Brian Jason Wainger - US grants

DESCRIPTION (provided by applicant): This proposal describes a five-year plan for Brian Wainger to achieve independence as an investigator who uses patient-derived motor neurons to study amyotrophic lateral sclerosis (ALS). The candidate is a neurologist at Massachusetts General Hospital. He has a strong research background in electrophysiology and molecular biology, as well as recent successful experience recording from patient-derived motor neurons, a technique that he will apply to the proposed project. Dr. Clifford Woolf, the primary mentor, is Professor of Neurology and Neurobiology at Harvard Medical School and Director of the Neurobiology Program at Children's Hospital Boston. He is a world expert in neuroscience, and has supervised numerous trainees who now hold academic faculty positions. Dr. Kevin Eggan, the co- mentor, is an HHMI Early Career Scientist and Professor in the Department of Stem Cell and Regenerative Biology and the Harvard Stem Cell Institute. He is a leader in the stem cell field and in ALS research. An advisory committee of senior scientists able to provide additional expert guidance includes also Dr. Merit Cudkowicz and Dr. Bruce Bean. The research will be performed at Children's Hospital. ALS is a devastating, untreatable neurological disease characterized by progressive weakness and death of neurons in the motor system. Most cases are sporadic, but about 10% are familial. The primary function of a motor neuron is to collect and integrate signals from the brain and spinal cord and transmit an outgoing electrical signal that results in muscle contraction. The researchers hypothesize that investigating the electrical properties of healthy and diseased motor neurons will help increase understanding of ALS and yield insight into how to treat the disease. Using recent advances in stem-cell technology, the investigators have derived motor neurons from ALS and control subjects and found that motor neurons derived from patients with familial ALS are hyperexcitable, meaning that they are prone to too much activity, and that the hyperexcitability may contribute to motor neuron death in ALS. The proposed project consists of evaluating hyperexcitability and motor neuron death in a range of inherited and sporadic variants of ALS. The project will yield a mechanistic understanding and characterization of the observed hypexcitability, specifically with regard to the role of voltage-gated potassium ion channels. Through the training program, the investigator will acquire critical expertise using techniques in stem cell biology and single cell RNA expression analysis, as well as didactic exposure to key components of translational medicine such as biostatistics and trial design. With these skills, the applicant will successfully complete the proposed project and transition to independence, where he can exploit this novel approach of modeling human neurological disease using human neurons more broadly.

Project Summary: The primary goal of this proposal is to develop and apply an in vitro model of the  neuromuscular junction (NMJ) using motor neurons and muscle both derived from human induced  pluripotent stem cells (iPSCs). The research will combine state-­of-­the-­art stem cell biology, gene  targeting, electrophysiology and genomic techniques. The main focus of the application will be in  amyotrophic lateral sclerosis (ALS), commonly known as Lou Gehrig?s disease, which is a devastating  disease of the motor nervous system with an onset often in the prime of life and an average survival of  only two to three years. ALS affects about 20,000 people in the US at any time. About 10% of cases  are due to dominant familial mutations and 90% are apparently sporadic. Degeneration of the NMJ is  the earliest pathological feature of ALS in both humans and mouse models. Involuntary muscle  contractions, known as fasciculations, are the first clinical disease symptom and reflect abnormal motor  neuron electrical discharges, which originate in the distal axon and NMJ. The NMJ novel model brings  to bear the advantages of human stem cell approaches, namely, to overcome limits of mouse models  including artifacts that result from heterologous over-­expression of human proteins in a mouse  background and inability to capture complexities of diverse human genetic backgrounds in mice.  Furthermore, mouse models are limited to the familial forms of the disease, while iPSC-­based models  have the potential to address sporadic disease, the vast majority of ALS cases. The applications of the  model described in the protocol include identifying morphological and genomic phenotypes of the ALS  NMJ, determining how the motor neuron and muscle each contribute to abnormal motor neuron  physiology and reciprocally how abnormal motor neuron excitability affects the NMJ. These questions  will be addressed across a range of familial and sporadic ALS variants. While still clearly reductionist,  the model goes beyond isolated iPSC-­derived motor neurons to add the disease-­relevant anatomical  context of the NMJ and thus give additional structural meaning to neuronal components like the initial  axon segment, axon and distal motor terminals and mechanistic processes implicated in ALS such  axonal transport. The human NMJ model developed in the proposed project will be useful to identify  specific ALS subgroups that share common disease mechanisms and to find targets and evaluate  candidate therapeutics appropriate for those individual disease subgroups. The model will also be  broadly applicable to other motor neuron diseases, such as spinal muscular atrophy, as well as motor  neuropathies and myopathies.