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2017 Research Progress


April, 2018



Douglas K. Anderson, Ph.D.

Eminent Scholar Professor and Chair Emeritus

Department of Neuroscience

University of Florida College of Medicine


Director of Research and Trustee

Facial Pain Research Foundation


Trigeminal Neuralgia (TN), is a prime example of neuropathic pain which is pain caused by injury to or malfunctioning of the nervous system.  TN is considered to be one of the most painful afflictions known to humans with debilitating effects on the activities of daily living of individuals with this painful condition.  The cost neuropathic conditions like TN adds to the U.S. health care system is greater than $100 billion/year yet funding for research from federal agencies, primarily the National Institutes of Health (NIH) and pharmaceutical companies continues to be woefully inadequate.  The meagerness of funding from 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 six years ago, the FPRF has raised over $4 million dollars to support six 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) all of the projects are not “cut from the same cloth”, i.e., 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 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 these other sources that came to our consortium of investigators emanating, all or in part, from FPRF seed funds, is estimated to be somewhere over $30,000,000.  The purpose of this yearly update is to briefly summarize these six projects and to highlight the progress that each achieved in 2017. 

 “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 3): May 2017-April 2018

Project 1: Cholesterol homeostasis in peripheral nerve myelin

Study staff: Ye Zhou, MS, graduate student

With Ye Zhou, a graduate student in Dr. Notterpek’s laboratory leading the way this group continued to make excellent progress on this project during the past year.  They gave presentations based on the results at several meetings.  Also they have submitted a manuscript titled “Peripheral myelin protein 22 is necessary for ABCA1-mediated cholesterol efflux’’ to PNAS Plus. However this article as written was not accepted for publication.  Consequently they are in the process of adding an additional data set to this manuscript and shortly will submit the revised paper to an alternative high impact journal.

Recent publications in the literature along with data from Dr. Notterpek’s lab continue to indicate that cholesterol homeostasis is critical for myelin stability in both the central (CNS) and peripheral (PNS) nervous systems.  They are focusing on mechanisms that involve cholesterol trafficking within the cells and cholesterol efflux from myelin-forming cells, including the potential exchange of cholesterol between the nerve and Schwann cells. Their studies indicate that the concentration and localization of cholesterol is tightly regulated within myelin forming cells, as in the absence of proteins that are involved in this process, both mice and humans develop neuropathies. They are also working on understanding how the correct amount of cholesterol is maintained in the myelin membrane, which is critical for membrane stability. Peripheral myelin protein 22 (PMP22), whose haploin sufficiency is linked with a compression induced neuropathy (that often is painful), is critical and necessary for assuring cholesterol transport to the membrane. Utilizing neuropathic mice with PMP22 deletion and point mutation, they discovered that PMP22 regulates cholesterol homeostasis and trafficking via the interplay with ABCA1, the major cholesterol efflux transporter. Considering the positive effects of drugs that facilitate cholesterol transport such as 2-hydroxypropyl-β-cyclodextrin (HβCD) in CNS myelination, along with the likely interaction between PMP22 and cholesterol (see below), they treated nerve explants from PMP22-deficient neuropathic mice with HβCD and found improvements in the myelination capacity of the affected cells in response to drug treatment, as compared to untreated samples. These studies will need to be repeated, but are leading us them toward potential therapeutic approaches on how abnormal levels of cholesterol in myelin could be corrected.

In another set of cellular/molecular studies, they have now identified the exact amino acid motif in PMP22 that likely binds cholesterol and are in the process of finalizing this study for publication. Related to this aspect of the project, Dr. Notterpek is in discussion with a membrane biophysicist at Vanderbilt University who has an assay to monitor the incorporation of normal and cholesterol-binding mutated PMP22 into lipid micelles and are working toward coordinating and/or combining their findings with his for publication purposes.  Working with another University of Florida scientist, Dr. Habibeh Khosbouei, they discovered that critical properties of the glial membrane, such as membrane resistance and capacitance, change when the levels of cholesterol are abnormal, e.g., as occurs when PMP22 is deficient.  To the best of their knowledge, no one else has performed similar single cell electrophysiological recordings of Schwann cells causing great excitement in the lab because understanding how abnormal membrane-associated cholesterol levels alter the ability of myelin to protect the axon and guide membrane depolarization (neuronal activity), are critical and fundamental aspects of neurobiology.  Moreover, Dr. Notterpek’s group is continuing their investigation of the impact of dietary cholesterol on peripheral myelin health using mice with different genetic backgrounds, and discovering strong impact of genotype and gender on the response of the animals to a high fat diet.  These studies are at various stages of sample and data analysis which when completed will serve as the foundation for future grant applications to the NIH.

Presentations in poster format

Zhou Y, Lee S, Miles J, Tavori H, Fazio S, Notterpek L. “A novel interaction between PMP22 and ABCA1 in regulating cholesterol trafficking”. This was presented at the 2018 Neurochemistry meeting.

Bazick H, Zhou Y, Salihi Mohammed, Fritz R, Tavori H, Fazio S, Notterpek L. “Investigating the impact of high fat diet-induced dyslipidemia on neuropathic mice.” This will be presented at the 2018 peripheral nerve society meeting

 Budget expenditures:

We have about $50,000 left of the total budget for this project, which I would like to extend into the 4th year to allow for Ye to finish her studies and graduate. Dr. Notterpek’s focus will have to be on getting this project funded from the NIH.

Project 2: Cholesterol homeostasis in peripheral nerve myelin, with a focus on statins

Year 1: May 2017-April 2018

Mohammed Al Salihi, MD, PhD, postdoctoral fellow is key investigator on this project

Myelin, a lipid-rich multilayered membrane made by oligodendrocytes (in CNS) or Schwann cell (in PNS), insulates axons and facilitates the transduction of neuronal signals (1). In the brain, cholesterol contained in myelin accounts for about 80% of total brain cholesterol with approximately 25% of the unesterified cholesterol of an entire adult mouse brain located in myelin, Cholesterol, therefore, plays an important role in the nervous system and is essential for brain and neuronal function (2). Statins are lipid lowering agents that are used in the treatment of dyslipidemia (abnormally elevated cholesterol or fats lipids in the blood) that contributes to the development of atherosclerosis.  Thus, statins are used to reduce the risks of cardiovascular diseases due to atherosclerosis (3).  According to the Centers for Disease Control and Prevention (CDC) and National Center for Health Statistics (NCHS), the usage of statins has increased from 16.3% to 23.2% between the years 2003 to 2012 in adults aged 40 and over, with Simvastatin being the most commonly used cholesterol lowering medication. The primary mechanism of action of statins is the inhibition of the HMG-CoA reductase enzyme, which is the rate limiting step in cholesterol biosynthesis (6). Despite the fact that statins have similar mechanism of action and the same effects on cholesterol profiles, they can be divided into 2 categories: Type 1, which are fungal derived statins, like Lovastatin, Pravastatin and Simvastatin; and Type 2, which include synthetically derived statins like Atorvastatin and Fluvastatin (3). There is a controversy regarding the effect of statins on peripheral nerves. Currently it is debated whether statins alleviate or aggravate neuropathic pain, and/or whether they cause the neuropathic pain. With these largely clinical observations as background, Dr. Nottterpek has developed the hypothesis that in certain individuals, statins alter the stability of myelin making it susceptible to a localized compression-induced demyelination with a subsequent painful neuropathy.

In order to test this hypothesis, they chose statins from each group and are testing their effects on cultured Schwann cells. Their results demonstrated that regarding cellular toxicity, the cells were still viable after 24h exposure to 500 uM of statins, however, morphologically they became rounded and started to lift up from the plate. In collaboration with Dr Khoshbouei’s laboratory, they were able to show that inhibiting cholesterol synthesis by statins altered the Schwann cell membrane electrical resistance, this effect being statistically significant even at low doses such as 1 and 2 μM. These studies are very novel and will likely be controversial.  Nonetheless, this approach allows them to examine the impact of cholesterol lowering drugs on Schwann cell biology at a cellular detail that was previously not possible.

Dr. Al Salihi is also testing the effect of statins on myelination of neurites in vitro and has found that statins lead to the fragmentation of myelin basic protein-positive myelin segments, an effect that is reproducible.  In statin treated cultures, they also detected pronounced changes in the expression of various proteins related to myelination and are now trying to understand the mechanism for these effects. Notably, when HMGCoA is inhibited by statins, some cholesterol transport proteins go down, while the levels of PMP22 robustly increase.  Although at present they do not understand the mechanism for this effect, the data supports our overall hypothesis that PMP22 is critical in the maintenance of lipid homeostasis in myelin-forming glial cells.

As an extension to this work, in the animal models they are “walking back” from the peripheral nerves into the CNS and will begin to examine how the neuronal cell bodies and surrounding CNS microenvironment may sense/respond to ongoing chronic peripheral myelin defects. In summary, Dr. Al Salihi is having had a good start to his postdoctoral training, he learned many of the techniques used in the laboratory and he is continuing to read and learn the literature.

Budget expenditures:

We have used about $110,000 of the total $360,000 budget (with $180,000 from FPRF and $180,000 from MBI over 3 years) for this project. The expenditures include salary and supplies.


  1. Snaidero N, Mobius W, Czopka T, Hekking LH, Mathisen C, Verkleij D, et al. Myelin membrane wrapping of CNS axons by PI(3,4,5)P3-dependent polarized growth at the inner tongue. Cell. 2014;156(1-2):277-90.
  2. Saher G, Stumpf SK. Cholesterol in myelin biogenesis and hypomyelinating disorders. Biochim Biophys Acta. 2015;1851(8):1083-94
  3. McFarland AJ, Anoopkumar-Dukie S, Arora DS, Grant GD, McDermott CM, Perkins AV, et al. Molecular mechanisms underlying the effects of statins in the central nervous system. Int J Mol Sci. 2014;15(11):20607-37
  4. Ovbiagele B, Schwamm LH, Smith EE, Hernandez AF, Olson DM, Pan W, et al. Recent nationwide trends in discharge statin treatment of hospitalized patients with stroke. Stroke. 2010;41(7):1508-13
  5. Qiuping Gu et al. Prescription Cholesterol-lowering Medication Use in Adults Aged 40 and Over: United States, 2003–2012. NCHC Data Brief. 2014. No. 177
  6. Schachter M. Chemical, pharmacokinetic and pharmacodynamic properties of statins: an update. Fundam Clin Pharmacol. 2005;19(1):117-25

 “Mapping Towards a Cure: Identification of Neurophysiologic Signatures of Trigeminal Neuralgia Pain.”

John K. Neubert, D.D.S., Ph.D., University of Florida (Project coordinator, animal modeling)

Mingzhou Ding, Ph.D., University of Florida (Human imaging)

Marcelo Febo, Ph.D., University of Florida (Animal imaging)

Robert M. Caudle, Ph.D., University of Florida (Therapeutics)

Todd Golde, M.D., Ph.D. (Gene Therapy)

This team of outstanding investigators at the University of Florida (UF) Colleges of Dentistry, Medicine, and Engineering and Evelyn F. and William L. McKnight Brain Institute (MBI) under the guidance of Dr. John Neubert continues to focus on identifying the neurophysiological signature of TN pain and providing promising new therapies. This group represents expertise in facial pain, functional imaging, molecular biology, pharmacology, and viral vectors.

This integrated project is investigating the cause of TN in both animals (rats) and humans. The purpose of this translational study is to determine if trigeminal injury models in rats replicates or is very similar to the neurobiology of TN in humans. Using state of the art magnetic resonance (MR) scanners and behavioral testing equipment, the UF team will identify and compare the individual imaging patterns caused by the activation of different areas in the brain and spinal cord in both TN patients and rats. These specific areas of the brain and spinal cord are called “signature centers of activity” that are activated by TN pain and present as targets for the new therapies their team is developing. In 2017, Dr. Neubert’s team made significant progress towards completing the goals of this project.

Identifying trigeminal nerve signaling within brainstem nerve tracts and brain regions is of utmost importance, as this will lead to novel therapies directed to relevant trigeminal targets designed to block TN pain. Dr. Neubert’s team of investigators already has access to existing therapies that they speculate will be successful in blocking TN pain and they evaluated some of these therapeutics this year (see below in Animal Studies). The human studies have identified brain regions important for both the genesis and processing of pain following a TN episode. Importantly, they have found critical brain regions that interact together in a network describe sites, interaction of networks, and inflammation.




TN with surgery

TN without surgery











  1. Recruitment (Table 1)

Twenty-nine TN patients (17 females) and three age and sex matched controls have been recruited and analyzed to date. One patient had surgery and no longer felt pain and was not scanned and 6 new subjects have been scanned but not analyzed yet and are not included in this report.



  1. Brain Activations during Trigeminal Pain

Figure 1. Brain regions activated during trigeminal pain.

Taking pain ratings while doing fMRI scans allow us to uncover brain regions that are activated during trigeminal pain. As shown in Figure 1, there are many regions involved with the processing of pain. The identification of such brain regions serves as the first step toward finding central-nervous system targets for effective therapeutic interventions in TN.

3. Novel MRI Analyses  

We have implemented novel analyses looking at neuroinflammation in the brain and completing machine learning methods for evaluating different structures that may be important for producing pain in TN subjects (Figure 2). The results suggest that structural MRI and diffusion MRI contain different information about the pathology of TN. This is an important as this information may translate into identification of comprehensive therapeutic targets and development of individualized pain therapies based on brain signatures.




  1. Trigeminal Pain and Neuroimaging

Dr. Febo’s group has completed approximately 50 high-resolution fMRI scans of rats with various orofacial pain conditions and treated with different therapeutic agents. The data generated is both huge and complex and presenting all of the data is beyond the scope of this report; as such, we are presenting some of the more relevant and exciting findings. Figure 3 demonstrates that animals with trigeminal nerve injury show increased neural-connectivity in pain sensitive structures within the brain. This is important because we will start comparing the human and rat imaging results to assess overlap of thesebrain regions following trigeminal pain.


Figure 3. Brain connectivity is altered with trigeminal nerve injury.  Nerve injury (CCI) produces increased brain connectivity in the brain.


  1. New trigeminal nerve injury model and novel therapy

 Figure 4. Novel CGRP-BoTox treatment targets trigeminal neurons. The red-stained structures represent the cell bodies of trigeminal neurons that were treated.

To address the issue of relating animal-to-human comparisons, we have improved our methods for inducing trigeminal nerve pain in rats (and mice). We were successfully in producing a trigeminal injury that has some of the same qualities as TN, such as touch sensitivity. This is exciting as it may provide a more robust and stable trigeminal nerve injury model. Dr. Caudle’s group has successfully designed and tested a novel therapy (CGRP-Botox) to specifically target and silence pain neurons. They have completed the in vitro (in cells) work and demonstrated the treatment is specific for sensory (pain) neurons (Figure 4) and we are now testing this treatment in rats. This is significant because if successful for blocking neuropathic pain in the animal studies, then we will continue development as a future therapeutic for TN patients.

 Viral-directed therapies

In a pilot study with Dr. Golde’s group, they have also used a fluorescently-labelled vector to demonstrate that they can specifically target the trigeminal system. This is important because they are confident that they have a strategy for treating trigeminal disorders, a finding they hope will allow them to enter the Phase 2 treatment studies sooner than initially thought.

Limitations. Dr. Neubert’s team continues to have issues regarding subject enrollment and staffing difficulties.  The FPRF has generously provided addition funding to increase enrollment and we have hired a research assistant to coordinate recruitment and have been working with Dr. Michael Pasternak of the FPRF to help spread the word that additional subjects are needed. The EEG & MRI animal studies were unsuccessful. We were not able to overcome EEG hardware failures and reliability was unproven.

Plans for the no cost extension period. Due to subject recruitment and staffing issues, we were unable to complete the proposed studies within the 3-year period. With permission of the FPRF and UF, we extended the project to a no cost extension period (2018-2019) that will allow us to use the remaining funds to complete the project.

  • Recruit and scan 25-30 more TN1 subjects.
  • Evaluate compounds from Dr. Golde’s and Dr. Caudle’s laboratories for pain relief.
  • Dr. Febo’s group will continue to analyze the animal imaging data set.

“The William H. and Leila A. Cilker Genetics Research Program to Find a Cure for Trigeminal Neuralgia”.  (“Finding the Genes that Predispose to Trigeminal Neuralgia”)

Investigators listed alphabetically

Kim J. Burchiel, M.D.

Professor Department of Neurosurgery

Oregon Health and Science University


Marshall Devor, Ph.D Professor and Chair

Department of Cell and Animal Biology

Institute of Life Sciences

Hebrew University of Jerusalem


Scott R. Diehl, Ph.D.*

Professor, Department of Oral Biology, Rutgers School of Dental Medicine

Professor of Health Informatics, Rutgers School of Health Related Professions

Rutgers University


Ze’ev Seltzer, Ph.D.

Professor of Pain Genetics

Faculty of Dentistry

Professor of Physiology

Faculty of Medicine

University of Toronto


*Genetics Project Coordinator


This project asks the question:  Are some individuals genetically predisposed to develop trigeminal neuralgia (TN) pain?  This question emanates from research which shows that although 17% of mature adults have a TN lesion (usually a vascular compression of the trigeminal nerve close to where it exits the brain) only a very small minority (0.01-0.2%) of these individuals actually experience TN pain. The disconnect between lesion and pain is an important clue pointing toward a genetic weakness for TN.  Other research shows that more than one third of patients with TN do not have vascular compression of the nerve, and the rare patients with bilateral TN do not generally have vascular compression of the nerve on both sides!  Further, it appears that younger patients (less than 45 years of age) are four times more likely to be women, than men.  All of these new findings point to the possibility of a genetic predisposition for TN. 

This project was conceived with the assumption that the risk for having classical TN has a genetic basis and the study’s primary objective is identifying genetic mutation(s) (aka, sequence variant(s) or polymorphism(s) associated with the development of TN pain.  To date, the genetic material (DNA) of 348 TN1 patients has been searched at over a million places where people are known to differ from one another in their DNA letters (A, T, G or C). On average, any two people have about five million DNA differences between them. Most differences have no effect on health, but some cause or predispose to disease. The challenge is to find the “needles in a haystack” that cause TN1, hidden among the millions of differences with no relation to TN1. This is done by comparing DNA of TN1 patients (cases) with that of people who do not have TN1 (controls).

Initial results have pointed to several regions in the human genome with promising evidence they harbor genes causing TN1. Some regions include only one gene, but there are 17 genes in the genome region with the strongest statistical support, including 3 genes with biological functions involving nerve degeneration, excitability of neurons or responses to nerve injury. These genes are strong suspects, but to determine which genes actually cause TN1 more genetic data are needed, including a new approach using high speed DNA sequencing, before the initial promising findings are ready for testing in animals or translation to the clinic.

These investigators will first check that the DNA differences observed between TN1 patients and controls aren’t simply due to chance, but occur because mutations in the regions actually cause TN1. To do this, they will repeat the experiment a second time and see if they get the same results. They will analyze a new set of TN1 patients and controls, more than doubling the total sample size. If the same genome regions are again different in TN1 patients compared to controls (like lightning striking in the same place twice), this replication will confirm that TN1 disease-causing genes are indeed located there. It’s very unlikely that all of the initially positive regions will be supported by the new data, but even if only one region is strongly confirmed this will be an extremely important advance, not only for TN1 but for neuropathic pain in general.

Moreover, it is not enough to just find regions of the genome that harbor genes that cause TN1 – it’s also necessary to identify the specific genes and mutations that cause the genes to biologically malfunction. This information is essential for translation to new therapies. Some mutations that cause TN1 may be extremely rare, found only in TN1 patients and their close relatives. To discover additional TN1 genes and TN1-causing mutations, including those that are very rare, the investigators will use the latest technologies for “reading” the DNA sequence of the entire human genome in a select subset of 100 TN1 patients. This expanded data set will enable the investigators to ask several very important questions such as: Do different genes cause TN1 in young patients versus those who experience their first symptoms when older? Are there different genetic effects in males versus females? Do different genes cause TN1 in patients without any vascular compression of their trigeminal nerve compared to patients with nerve damage from years of compression?

The findings obtained thus far suggest that the genetics approach may reveal TN1 clinical subtypes that currently are phenotypically indistinguishable. This has been found in numerous other neurological conditions including the epilepsies and ataxias, where mutations in different genes cause different clinical symptoms. Progress towards a cure (or, more likely, “cures”) will surely be more rapid and have greater chance of success if built on a foundation of knowledge about what is really going on biologically in different TN1 patients rather than forced into a “one size fits all” therapeutic approach. This is a one year project with a target of late Spring 2019 for full completion.

“Cell Replacement Therapy as a Treatment for Injury-Induced Neuropathic Pain”

Allan Basbaum, Ph.D., FRS

Professor and Chair

Department of Anatomy

University of California San Francisco

As described in a previous report, Dr. Basbaum’s laboratory has developed a powerful new approach to reversing the abnormal hyperexcitability that underlies many neuropathic pain conditions including trigeminal neuralgia.  Although there are many explanations for the hyperexcitability that occurs after nerve injury, Dr. Basbaum’s laboratory has focused on the loss of inhibitory controls interposed between the peripheral nerve fiber input and the central nervous system circuits that transmit that input to the brain where pain is perceived.  There is now considerable evidence that the loss of inhibitory control results from a significantly reduced function of a population of nerve cells that synthesize the inhibitory neurotransmitter, GABA.  With these findings confirmed, Dr. Basbaum’s group took on the task of developing a strategy for rebuilding those lost or dysfunctional circuits.  They chose a transplant paradigm and, to date, have documented the survivability and functionality of precursors of GABA nerve cells that have been transplanted into the spinal cord of a mouse model of neuropathic pain created with a hindlimb peripheral nerve injury.  Results reported in a series of publications from Dr. Basbaum’s laboratory established that these nerve cells integrate incredibly well into host spinal cord circuits and restore inhibition, the result of which is a near complete loss of the hyperexcitability of spinal cord neurons and most importantly of the mechanical hypersensitivity that occurs in this neuropathic pain model. 

Building upon these studies, Dr. Basbaum and his team have now turned their attention to a chemotherapy-induced neuropathic pain model.  it is significant that Dr. Basbaum’s group has shown that these GABA precursor cell transplants can also reduce both the mechanical and thermal hypersensitivity that occurs after chemotherapy treatment. 

Dr. Basbaum continues his exciting studies with mouse transplantation of progenitor cells, which have the ability to develop into functioning GABAergic inhibitory nerve cells.  Previously, his laboratory demonstrated that transplantation of these cells into the spinal cord can overcome the mechanical hypersensitivity that occurs in a mouse model of peripheral nerve injury-induced neuropathic pain. Their new studies, in a chemotherapy-induced neuropathic pain model, are built on the scientific literature indicating that dysfunction of inhibitory circuits in the anterior cingulate cortex (ACC) is a major contributor to the affective (aversive), but not the sensory-discriminative features (mechanical hypersensitivity), of the pain experience. As after spinal cord transplants, they found that the transplants integrated into the host cortical circuits and profoundly inhibited host neurons. As predicted, mechanical hypersensitivity, which they believe is processed at the level of the spinal cord, was not reduced. However, the cortical transplants did block the animal’s preference for a pain-relieving drug (gabapentin).  From these findings they conclude that selective and long lasting inhibition of the ACC can reduce the ongoing pain and aversiveness that contribute to neuropathic pain’s most negative influences on quality of life. In ongoing studies, Dr. Basbaum’s laboratory, in collaboration with scientists at Neurona, are evaluating the utility of transplanting human embryonic stem cells that have been modified to take on properties of GABAergic nerve cells. Those studies are critical first steps in the development of an approach to treatment clinical pain conditions.

Neurona Therapeutics

Cory R. Nicholas, PhD Co-Founder and Chief Scientific Officer; Neurona Therapeutics Assistant Professor, Adjunct; University of California, San Francisco

Neurona was conceived in 2008 to realize the therapeutic potential of neuronal cell-based therapy. The rationale for pursuing this translational effort was based on many years of rigorous scientific discovery. In the late 1990’s, co-founders John Rubenstein, MD, PhD, and Arturo Alvarez-Buylla, PhD, were among the first to demonstrate that interneurons, a type of nerve cell, in the cerebral cortex mostly originate from precursor cells in the medial ganglionic eminence (MGE), a transient structure during fetal brain development. In these early studies, MGE cells were isolated from fetal mouse brain and injected into the brains of post-natal mice. In contrast to other neural stem and progenitor cells from neighboring fetal brain regions that remained as large cellular masses post-injection, the MGE cells rapidly dispersed away from the injection site and appeared to seamlessly integrate into juvenile and adult mouse recipient brain tissue. The injected MGE cells appropriately matured into various sub-lineages of cortical interneurons that secreted GABA, the major inhibitory neurotransmitter in the central nervous system. Electrophysiology confirmed these cells to be functional neurons that both received and formed synapses, validating their integration post-transplant. The interneurons persisted long-term following transplantation. Moreover, the transplanted interneurons augmented functional inhibition in the brain regions where they were placed. Therefore, MGE interneuron precursor cells had unique properties that could be advantageous if developed into a transplantation cell-based therapeutic. These cells could potentially provide targeted inhibition to hyper-excitable neural networks in relevant neurological disorders. Rather than acting as a transient point source of GABA or a drug pump, the injected interneuron precursor cells integrated into neural circuitry, possibly allowing for stable repair of the injured nervous system.

All of the studies above were performed with mouse MGE interneuron precursor cells. However, a human cell source is required to facilitate the translation of an interneuron therapeutic for patients. Since the MGE structure disappears before birth and human interneurons in the adult brain are not accessible, they turned to human pluripotent stem cell (hPSC) lines. These hPSC lines can be expanded to meet clinical demand and can be coaxed into becoming virtually any cell type in the body. They developed methods to direct hPSCs into the MGE-type interneuron lineage in petri dish cell cultures. These human interneurons produced in petri dishes were then isolated and provided to collaborators at UCSF for testing in many of the preclinical animal models described above. Encouragingly, Dr. Basbaum found that version 1.0 of the human interneurons also could reduce pain hypersensitivity in the sciatic nerve injury mouse model. Arnold Kriegstein, MD, PhD, in collaboration with another laboratory, next demonstrated that the human interneurons mitigated neuropathic pain and bladder dysfunction following spinal cord injury in mice. Thus they believe that multiple neurological disorders are associated with interneuron dysfunction and/or imbalances in neuronal activity. Because they do not innately regenerate, stem cell technology is required to generate inhibitory interneurons to re-balance the nervous system. Technology to produce human interneurons was developed at UCSF using research-grade stem cell lines, materials, and methods. The company is now tasked with extending this work and developing clinical-grade product candidates. They have assembled an all-star team of 35 researchers from around the world to advance this technology toward the clinic and have made tremendous progress and continue to passionately pursue regenerative neuronal cell-based therapies with the potential to successfully treat patients who have neurological disorders.

Evaluation of adeno-associated virus (AAV) constructs directed at pain pathway targets – a rapid approach for identification of effective therapies for trigeminal neuralgia

Principal Investigators (PIs):

Todd E. Golde, M.D., Ph.D., University of Florida (Therapeutics)

John K. Neubert, D.D.S., Ph.D., University of Florida (Behavioral pain testing)

Other Significant Contributors:

Yona Levites, Ph.D., University of Florida (Therapeutics)

Robert M. Caudle, Ph.D., University of Florida (Behavioral pain testing)

The FPRF is pleased and excited to add Todd Golde,M.D., Ph.D. to the outstanding international group of investigators that comprises the FPRF scientific consortium.  Dr.  Golde is a professor in the Department of Neuroscience at the University of  Florida Colllege of Medicine and Executive Director, University of Florida McKnight Brain Institute. His is recognized as an international thought leader in the general research areas of neurodegeneration and neuroinflamation.  In 2017 Dr. Golde submitted a proposal to the FPRF entitled “Evaluation of adeno-associated virus (AAV) constructs directed at pain pathway targets – a rapid approach for identification of effective therapies for trigeminal neuralgia” The purpose of Dr. Golde’s project is to investigate novel viral-based delivery therapeutics that will be aimed at specific targets within the pain processing network.  He hypothesizes that advances in specific gene-targeting vectors will allow for the rapid identification of potential therapies for the treatment of trigeminal neuralgia (TN).  Importantly, Dr. Golde’s team already has expertise in designing and testing viral-based delivery systems in established pain animal models.  Not only is testing new therapies in animal models absolutely mandatory before considering their use in humans, animal testing of new therapeutic approaches can be used as a way of rapidly screening for therapies with significant efficacy to block and/or cure TN pain.

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 safe and the most efficacious in doing so…initially in animals.  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. 

Since gene therapy is a central character in this program perhaps a review of the fundamentals of gene therapy would be in order to assure that everyone has some understanding of this technology. 

Viruses are simple organisms consisting primarily of viral DNA encased in a protein coat.  In order to make a viral construct that can be used to introduce corrective DNA.                                                                                         

In to animals/humans, it is necessary to first remove the wild-type viral DNA leaving only the protein coat.  Without its wild-type DNA, this viral protein shell can serve as a modified delivery system or a viral vector. Placing exogenous therapeutic genes into the empty protein shell creates what is termed a viral construct.  When these viral constructs are injected into the brain, spinal cord, trigeminal ganglion, etc., the therapeutic genes are released into the interior of cells through existing receptors in the cell membrane because the viral protein shell can alter its structure and lock on to these vacant receptors.  The therapeutic DNA can either integrate with the cell’s native DNA or remain as an independent strip of DNA and either way will produce the desired protein product in the injected tissue.  In short, gene therapy or delivery is the use of a viral construct to put in a copy of a gene that replaces a missing or defective gene; overproduces a protein that promotes cell growth and/or is “protective”, e.g., can control pain; and bioengineer stem cells and other types of cells to produce missing or protective proteins.

Dr. Golde has designed a solid, well thought out strategy for attempting to identify genes that are involved in TN pain.  Based upon the published findings from other laboratories in addition to his own, Dr Golde has chosen six AAV constructs to test that represent a number of different targets within the pain-sensing system. His initial strategy are proposing to attack both excitatory and inhibitory pathways – an inhibition of excitatory signals or an enhancement of inhibitory pathways is expected to produce pain relief.  While the goals and the experimental design used to achieve these goals are major strengths of this application, what excites me about this application is the intended consequence of the very real possibility of using Dr. Golde’s gene therapy expertise and technology to determine if gene(s) identified by our genomics team are causally related to the genesis of TN pain or merely an epiphenomena.  The genes they identify and then replicate to confirm should be genes directly associated with TN pain. Using gene therapy technology, these candidate genes can either be eliminated (knocked out)or added back (knocked in) depending on whether TN pain is caused by excessive excitation or loss of inhibition. Dr. Golde and his team with their considerable expertise in designing and synthesizing AAV constructs for identifying specific genes related to pain processing and then, using accepted animal pain models, determine which of these constructs are the best at limiting TN pain could play an important and pivotal role in moving from the discovery of candidate genes to determining their functional role in the genesis of TN pain.

“Novel Ways to Deliver Compounds That Can Eliminate the Pain of Trigeminal Neuralgia”

Wolfgang Liedtke M.D., Ph.D.


Department of Neurology

Duke University

In 2015, Dr. Wolfgang Liedtke approached the Facial Pain Research Foundation (FPRF) with a proposal to develop novel techniques for direct anatomic targeting of specific components of the trigeminal system. The primary advantage of this approach is with delivery of the drug directly to the lesioned area, only a small fraction of the normal systemic dose of the drug will be needed thereby eliminating the unwanted side effects associated with systemic or whole body delivery of the drug.  While, development of techniques that allow for the direct delivery of therapeutic substances to the diseased or injury site is laudable and needed, often the proposed strategy is highly speculative using materials and delivery systems in new and untested ways, Without an indication in the way of solid preliminary data that the proposed  system will work as advertised, a speculative proposal not supported by positive preliminary findings submitted to federal funding agencies (NIH, VA), drug companies, and venture capitalists would in all likelihood would be DOA.  Thus, in order to get the seed funding needed to acquire important preliminary data demonstrating that either he or his co-investigators have the necessary expertise to perform the proposed experiments and that the approach espoused by Dr. Liedtke stands a good chance of being successful. To this end Dr. Liedtke submitted a proposal for seed funding  that after review, was recommended for funding to the FPRF Board of Trustees BoT who subsequently awarded him a one year $50,000 seed grant to initiate his program.

To develop his very unique and innovative delivery system, Dr Liedtke proposes to combine material science with cell engineering.  This idea of blending a unique porous material to encase specially engineered cellular pumps will provide focused delivery of molecules or drugs that have previously shown the capacity to block or eliminate pain in other models and/or primitive models of TN pain.  Thus, the objective of his approach is to deliver molecules that have displayed the capacity to shut down transmission of the aberrant neuronal signals arising from injured site(s) on the trigeminal system.  These little assemblies are designed to be placed around peripheral trigeminal nerve branches and around the trigeminal nerve root at the point where this root exits the brainstem. The experiments described in this proposal are based on previous work by Dr. Liedtke and collaborators using a specifically-created kind of carbon nanomaterial, i.e., a few-walled carbon nanotubes with high electrical conductance that has been highly purified to the point that it is devoid of any neurotoxic contaminating by-products

Dr. Liedtke also proposed to develop another variant of a topical drug delivery vehicle, i.e., one that will be made of immunologically inert biocompatible polymers designed to hold human fibroblast-like cells. Using gene therapy techniques, Dr. Liedtke proposes to replace the native DNA from a wild-type virus with genes harvested from fibroblasts (which are readily obtainable from skin) that code for pain inhibiting substances like GABA or glycine. This switch in genetic material will should effectively reengineer the fibroblasts turning them into anti-pain generating pumps.  The biocompatible polymer devices will prevent the cells from dispersing but will be made sufficiently porous to allow the pain inhibiting molecules to escape.  Dr. Liedtke’s complete progress reports are on file with the FPRF and are available upon request.

Dr. Liedtke has made good progress in developing his delivery systems given that his starting point was very close to the bottom of the development curve for both device.  His future plans include using these two approaches to deliver substances to the     . cisternal trigeminal nerve root in an attempt to create a valid pre-clinical model of TN which can then be used to obtain the necessary and sufficient pre-clinical data to move these two technologies towards clinical studies.  Finally, Dr. Liedtke has indicated a   desire to leverage these preliminary findings (collected with funding from the FPRF) to acquiring a substantial NIH grant or additional funding.



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