplines.

At Harvard Medical School, Dr. Boulis worked with Huntington Potter, PhD on the molecular biology of Alzheimer's disease. In Dr. Potter's laboratory, he gained his initial exposure to molecular biology, attempting to utilize polymerase chain reaction to study the proteases important in the development of neuritic tangles and amyloid plaques. Next, Dr. Boulis worked with Larry Benowitz, PhD at Boston Children's Hospital on primary goldfish retinal cell culture and protein chemistry attempting to characterize the native neural growth factors of the fish optic nerve that might trigger regeneration. He completed a one year fellowship funded by the Howard Hughes Institute in the Benowitz laboratory. For this work, Dr. Boulis was awarded the Harold Lamport Biomedical Research Prize, for the best graduating thesis from Harvard Medical School in 1994.

Over the past six years at the University of Michigan, Dr. Boulis has developed a research group in the laboratory of Eva Feldman, MD, PhD for the application of viral gene therapy to the rat spinal cord. This group has focused on mechanisms of gene delivery to the spinal cord, vector development, and the delivery of neural growth factors through viral gene therapy. Dr. Boulis' initial interest in applying gene therapy to the spinal cord was an extension of his previous work with growth factors. He hypothesized that the delivery of neurotrophic genes might stimulate regeneration following spinal cord trauma and motor neuron disease. In order to minimize the parenchymal damage that was associated with the inflammatory response to direct vector injection, he developed a model for spinal cord gene delivery that depends on retrograde axonal transport after peripheral nerve injection. He has since demonstrated that no neuronal cell death occurs following this type of delivery, though expression remains transient. Throughout this time period, Dr. Boulis received training in vector development in the laboratory of Michael Imperiale, PhD. In Dr. Imperiale's laboratory, he has constructed two adenoviral vectors and one adeno-associated viral (rAAV) vector for the delivery of IGF-I. The work in both vector design and delivery is discussed in the preliminary data section of The Research Plan. Dr. Boulis' research throughout his residency was rewarded with both of the Department of Surgery's major prizes for research excellence; The Young Investigator Award, and The Crudup Award.

The Program For Understanding Neurological Disease (PFUND) at the University of Michigan, will continue to build on this work to apply gene based neurotrophic therapy to both traumatic and neurodegenerative disease. In particular, the program hopes to utilize peripheral nerve injection to deliver IGF-I vectors to impact on motor neuron survival in SOD-1 mutant mice, and DRG sensory neuron survival in diabetic animals. In parallel, an apparatus for inducing controlled spinal cord contusions has been constructed. Dr. Boulis will oversee experiments conducted by PFUND to apply viral vectors expressing neural growth factors to animals with these injuries assaying both recovery of limb function with behavioral assays as well as axonal regeneration using histological assays. Dr. Boulis will maintain his affiliation with PFUND in his new appointment at the Cleveland Clinic, acting as a consultant on administration of growth factor gene therapy.

In addition, he serves as a consultant to the ALS Therapy Development Foundation in the development of a clinical trial for rAAV vector gene therapy for ALS. In this trial, a vector for EAAT2, an excitatory amino acid transporter has been designed to reduce potential excitotoxicity in the spinal cords of ALS patients.

Career Goals: Dr. Boulis has accepted a staff position in the Department of Neurosurgery at the Cleveland Clinic with a clinical focus on deep brain stimulation for targeted neuromodulation. His clinical partner, Alai Rezai, MD, has published data suggesting that local injection of a GABA agonist is capable of reproducing the effects of deep brain stimulation on the basal ganglia in humans. Muscimol injections suppress human tremor and rigidity in Parkinsonian patients when delivered locally to the deep brain targets used in the treatment of movement disorders. While transient, this finding demonstrates the therapeutic potential of stereotactically delivered pharmacologically specific agents. Gene therapy has the capacity to create focal changes in synaptic function that will allow for controllable system specific neuromodulation without electronic prosthesis. Together with Thyagarajan Subramanian, MD, who will focus on the gene-based modulation of the dopamine system, Dr. Boulis will develop a laboratory for gene-based neuromodulation.