Facial Pain Research Foundation Awards Grant

To Dr. Todd Golde For Ongoing Project:

Towards Gene Therapy for Trigeminal Neuralgia 

The Facial Pain Research Foundation is pleased to have approved a research grant to Dr. Todd Golde.  The funding is a continuation of his efforts to find cures for trigeminal neuralgia and related neuropathic pain. His research project is entitled “Towards Gene Therapy for Trigeminal Neuralgia.  The Trustees of the Facial Pain Research Foundation believe that this research has the potential to make a significant difference in the lives of many in pain. 

Todd E. Golde, M.D., Ph.D., is executive director of the Evelyn F. and William L. McKnight Brain Institute of the University of Florida. He oversees neuroscience and neuromedicine research programs across the University of Florida campus. A professor of neuroscience and neurology, Dr. Golde joined the University of Florida faculty in 2009 and became founding director of the university’s Center for Translational Research in Neurodegenerative Disease, which he led until taking charge of the McKnight Brain Institute in December 2016. He is also director of the NIH funded 1Florida Alzheimer’s disease Research Center group of institutions. 

An internationally known expert in the scientific understanding of Alzheimer’s disease, Dr. Golde has published more than 240 papers that have been cited more than 30,000 times, (https://scholar.google.com/citations?user=X_9xacsAAAAJ&hl=en), and has expanded his leading-edge research to include other neurodegenerative diseases, cancer and even malaria. He is lead investigator for the Facial Pain Research Foundation’s Gene Therapy Research Project, at the University of Florida, to find the cures for TN.  

Focal delivery of rAAV vectors to the trigeminal nerve has a huge upside, the capacity to directly deliver to various places on the trigeminal “waterway” large therapeutic doses of compounds that inhibit/block pain signaling with minimal if any side effects. Perfecting delivery of rAAV to peripheral nerve and trigeminal nerve together with an ability to evaluate a large number of possible biotherapeutic “cargos” encoded within the rAAV vector will enable the development of a gene therapy that they hope will result in a permanent cure for TN. 

The primary objective of this project is to design and synthesize a number of viral vectors directed at specific genes related to pain processing and then determine which of these constructs are the most efficacious in doing so…initially in animals.  Dr. Golde points out that animal testing is an essential requirement mandated by the Food and Drug Administration (FDA) to assess safety in translating a therapeutic strategy from animals to humans.  Further, he posits that in addition to determining safety, this approach should decrease the time needed to discover novel new therapies for the treatment of TN.  Dr Golde’s team will utilize established expertise in  Adeno Associated Virus (AAV) delivery systems to evaluate the effectiveness of these vectors for reducing pain.  

Dr. Golde’s team fully recognizes the need to be substantially more rigorous about monitoring the delivery of the therapeutic to the nerve. Using cargos that are expressed by the rAAV vector and are functionally within nerves, they will evaluate whether there is the ability to block pain in several models including models of trigeminal nerve pain developed by Drs. John Neubert and Rob Caudle at the University of Florida’s McKnight Brain Institute. 

Mervyn Rothstein Interviews

Cory R. Nicholas Ph.D.

 

 

Cory R. Nicholas, Ph.D., is a co-founder and chief scientific officer of Neurona Therapeutics. He is also an adjunct assistant professor at the University of California, San Francisco. Before co-founding Neurona, Dr. Nicholas was a faculty member in the Department of Neurology at the University of California, San Francisco, where his research program was focused on investigating the development of human cortical interneurons from their earliest stages to maturity.

Dr. Nicholas pioneered methods to obtain interneuron forerunners from human immature stem cells that were capable of being transplanted into multiple animal models of neurological disease. Neurona was conceived in 2008 to investigate the potential of neuronal cell-based therapy – using such cells as a basis for eliminating neurological pain. Dr. Nicholas and his co-founders, John Rubenstein, M.D., Ph.D., and Arturo Alvarez-Buylla, Ph.D., were among the first to do research that led to the discovery that these cells could potentially inhibit hyper-excitable neural networks in relevant neurological disorders – that the injected interneuron precursor cells became integrated into neural circuitry, possibly allowing for stable repair of the injured nervous system.

The Facial Pain Research Foundation recently agreed to finance a cooperative research project between Neurona and the University of Florida’s McKnight Brain Institute to test the Neurona neuro-stem cells to see if they will stop neuropathic pain in animals. The study should take from 12 to 24 months. If it is successful, Neurona plans to seek Food and Drug Administration approval to begin studies in humans that hopefully will lead to clinical use. 

Dr. Nicholas spoke by telephone from California with Mervyn Rothstein, who was an editor and writer at The New York Times for nearly 30 years before retiring in 2010. Mr. Rothstein has also written for Playbill Magazine, and now playbill.com, since 1991. He currently writes the Stage Directions column for the Playbill web site. He was diagnosed with Trigeminal Neuralgia in 2005. Dr. Nicholas spoke about his research, Neurona, the agreement with the Facial Pain Foundation, and his future hopes.

Why did you decide to focus on pain in your career?

I’m a developmental biologist and I was studying the development of the nervous system, specifically the development of inhibitory nerve cells that are the basis of the collaborative work with the foundation. And we had a colleague at the University of California at San Francisco, Allan Basbaum, who was very interested in seeing if these inhibitory cells could be applied to the treatment of pain, which is his expertise. So our groups joined forces. We were able to work together, and Allan in his laboratory took this and really pioneered application of the cellular transplants that we were studying in the brain and applied them to the spinal cord. And he was able to show that the transplantation of those cells could ameliorate and in some cases completely reverse the experimental pain sensitivities in rodent models of neuropathic pain. 

What is the importance of this Trigeminal Neuralgia research study?

I think this is a tremendous opportunity to take the next step and to build upon the work from Allan and others in our group and really to leverage the resources from the network at the Facial Pain Research Foundation. Because we’ve not yet tried putting these human nerve cells that we think can be restorative and potentially curative into any type of animal model of Trigeminal Neuralgia. One of the reasons we weren’t able to do this on our own is that we didn’t have access to the animal models of this disease. These animal models are quite nuanced and can be difficult to establish on our own. That was when we were introduced by Michael Pasternak of the Foundation to John Neubert at the University of Florida, who had this model. He’s been studying it for a while and had it up and running at his lab. Thankfully Michael Pasternak and the Foundation were able to make that introduction.

How would you describe the work being done at Neurona and its future and its progress?

We submitted a proposal to the Foundation that was just funded where we can do three things. Number one, we can see if the pain phenotype – the observable pain characteristics –  that John has studied for a long time in his animal model can be extended to an animal that does not have an immune system. We need an animal that does not have a functioning immune system to allow for the long-term persistence of the human cells in a rat, because the same kind of immunosuppressants that we will be using in the future, potentially in clinical trial for humans, those drugs don’t always work in rodents. So we had to change the animal strain – so that is the first aim, to see if we can produce a stable pain phenotype using the animal model of Trigeminal Neuralgia that John has studied.

The second aim to see whether our human interneuron cells could be delivered using neurosurgical stereotaxis – a method of locating points within the brain – specifically into the regions where we think the pain triggers are in this disease, and to see if those cells will persist with long-term stability in those rodents.

And then finally, if we succeed with the first two, to see whether the human cells can demonstrate disease-modifying activity and significant compelling pain reduction in this rodent model. If we satisfy all those conditions, then I think we really have considerable support and rationale to pursue a clinical trial in humans for treatment of drug-resistant, intractable Trigeminal Neuralgia. And of course we then have to work with the Food and Drug Administration to run a battery of safety and toxicology studies before doing the first trial. But those would be the next steps.

When do you envision (or hope) that cell-replacement treatments for neuropathic pain (including Trigeminal Neuralgia) will be tested on humans and eventually be ready for use in humans?

The first study, with John, is between one and two years long. And then it would probably be another two to three years of working with the F.D.A. to set up the clinical trial.

What are the biggest obstacles that you face in translating your preclinical findings to the clinic?

That’s a really good question. It’s one that is common not just to pain but just about to any disease, especially in the neurological disease space. And that’s quite simply because rodents are quite different from humans in many aspects. There are different anatomies, different neural circuitries. And especially with pain, where you don’t know how an animal such as a rodent is feeling. We use instead, in lieu of an actual communication as to how a rodent is perceiving pain, we use surrogate experiments to measure aversive behavior or hypersensitivities, in terms of how the animals are moving their limbs or responding to a stimulus. Those surrogates are of course never the same thing as asking a person if they’re experiencing pain. So it’s both the anatomy and the ability to be able to communicate.

On top of this, there are many confounding aspects of pain – psychological, emotional – as well as a very high placebo effect. Which makes all of this very difficult to translate from rodents into humans.

The Facial Pain Research Foundation is moving forward with a sense of urgency to find cures for Trigeminal Neuralgia and related neuropathic pain.  Why have you linked your efforts to such a new organization is the field of pain research?

We were looking for ways to apply this stem-cell technology. First of all, pain was an important potential indication for this kind of cellular transplant. But when we heard from the Foundation the stories of people who are suffering with this disease, it really pulls at your heartstrings and you want to do whatever you can to help.

It just seems like pain is an indication that really does possibly lend itself to testing this proposed cellular therapy. Because it’s a disorder that’s focal in nature – there’s a focus of pain in the facial region that can be targeted, so we have a target to deliver the cells, which is one of the criteria we have for applying the cellular therapy. Because the cells are delivered directly into a lesioned area. They’re not delivered systemically. So when we think about diseases that we can address with this technology we think about a disease that has a well-described focus – an area such as the trigeminal ganglion or other aspects near the brain stem where we think the pain trigger exists in Trigeminal Neuralgia.

This is still quite controversial in the field and nobody knows for sure. So because we don’t understand the cause, it gets back to your question about what makes it difficult to translate to humans. That’s another important aspect as to why this is a tough bridge – because we don’t really know yet what causes this disease. Which is true for many diseases. So we do our best to recapitulate using artificial triggers in rodent models.

Every day many people are contacting the Facial Pain Research Foundation asking if it is truly possible to find a cure for Trigeminal Neuralgia. How would you respond to their questions?

Absolutely I think it’s possible. And I think the Foundation is funding some tremendous work. The Foundation has wisely spent its dollars on leading-edge research to understand the causes of the disease, looking very carefully at human genetics. And complementing this basic understanding with attempts at a cure using gene therapies, and cell therapies like ours. So I think there have been very rationally thought-out efforts to try to cure the disease with the technologies we have and the information we have, along with efforts to understand the causes of the disease.

What keeps you awake at night?

It’s always how do we continue to fund this important research so that we can truly try to make a difference and help people who are suffering from this terrible disease. There are just not enough research dollars available from the National Institutes of Health. There are very few means to acquire dollars for cutting edge moon-shot types of ideas. I think this was a perfect opportunity for the Foundation to connect a couple of different research efforts, one on animal models and the other on stem cell technology, with one another to give  it a shot and see if we can see some efficacy in these animal models. And then if we do, the next step will be trying to figure out how to garner the dollars for the next step, to march this toward the first clinical trial in humans. That’s where it gets very difficult, because the dollar amounts go up. But I think the Foundation has really done a terrific job in seeding these efforts to take that important first step. And if we can determine that this can potentially represent a very effective, long-lasting therapy, possibly a cure, then there’ll be more evidence and rationale and support and enthusiasm for this to snowball. But we’re going to need all the stakeholders to come together on this and continue to raise the dollars to keep this going. And that’s always a source of anxiety – how do we get the dollars to investigate the feasibility part of this, which is what we’re talking about with this first project. But then once we demonstrate feasibility, how do we sustain that, to make this a real human-therapy clinical trial for patients who are suffering. That’s the two-fold anxiety that keeps me up. 

Facial Pain Research Foundation

Awards Grant To Dr. Allan Basbaum

The Facial Pain Research Foundation is pleased to have approved a research grant to Dr. Allan Basbaum at the University of California San Francisco. The funding is a continuation of his efforts to find cures for trigeminal neuralgia and related neuropathic pain. His research project is entitled “Novel sensory target genes for novel trigeminal neuralgia pharmacotherapies.” The Trustees of the Facial Pain Research Foundation believe that this research has the potential to make a significant difference in the lives of many in pain.

The approach that Dr. Basbaum is taking differs considerably from genome wide screens that are seeking to identify genes that underlie the etiology of or predispose to TN. The approach will identify genes that can be targeted to regulate the ongoing pain and particularly the paroxysmal pains that are so debilitating to the TN patient. Rather than the traditional therapeutic approach that ameliorates its pain manifestation, identifying such genes is of great interest and could lead to a novel approaches to polypharmacological or even gene therapy approaches to the management of TN.

Dr. Basbaum is a Professor and Chair of the Department of Anatomy at the UCSF School of Medicine. He is considered by many as “the” outstanding researcher in the field of pain. Allan has provided significant leadership in the development of Stem Cell Replacement Therapy for TN and related neuropathic pain. Allan has been an important member of the Facial Pain Research Foundation’s International Consortium of Scientists. He has provided his expertise and support to a number of the FPRF research projects to find the cures.

As Dr. Doug Anderson says about Allan Basbaum working on FPRF projects: “Having a scientist of Dr. Basbaum’s caliber on your team is like having Justin Verlander, Babe Ruth, Derick Jeter and Mickey Mantle join your Little League Team!”

First Report

Fifth Science Meeting At

University Of Florida Orthopaedics & Sports Medicine Institute

Gainesville FL

 Kim Burchiel, M.D., F.A.C.S 

The Facial Pain Research Foundation convened its Fifth Scientific Meeting in Gainesville Florida March 7-8, 2019.  The FPRF has assembled a group of world-class investigators as a consortium to find a cure for Trigeminal Neuralgia.  Clinicians and scientists from the US, England, Canada, and Israel were on site, to lend their perspectives on the state of research on TN.  The agenda was ambitious, but the science and discussions lived up to the promise that we are closing in on the new knowledge that will help us find the cure.  Many new ideas were presented and as is true of high functioning groups, some of the best insights were developed during the dinners and social hours after the formal meeting. 

I will summarize some of the exciting concepts discussed at the scientific sessions, by topic.  I present not so much as a full representation of the work that is currently underway, but as an indication of the high-level science that is being devoted to finding a cure for TN.  The conference commenced with discussions of FPRF funded research, and related investigations, of which I am most familiar.

 Scott Diehl, Ze’ve Seltzer, and Kim Burchiel presented the latest on their project “Genes That Predispose to TN”.

 Kim Burchiel from Oregon Health & Science University discussed new insights into the origins of TN, which relate to different populations of patients with TN. Data is emerging that younger patients, particularly females, develop TN without the requirement for neurovascular compression (NVC) of the nerve.  Older patients do frequently have NVC, and their prognosis from MVD is directly related to the severity of that compression.  The size of the skull compartment that houses the trigeminal nerve (posterior fossa) seems also to be directly related to the incidence of NVC.  This new knowledge will help us direct our genetic search.

Scott Diehl from Rutgers University reviewed the latest analysis of the genetic work on TN. A number of genes are now coming into focus, all of which appear to be relevant to the development, or function, of nerves.  Now that we have identified candidate genes, the next step is to “sequence” these genes to determine if the preliminary findings stand up, and to find the exact mutations in these candidate genes.  The hope is that soon we will be able to determine the function of these genes to either discover or develop new drugs for TN, or even replace defective genes with “gene therapy”. Ze’ve Seltzer from the University of Toronto gave a broad overview of the role of genetics in the development of neuropathic pain, and how findings from his studies on phantom limb pain can provide insights into the genes that predispose to TN. Some preliminary data was presented which suggests that this strategy of comparison of large data sets from patients with neuropathic pain may well provide clues to the origins of TN. 

Allen Basbaum from the University of California, San Francisco discussed his work to identify novel gene targets in pain processing, what he termed the "dark genes.”

Lucia Notterpek from the University of Florida presented her work on dysregulated lipid metabolism as a disease modifier in peripheral neuropathies.

Wolfgang Liedtke from Duke University discussed his work on the Trigeminal nerve root as a rational target for safe and effective treatment of trigeminal nerve pain.

Cory Nichols from Neurona, described the work of his company targeted on developing a human GABAergic interneuron cell therapy to treat refractory epilepsy and neuropathic pain disorders.

The second conference day was devoted to a range of new and innovative potential therapies for TN.

Todd Golde from the University of Florida talked about the possibility of gene therapy for Trigeminal Neuralgia.

Joanna Zakrzewska from the University of London, discussed what happens to our patients with TN over time.

John Neubert from the University of Florida provided an overview of his FPRF project on identifying the neurophysiologic signatures of trigeminal neuralgia pain. 

Mingzhou Ding from the University of Florida related his findings on trigeminal neuralgia and multimodal neuroimaging.

Rob Caudle from the University of Florida presented his findings on a approach to ending neuropathic pain with a combination of substances for blocking pain transmission. A novel and unique approach using old compounds to inhibit neuropathic pain.

Allan Basbaum presented an update on the work of a company he is affiliated with which is working on a genetic “switch” that can potentially selectively turn off pain generation in the trigeminal system, using a benign oral drug.

Mike Iadarola from NIH expanded on the use of a compound to potentially control facial pain and plans to test for ending TN and related neuropathic pain.

Jerry Krbec concluded the day with a discussion on funding opportunities from other sources to support the work of the FPRF.

What I would like to leave you with is the sense of the awe I have for the FPRF, having taken on this mission of finding a cure for TN.  Through hard volunteer work, diligent fund raising, and the assembling of scientific expertise, the FPRF has accomplished something which in my experience is very unique.  The FPRF scientific consortium has become the “place to be” in the neuroscience of TN.  Groundbreaking work is being accomplished, and I came away from this meeting with a sense of optimism that we are close to an understanding of how we may cure TN.  In fact, the most encouraging aspect of this meeting was, to me, how many different avenues of new knowledge are opening up.  I am proud to be part of this work, and am excited to see the next levels of understanding that await!

 Kim Burchiel, M.D., F.A.C.S

 Introduction 

by

Lucia Noterpek Ph.D

Oct 2, 2019 

In 1826, French lawyer and politician Jean Anthelme Brillat-Savarin coined the aphorism: “Tell me what you eat, and I will tell you what you are.” This idiom—boiled down to “you are what you eat”— has been repeated for decades, by parents and peers alike, who remind us to eat healthy. While it is not well understood how peripheral nerves benefit from dietary lipids, laboratory investigations in animal models of neurological disorders indicate that dietary lipid supplementation, or substitution (replacing certain lipids with those that are being studied), can positively impact the nervous system. The beneficial effects of specific lipids have been observed in models of multiple sclerosis and inherited peripheral neuropathies. Which was the subject of our recent study.

Our nerves are protected by a lipid-enriched membrane called myelin, which is fundamental in supporting the proper conduction of electrical impulses in the nervous system. Myelin contains high levels of lipids, including phospholipids and neutral lipids such as cholesterol. Changes in the lipid composition of nerves have been described in neurological diseases. Therefore, as a proof-of concept exploratory study we utilized a commercially available high fat diet (HFD) formulation, to examine the influence of dietary lipids on neuropathy severity in a mouse model, called Trembler J (Zhou et al., 2019). The nerves of these animals have severe deficits in myelin, which lead to pronounced functional effects with age. Mice were assigned at a young age to a control diet (CD) or HFD for 6 weeks, while their nerves were still being myelinated and the myelin deficits just begun to exhibit symptoms. At the end of the study, we examined the peripheral nerves and found the animals on the HFD had healthier nerves, with more myelinated fibers and markers of nerve inflammation and myelin pathology were reduced. While this pilot study did not utilize an optimal healthy lipid-rich diet, it demonstrated to us that dietary lipids have a discernable impact on nerves, which could be optimized to help protect and repair damaged myelin.

Our effort in developing a healthy lipid-enriched diet, or a supplement, to protect and repair myelin is also supported by numerous epidemiological studies that show benefits of good dietary fats on the nervous system, and on the management of chronic pain. Our long-term goal is to develop a food-based lipid supplement that will not only help peripheral nerves, but will also be part of a well-balanced diet. An optimal diet formulation should promote longevity and the maintenance of normal body weight, without increasing the risk for cardiovascular disease.

For further details please see

Zhou Y, Lee S, Bazick, H, Miles J, Fethiere AI, Salihi MOA, Fazio S, Tavori H, Fazio S, Notterpek L. A neutral lipid-enriched diet alleviates peripheral nerve pathology and improves myelination in neuropathic mice. Experimental Neurology. 2019 Nov; 321:113031

Facial Pain Research

Foundation

2018 Research Progress

Compiled and Submitted

April, 2019

By Douglas K. Anderson, Ph.D.

Eminent Scholar Professor and Chair Emeritus Department of Neuroscience

University of Florida College of Medicine

And Director of Research and Trustee

Facial Pain Research Foundation

Trigeminal Neuralgia (TN), is a prime example of neuropathic pain which is pain caused by inmalfunctioning of the nervous system and is considered to be one of the most painful disease conditions known to humans. In addition to the debilitating effects on the lives of those afflicted with neuropathic pain, the cost disorders like TN adds to the U.S. health care system exceeds $100 billion/year. Yet despite the very obvious escalating and pain fueled humanitarian and economic crises, funding targeted at discovering the basic molecular mechanisms underlying neuropathic pain conditions from traditional federal agencies, continues to be inadequate.

The meagerness of funding from these traditional sources has led the growing realization that discovering the root cause of and treatment for TN and other neuropathic pain conditions is going to have to be accomplished with funding from the private sector. To this end, The Facial Pain Research Foundation (FPRF) was created in January, 2011 to provide the critical elements that are necessary and sufficient to study these facial pain conditions. Consequently, the sole mission of the FPRF is “…to establish a well-funded translational (i.e., fundamental discovery to clinical application) research continuum that is dedicated to identifying the mechanisms underlying neuropathic facial pain and to develop novel new therapeutic strategies that will permanently stop the pain of TN and related neuropathic pain syndromes”.

Since its inception a little over eight years ago, the FPRF has raised millions of dollars to support eight separate and distinct research projects. The FPRF uses three fundamental criteria in choosing which proposals are to be considered for funding: (1) the projects are novel and unique; (2) the investigator(s) responsible for each project are among the leading researchers in their respective fields; and (3) there is significant diversification in the research strategies among the projects. It is important to note that these FPRF funds have also served in a multiplier capacity in that some of our investigators have used FPRF support as seed funding to jury to or generate the necessary preliminary data which contributed to these investigators acquiring large grants from the NIH and other foundations and, in some cases, the acquisition of venture capital. Total dollars from other sources that came to our consortium of investigators emanating, all or in part, from FPRF seed funds, is estimated to be somewhere over $40,000,000. The purpose of this yearly update is to briefly summarize these six projects and to highlight the progress that each achieved in 2018.  I have also provided summaries of two new FPRF research projects beginning this Summer 2019.

"Investigating Protein-Lipid Interactions in Peripheral Nerve Myelin"

Lucia Notterpek, Ph.D. (Principal Investigator) Professor and Chair Department of Neuroscience
University of Florida College of Medicine

Progress report (Year 4): May 2018-April 2019

Project 1: Cholesterol homeostasis in peripheral nerve myelin Study staff: Ye Zhou, MS, graduate student

We made excellent progress on this project during the past year, with Ye Zhou the graduate student leading the work in the lab. Because of the great progress, Ye has successfully defended her PhD in March 2019, and will be graduating in May 2019. At the bottom of the progress report, I list publications related to this work that are now in press, or under review.

As described previously in my progress reports, the absence of functional peripheral myelin protein 22 (PMP22) is associated with shortened lifespan in rodents, and severe peripheral nerve myelin abnormalities in several species including humans. Schwann cells and peripheral nerves from PMP22 knockout (KO) mice show deranged cholesterol distribution and aberrant lipid raft morphology, supporting an unrecognized role for PMP22 in cellular lipid metabolism. To examine the mechanisms underlying these abnormalities, we studied Schwann cells and nerves from male and female PMP22 KO mice. Whole-cell current clamp recordings in cultured Schwann cells revealed increased membrane capacitance and decreased membrane resistance in the absence of PMP22, which was consistent with a reduction in membrane cholesterol. Nerves from PMP22-deficient mice contained abnormal lipid droplets, with the levels of proteins involved in cholesterol transport, apolipoprotein E (apoE) and ATP-binding cassette transporter A1 (ABCA1), highly upregulated. Despite the upregulation of ABCA1 and apoE, the absence of PMP22 resulted in reduced localization of the cholesterol transporter at the cell membrane and diminished secretion of apoE-cholesterol. In nerves from normal mice, we identified overlapping distribution of PMP22 and ABCA1 at the Schwann cell plasma membrane. Together, these results reveal a novel role for PMP22 in regulating lipid metabolism and cholesterol trafficking through functional interaction with the cholesterol efflux regulatory protein, ABCA1.

Significance of the described results (lay terms):
Understanding the subcellular events that underlie abnormal myelin formation is critical for advancing therapy development for neuropathies. PMP22 is an essential peripheral myelin protein, as its genetic abnormalities account for approximately 80% of hereditary neuropathies. Here we demonstrated that in the absence of PMP22, the cellular and electrophysiological properties of the Schwann cells plasma membrane are altered, and cholesterol trafficking and lipid homeostasis are perturbed. The molecular mechanisms for these abnormalities involve a functional interplay between PMP22, cholesterol, apoE and the major cholesterol-efflux transporter protein ABCA1. These findings establish a critical role for PMP22 in the maintenance of cholesterol homeostasis in peripheral nerves and provide new knowledge for efforts toward therapy development.

 Project 2: Cholesterol homeostasis in peripheral nerve myelin, with a focus on statins Year 12: May 2018-April 2019

Mohammed Al Salihi, MD, PhD, postdoctoral fellow was the key investigator on this project, but he resigned in March 2019, and returned to training in medicine. I will be hiring a new person shortly.

Statins are lipid-lowering agents that are used in the treatment of dyslipidemia by the inhibition of the HMG-CoA reductase enzyme, which is the rate-limiting step in cellular cholesterol biosynthesis. The influence of statins on the nervous system is controversial, particularly concerning peripheral nerves. Currently it is debated whether statins alleviate or aggravate neuropathic pain, and/or whether they cause the neuropathic pain. Based on our investigations of lipid metabolism in peripheral nerves, and these mentioned clinical observations, we hypothesize that in certain individuals, statins alter the stability of myelin making it susceptible to localized compression-induced demyelination with a subsequent painful neuropathy. Dr. Al Salihi has been testing the effects of statins on Schwann cells and on myelination of neurites in vitro, and has found that statins lead to myelin fragmentation. With Mohammed leaving the lab, we are now repeating these results and expanding the studies to more in depth analyses of myelin and Schwann cells viability. Related to the statin project, Ye is exploring the influence of exogenous cholesterol supplementation on myelinating cultures from heterozygous PMP22 knock out mice, which model compression-induced neuropathy. Quantification of internodal myelin segments and internode numbers indicate beneficial influence of cholesterol supplementation. Together, these studies will help us unravel how intracellularly made cholesterol and exogenously supplied cholesterol impact Schwann cell biology, specifically myelin formation and stability.
In a related project, we examined the influence of a neutral lipid-enriched diet on myelination in neuropathic mice. For our studies we used Trembler J (TrJ) mice that carry a spontaneous mutation in peripheral myelin protein 22 (PMP22) and model early-onset, severe neuropathy. While it is known that lipid metabolism is critical for myelination and myelin maintenance, the impact of a cholesterol-enriched diet on early-onset neuropathies has not been examined. Furthermore, the mechanisms by which dietary lipid supplementation and/or substitution (no increase in overall calorie intake) benefit nerve myelin are unclear. To address these questions, we examined the lipid profile of peripheral nerves from 6-month old TrJ mice and tested the impact of a 6-week long neutral lipid-enriched high-fat diet (HFD) on neuropathy progression in young, newly-weaned mice. Oil Red O staining showed pronounced neutral lipid accumulation in nerves from affected, neuropathic mice, along with elevated levels of key cholesterol and triglyceride transport proteins including apoE, LRP1 and ABCA1, compared with wild type (Wt). In young mice, the short-term HFD intervention increased serum cholesterol levels, without impacting triglycerides, or body and liver weights. Nerves from neuropathic TrJ mice showed improvements in the maintenance of myelinated fibers after the 6-week long dietary intervention. Concomitantly, aberrant Schwann cell proliferation was attenuated, as detected by reduction in mitotic markers and in c-Jun expression. Nerves from HFD fed TrJ mice also contained fewer macrophages, with a normalized count of inflammatory CD11b+ cells. In addition, we detected an increase in neutral lipids in the nerve endoneurium and a trend toward normalization of apoE, LRP1, and ABCA1 expression, after the HFD feeding. Together, these results demonstrate a beneficial influence of a neutral lipid-enriched diet on neuropathy progression in young TrJ mice, and support further work in investigating the potential benefits of dietary lipids on demyelinating neuropathies. In our next set of studies, we plan to further optimize the lipid-enriched diet and examine sensory deficits in the various neuropathic models, with and without dietary intervention.

Related to these studies we have the following manuscripts at various stages of publication:
1. Zhou Y, Lee S, Bazick, H, Miles J, Tavori H, Landreth GE, Fazio S, , Notterpek L. PMP22 regulates cholesterol trafficking and ABCA1-mediated cholesterol efflux. J Neuroscience, 2019 (in press).
2. Zhou Y, Lee S, Bazick, H, Miles J, Fethiere A, AISalihi M, Tavori H, Fazio S, , Notterpek L. “A neutral lipid-enriched diet improves myelination and alleviates peripheral nerve pathology in neuropathic mice. Experimental Neurology (under review)
3. Zhou Y, et al., Steatosis and cholesterol mislocalization in the liver of PMP22-deficient mice (In preparation for Journal of Lipid Research)
4. Zhou Y, et al., PMP22 senses cellular cholesterol homeostasis and regulates cholesterol subcellular trafficking via CRAC domain(In preparation for Cellular Neuroscience)
5. AlSalihi, Bazick et al., HMG-CoA reductase inhibitors and their effect on Rat Schwann cells and myelination (In preparation but still collecting data)

"Mapping Towards a Cure: Identification of Neurophysiologic Signatures of Trigeminal Neuralgia Pain"

Summary:

A University of Florida (UF) team consisting of John K, Neubert, D.D.S.,Ph.D. (Project Leader), Mingzhou Ding, Ph.D., Robert Caudle, Ph.D., Marcelo Febo, Ph.D., and Todd Golde, M.D., Ph.D. are investigating the neurophysiological signature of trigeminal neuralgia (TN) pain and developing promising new therapies. This group represents expertise in facial pain, structural and functional neuroimaging, neurophysiology, molecular biology, pharmacology, and viral vectors. Initially, this project was developed to include both preclinical animal models of trigeminal nerve pain and human studies using imaging studies from TN subjects to investigate the cause of TN. The preclinical animal studies have yielded the novel therapeutic agent, CGRP-Botox (see Dr. Caudle’s separate report). Dr. Febo has provided additional animal imaging analyses and has identified unique connections within the brain following trigeminal nerve injury. These findings confirm unique pathways relevant to trigeminal nerve pain and can provide a foundation for studying humans using the same imaging analyses. The human studies have found critical brain regions important in transmitting pain in TN patients and we identified an exciting relationship relating to neuroinflammation, chronicity, and surgery (see below in Human Studies). These findings provide the foundation for the journal article that is in preparation and will be submitted in the next few months. We have made significant progress towards completing the goals of this project and will expand on these exciting results (see Future Directions) to provide a pathway for finding a cure for TN.

Read more ...