Monday, September 29, 2014

Highlights From the 2014 Annual Scientific Meeting of the American Pain Society: Part 2

More on the latest research presented in Tampa
This is Part 2 of a two-part report on selected talks from the 33rd Annual Scientific Meeting of the American Pain Society (APS), held April 30-May 3, 2014 in Tampa, Florida, US. See also Part 1.

Inflammatory pain: novel mechanisms and mediators
Yuriy Usachev, University of Iowa, Iowa City, US, is examining the role of the complement system, a key component of innate immunity, in inflammatory pain. The idea that the complement system could be important in pain signaling derives support from microarray gene expression studies of pain in rodents in which several complement genes surfaced as statistically significant “hits” (Lacroix-Fralish et al., 2011). A notable recent study implicated C1q, a key molecule in the complement cascade, in persistent inflammatory pain signaling in mice (see PRF related news story).

Usachev is interested in another complement molecule, the proinflammatory complement fragment C5a. Previous research showed that blocking the C5a receptor with an antagonist reduced mechanical allodynia, edema, and skin cytokine levels in the mouse hindpaw incision model (Clark et al., 2006). At the meeting, Usachev reported unpublished data showing that complete Freund’s adjuvant (CFA)-induced thermal hyperalgesia was significantly reduced in C5a receptor knockout mice. Administration of C5a receptor antagonists also decreased CFA-induced hyperalgesia. Meanwhile, intraplantar injection of C5a itself resulted in thermal and mechanical hyperalgesia that could be reversed by a TRPV1 antagonist or by TRPV1 deletion, suggesting that the ion channel may mediate the effects of C5a on pain.

To more precisely delineate the signaling mechanisms by which C5a contributes to inflammatory pain, Usachev has turned his focus to macrophages. Macrophages in skin tissue express the C5a receptor, and macrophage cell lines respond to C5a with calcium transients. Thermal and mechanical hyperalgesia induced by C5a were eliminated in transgenic mice lacking macrophages in the periphery, further implicating these white blood cells in C5a-induced pain hypersensitivity.

Could macrophages communicate with neurons via C5a signaling to cause pain, and if so, how? Here Usachev pointed to nerve growth factor (NGF), which macrophages express. NGF levels in the skin are elevated after C5a injection, and Usachev’s group has found that an NGF-neutralizing antibody reduced C5a-induced thermal hyperalgesia, as did an NGF receptor (Trk) inhibitor.

Based on these preliminary findings, Usachev is proposing a model in which C5a binding to its receptor on macrophages increases NGF release from macrophages, which then sensitizes TRPV1 on neurons, leading to pain sensitivity. Usachev’s work thus provides further evidence that the role of immune system components such as complement factors extends far beyond the immune system into pain signaling, too.

David Clark, Stanford University School of Medicine, Stanford, US, presented his group’s work on the role of epigenetic mechanisms and mediators in inflammatory pain. This research has important clinical relevance for postoperative pain, a significant problem for many patients, especially for those on chronic opioid treatment.

Enzymatic modification of histones, the proteins that package DNA, by histone acetyltransferases (HATs) and histone deacetylases (HDACs) is one key epigenetic mechanism that may contribute to pain states (for a review, see Crow et al., 2013). Clark previously reported that treatment of mice with the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) worsened mechanical hypersensitivity in that model, but did not affect thermal sensitivity (Sun et al., 2013). Conversely, administration of the HAT inhibitor anacardic acid ameliorated mechanical hypersensitivity, also without impacting thermal sensitivity. Using an incision model of hyperalgesic priming, Clark also found that treatment with the HAT inhibitor around the time of incision partially blocked hyperalgesia when the animals were challenged two weeks later with prostaglandin E2, whereas the HDAC inhibitor had no effect.

The results suggested that incision increased histone acetylation levels to cause pain, and further experiments looking at spinal cord tissue after incision indeed revealed increased acetylation of an important histone subtype, H3K9, in dorsal horn neurons; SAHA treatment further increased levels of acetylated H3K9. Clark and colleagues further found that incision increased chemokine signaling involving CXC chemokine receptor 2 (CXCR2) and its ligand, CXCL1, a pathway known to play a role in pain and inflammation; SAHA treatment further increased that signaling. Together, the findings suggested that epigenetic processes affecting CXCL1/CXCR2 signaling could be a target for postoperative pain.

Clark’s latest work, published shortly after the meeting, shows that hyperalgesia induced by chronic opioid use also involves epigenetic alterations of CXCL1/CXCR2 signaling (Sun et al., 2014). Again using the mouse hindpaw incision model, Clark showed that chronic morphine administration, which produced mechanical allodynia and thermal sensitivity pre-incision, and worsened incision-induced mechanical allodynia, also increased expression of CXCL1/CXCR2 in the wound area of the skin (dermal layer), though not in the spinal cord; neutrophils in the wound area were the likely source of CXCL1. Chronic morphine further increased the acetylation of H3K9 observed after incision in dermal neutrophils infiltrating the wound area, and acetylation of the CXCL1 promoter was also elevated. In addition, SAHA had effects on CXCL1 levels in the skin similar to those seen with chronic morphine administration.

The recent results suggest that opioids may regulate CXCL1 expression and function via epigenetic processes in injured tissues. Compounds targeting those processes could have value to dampen postoperative pain, especially for patients taking chronic opioids.

Rheumatoid arthritis: a spinal player in post-inflammatory pain
Historically, pain in rheumatoid arthritis has been attributed to inflammation, but many patients with the condition continue to experience pain despite treatment with disease-modifying antirheumatic drugs (DMARDs) that effectively control inflammation. In a plenary lecture, Camilla Svensson, Karolinska Institute, Stockholm, Sweden, focused on mechanisms that may drive persistent pain in rheumatoid arthritis not only during inflammation but also after inflammation has subsided. A particular highlight was discussion of her work, published shortly after the meeting, on the role of extracellular high mobility group box-1 (HMGB1) protein in arthritis pain (Agalave et al., 2014).

HMGB1 is a damage-associated molecular pattern (DAMP) molecule that helps mediate the response to tissue damage and inflammation (Harris et al., 2012) and has been linked to neuropathic pain, low back pain, bone cancer-induced pain, diabetes-induced pain, and migraine in experimental models (e.g., see Shibasaki et al., 2010). Svensson’s path to HMGB1 stemmed from earlier work in which she and her colleagues used a serum transfer arthritis model in mice where mechanical hypersensitivity persists for several weeks after joint inflammation resolves (so-called “late phase” hypersensitivity; Christianson et al., 2011). The investigators found that spinal deficiency of the innate immune receptor toll-like receptor 4 (TLR4), as well as intrathecal administration of TLR4 antagonists, prevented the development of persistent mechanical hypersensitivity in the animals. Because HMGB1 is an endogenous TLR4 ligand, it seemed plausible that HMGB1 could play a part in TLR4-mediated arthritis pain.

In her recently published research, Svensson used a collagen antibody-induced arthritis (CAIA) model in mice to study HMGB1 function. Immunohistochemistry findings revealed the expression of HMGB1 in spinal cord dorsal horn neurons and glia of naïve animals, and that HMGB1 levels were significantly increased in the spinal cord in CAIA animals. Furthermore, blocking HMGB1 with an intrathecal neutralizing monoclonal antibody or with a recombinant peptide known to block extracellular HMGB1 activities reversed mechanical hypersensitivity during both the inflammatory and late phase in the CAIA mice. The investigators also discovered that only a particular, partially oxidized isoform of HMGB1 (disulfide HMGB1) caused mechanical hypersensitivity when administered intrathecally to naïve mice; it is only the disulfide form that activates TLR4. Furthermore, while intrathecal disulfide HMGB1 caused mechanical hypersensitivity in wild-type mice, as well as in animals missing immune receptors other than TLR4, TLR4 knockout mice did not show the same response, indicating that TLR4 mediated HMGB1-induced hypersensitivity. Finally, as activation of TLR4 is associated with activation of glial cells and production of cytokines, the researchers examined the effects of HMGB1 on glial and cytokine gene expression. Only the disulfide form of HMGB1 induced expression of cytokine and glial genes, further underscoring the importance of that particular isoform in rheumatoid arthritis pain signaling.

Collectively, the results show that disulfide HMGB1 expressed in spinal neurons and glial cells acts through TLR4 to enhance glial activity and drive rheumatoid arthritis-induced pain, even in the absence of inflammation, and that compounds that interfere with this pathway could have salutary effects on pain. Svensson is now investigating whether levels of HMGB1 (either alone or in complex with other factors) are altered in cerebrospinal fluid from patients with rheumatoid arthritis.

Next year’s APS Annual Scientific Meeting will take place in Palm Springs, California. For more information, see the APS website.

Highlights From the 2014 Annual Scientific Meeting of the American Pain Society: Part 1

Recent conference featured diverse slate of talks on pain science and treatment
Pain researchers and clinicians headed to Tampa, Florida, US, for the 33rd Annual Scientific Meeting of the American Pain Society (APS), held April 30-May 3, 2014. A wide range of plenary talks, symposia, award lectures, and poster sessions, delivered by leading experts in the pain field, generated great excitement about the latest directions in pain science and treatment. This two-part report summarizes selected presentations from the meeting. Part 1 is below; see Part 2.

Chronic low back pain: an integrated view
Is chronic low back pain (LBP) a disease of the spine, the connective tissue, or the brain? The consensus of a session on this common and often debilitating condition was that alterations in all three play important pathophysiological roles.

Laura Stone, McGill University, Montreal, Canada, focused on the spine, and the contribution of intervertebral disc degeneration to chronic LBP—disc or joint degeneration is present in 10 to 25 percent of patients with the condition. Animal models in which disc degeneration is induced by injury (e.g., with a needle into the disc) have been developed, but those models do not faithfully mimic the slow, progressive, and age-dependent disc degeneration observed in humans. To better model intervertebral disc degeneration in people, Stone uses mice missing the extracellular matrix protein SPARC (secreted protein, acidic and rich in cysteine). Decreased SPARC levels have been observed in people with disc degeneration (Gruber et al., 2004), and the SPARC-null mice show accelerated, age-dependent disc degeneration as well as behavioral signs of chronic back pain (Millecamps et al., 2011).

Previously, Stone’s group found that mice lacking SPARC exhibited signs of both low back and spine (axial) pain, as well as pain down one or both legs (radiating pain) (Millecamps et al., 2012). More recently, Stone and colleagues showed that SPARC-null mice have age-dependent increases in sensory nerve fiber innervation in and around the discs, compared to control animals (Miyagi et al., 2014). That finding suggests that pathological sensory innervation is one mechanism by which disc degeneration could lead to pain.

At the meeting, Stone presented unpublished data indicating that axial pain and radiating pain could have different underlying mechanisms in the SPARC-null mice. She showed that the degree of histologically assessed disc disruption correlated with axial pain, while reduced disc height was associated with radiating pain. Furthermore, the axial pain could be relieved with morphine, pregabalin, or ibuprofen, suggestive of a “mixed” pain phenotypic profile, whereas radiating pain responded only to pregabalin, indicative of a neuropathic pain profile. That is an important concept, since treatment approaches to chronic LBP, Stone said, should be mechanism based. Stone also reported that an anti-nerve growth factor (NGF) antibody reversed radiating pain, but not axial pain, in SPARC-null animals.

Finally, Stone and colleagues discovered that exercise (voluntary running on a plastic wheel) reversed radiating, but not axial, pain in the SPARC-null mice. The researchers also observed that exercise gave partial protection against disc pathology including loss of disc height, and against increased disc innervation. The findings suggest that exercise has beneficial effects on disc health, but Stone stressed that exercise also has beneficial effects on other tissues, including muscle, tendon, and brain, so the results should be interpreted with caution.

Helene Langevin, Harvard Medical School and Brigham and Women’s Hospital, Boston, US, and University of Vermont College of Medicine, Burlington, US, turned her attention to a new area of interest: the role of connective tissue as a potential pathophysiological contributor to chronic LBP.

Langevin’s previous work established that people with chronic LBP exhibit thicker connective tissue in and around the muscles of the low back compared to pain-free controls (Langevin et al., 2009). To understand the role of such connective tissue changes in chronic LBP, Langevin studies the thoracolumbar fascia (TF), a type of connective tissue that covers the muscles of the back of the trunk. TF consists of layers of dense connective tissue separated by layers of “loose” (termed areolar) connective tissue. It is this loose tissue that allows the dense layers to glide past one another. To examine whether alterations in TF layers contribute to chronic LBP, Langevin employs ultrasound imaging while subjects are placed on a motorized table that moves the trunk. Her research has shown that in control subjects, TF tissue layers move in opposite directions in a fluid, gliding manner, but in patients with chronic LBP, the layers do not exhibit as much gliding—rather, they look stuck together. Shear strain measurements indicated that shear strain within the TF was reduced in patients, compared to controls with no LBP, indicative of possible connective tissue pathology (Langevin et al., 2011).

To better understand the contribution of connective tissue alterations to chronic LBP, Langevin began to study the sensory innervation of that tissue in animals. In 2011, she and her colleagues documented for the first time the presence of calcitonin gene-related peptide (CGRP)-positive sensory nerve fiber endings in the connective tissue of the lower back in normal rats (Corey et al., 2011). Based on their small diameter, the neurons appeared to be Aδ and/or C fibers, suggesting that sensory innervation of connective tissue may contribute to pain perception in chronic LBP.

Could therapeutic interventions that target connective tissue improve chronic LBP? To address that question, Langevin’s group uses a model of connective tissue inflammation in the rat, specifically to study the effects of stretching; clinical trials of low back pain have shown that stretching can have therapeutic benefits, although the mechanisms underlying those beneficial effects have remained unclear. In the model, injection of carrageenan into subcutaneous connective tissue of the low back results in gait changes (such as decreased stride length) and increased mechanical sensitivity in the low back as assessed by von Frey testing (Corey et al., 2012). Langevin found that a stretch treatment that encourages the animals to stretch the full length of their bodies mitigated those alterations. The findings suggest that stretching has beneficial effects, but what about the converse? Does a lack of stretching have adverse consequences? Langevin is now addressing that question by using a pig model in which animals wearing a hobble to restrict back movement are subjected to a connective tissue injury. Early results suggest that movement restriction worsens connective tissue structure in the animals. Langevin is also using this model to study whether stretching can reverse the effects of movement restriction, and if so, what the right dose might be.

In the final talk of the session, David Seminowicz, University of Maryland School of Dentistry, Baltimore, US, reviewed several lines of evidence from imaging studies suggesting that chronic LBP is a disease that affects the brain. First, it has long been known that chronic LBP is associated with altered brain structure and function (Apkarian et al., 2004; Baliki et al., 2008; for a recent review of gray matter changes in chronic pain, see Davis and Moayedi, 2013).

Second, treatment can reverse such structural and functional changes. For example, Seminowicz previously showed that effective surgical treatment or facet joint injections in chronic LBP patients resulted in increased cortical thickness in the dorsolateral prefrontal cortex (DLPFC), which correlated with reductions in pain (Seminowicz et al., 2011). Treatment also normalized altered cognitive task-related DLPFC brain activity observed in the patients before treatment.

More recently, Seminowicz and colleagues showed that cognitive behavioral therapy (CBT) increased gray matter volume and density in a number of brain regions, including the DLPFC, in patients with chronic musculoskeletal pain, including six with LBP (Seminowicz et al., 2013; and see PRF related news story). The changes were associated with decreased pain and decreased pain catastrophizing, two of the goals of CBT. Finally, Seminowicz pointed to his unpublished data showing that surgical treatment or facet joint block in chronic LBP patients could normalize altered structural and functional connectivity in the insula, a brain region that is particularly important for processing emotional aspects of pain.

Finally, data from clinical studies of the transition from acute to chronic LBP also support the idea that chronic LBP affects the brain. A one-year study found that functional connectivity between the nucleus accumbens—a key mesolimbic brain structure—and the medial PFC predicted which patients with subacute back pain would go on to experience chronic back pain (Baliki et al, 2012). A follow-up study showed that in patients with acute or subacute LBP, brain activity representing back pain was limited to nociceptive regions that play a role in acute pain, while in those with chronic LBP (pain for over 10 years), that activity was limited to regions involved with emotional aspects of pain (Hashmi et al., 2013; and see PRF related news story).

Because chronic LBP affects sensory, cognitive, and emotional brain circuits, Seminowicz sees a future where pharmacotherapy and non-pharmacological approaches (including CBT, yoga and other exercise, meditation, hypnosis, and non-invasive brain stimulation) are used together to target those circuits. But the condition is not one that solely affects the brain, as the other talks in the session made clear. Chronic LBP, he thus concluded, is best viewed in an integrative fashion that considers the pathophysiology present not only in the brain, but also in the spine and connective tissue.

Headache pain: a role for dural fibroblasts
In a session on emerging concepts in headache research, Greg Dussor, University of Texas at Dallas, US, described his recently published work demonstrating a role for fibroblasts as important contributors to headache pathophysiology. Fibroblasts, which synthesize the extracellular matrix, are the main cell type in the meninges that surround the brain, but unlike other resident meningeal cells, such as mast cells and macrophages, they had not previously been linked to headache pain.

As one of the only pain-sensitive structures within the skull, it is thought that the meninges contribute to headache pain, but the exact mechanisms have remained unclear. In previous work, Dussor and colleagues showed that afferent nerves innervating the dura mater (the outermost layer of the meninges) express acid-sensing ion channels (ASICs) and are sensitive to pH (Yan et al., 2011). Upon stimulation of the dura mater with low pH, awake rats exhibited cutaneous (facial and hindpaw) allodynia, suggesting a role for low meningeal pH in migraine-related behaviors. Follow-up work implicated ASIC3 as the specific ASIC mediating the effect of low pH on dural afferent signaling (Yan et al., 2013).

Interestingly, immunostaining and PCR experiments revealed the expression of ASIC3 not only in nerve endings within the dura mater, but also in non-neuronal dural fibroblasts. In their most recent study (Wei et al., 2014), Dussor and colleagues discovered that conditioned media from lipopolysaccharide (LPS)-treated dura mater fibroblasts induced hyperexcitability of trigeminal neurons in vitro, as assessed by patch-clamp recordings. Next, using a model of migraine in rats, the investigators found that application of fibroblast conditioned media to the dura mater produced cutaneous allodynia in the animals, as indicated by a decrease in facial and paw withdrawal thresholds in response to tactile stimuli. Further experiments revealed that dural fibroblasts released interleukin-6 (IL-6) in response to LPS stimulation, suggesting a possible mediator of the fibroblast contribution to headache pain—previously, it had been shown that application of IL-6 to the dura mater in rats causes cutaneous allodynia (Yan et al., 2012), and that IL-6 is elevated during migraine attacks in people. Dussor also found that low pH and ATP promoted IL-6 release from dural fibroblasts, and that conditioned media from low pH- and ATP-treated cells produced allodynia.

Dural fibroblasts may respond to low pH and ATP by releasing pro-nociceptive mediators, but what is the source of those conditions within the meninges? Here, Dussor became interested in a possible role for stress and sympathetic efferent signaling—norepinephrine promotes IL-6 release from fibroblasts, and norepinephrine-conditioned media produces allodynia, which made Dussor curious about how such signaling may be involved. Sympathetic activity is known to release norepinephrine, ATP, and protons in the meninges, and when that happens those substances will activate receptors on fibroblasts. Dussor is now undertaking early studies to tie fibroblast activity directly to stress in vivo.

What other agents do fibroblasts release that could cause pain? To what other stimuli can fibroblasts respond? How do ASICs, as well as P2X and norepinephrine receptors (which fibroblasts also express), function to contribute to pain? And can fibroblast-mediated mechanisms be targeted for human headache therapeutics? These are some of the important questions, Dussor said, for future research to address.

For more, see Part 2.

Image credit: American Pain Society

Repurposed Drugs Show Promise to Treat Chemotherapy-Induced Peripheral Neuropathy

Diabetes and multiple sclerosis drugs produce positive results in animal models, are poised for human testing
One of the most common reasons cancer patients stop chemotherapy early is because of a single side effect, chemotherapy-induced peripheral neuropathy (CIPN). Characterized by a gradual destruction of sensory nerves of the extremities, CIPN results in a combination of tingling, numbness, shooting and burning pain, and sensitivity to temperature. There is currently no way to prevent CIPN, but two recent studies demonstrate that FDA-approved drugs already in use for other purposes provide protection against the condition in mouse models. The studies open up a fast track to clinical testing of potential new treatments for a debilitating and ultimately life-threatening complication of cancer therapy.

A two-pronged approach
In searching for the biological underpinnings of chronic pain, researchers have focused on DNA, the proteins and enzymes it encodes, and finally the metabolites those enzymes affect. Recently, data have pointed to the central role of these metabolites, in particular, the pro-inflammatory sphingomyelin/ceramide pathway (see PRF related news story). Over the past five years, Daniela Salvemini, a pharmacologist and physiologist at St. Louis University School of Medicine, Missouri, US, and her team have delineated how sphingosine-1-phosphate (S1P), a derivative of ceramide, activates the S1P receptor type 1 (S1PR1) in dorsal horn neurons in the spinal cord and peripheral sensory neurons, sensitizing them and initiating a cellular cascade that results in neuropathy, neuroinflammation, and pain (for a review, see Salvemini et al., 2013).

Interestingly, production of ceramide is part of the main mechanism of cell death induced by cancer chemotherapeutic agents, providing a possible route to CIPN—in the course of killing tumor cells, a chemotherapeutic agent may also boost production of S1P and activate pain pathways.

For the first time, in a study published May 29 in the Journal of Biological Chemistry, Salvemini and colleagues report that S1P, acting via S1PR1, is critical for triggering symptoms of CIPN in a rat model treated with the taxane paclitaxel, a first-line chemotherapy treatment for breast and other cancers, or oxaliplatin, a treatment for colorectal cancer.

That was exciting because there are several marketed drugs that target S1P action at its receptor. FTY720 (also called fingolimod) was approved by the FDA in 2010 for the treatment of multiple sclerosis (MS) and is a potent functional antagonist of S1PR1. “As soon as we found that S1PR1 is involved, we tested FTY720,” Salvemini said.

In the study, lead author Kali Janes and coworkers found that FTY720 could prevent or reverse the signs of neuropathic pain in the rats without diminishing the anti-cancer properties of paclitaxel or oxaliplatin.

Based on these results, “Clinical evaluation of this compound needs to happen fast,” said Salvemini. “We have a drug that is already used in MS patients for several years now that could be translated for chronic neuropathic pain states, like CIPN.”

By repurposing FTY720, any of its second-generation compounds being tested for MS could be tested for CIPN as well. Additionally, FTY720 itself is being tested as an anti-cancer agent, as it blocks anti-apoptotic actions of S1P. The hope is that while preventing CIPN, clinicians could also enhance the anti-cancer effects of chemotherapeutic agents—a dual-pronged approach (Zhang et al., 2013).

“It’s a finding that can’t be put on the side,” Salvemini said.

Salvemini told PRF her team at St. Louis University will be initiating clinical trials of FTY720 for CIPN specifically. In addition, the group is already conducting rat studies to determine if FTY720 reduces peripheral neuropathy induced by other common chemotherapeutic agents.

More dual action
For cancer patients, hope for CIPN might lie in another familiar drug. Metformin is a widely used, FDA-approved anti-diabetic drug. For type 2 diabetics, it stabilizes blood glucose levels and reduces blood lipid levels.

Cobi Heijnen, a neuroimmunologist at the University of Texas M.D. Anderson Cancer Center, Houston, US, recognized metformin’s potential after Theodore Price and his team, then at the University of Arizona, Tucson, US, demonstrated that metformin prevented neuropathic pain in rodent models of nerve injury (Melemedjian et al., 2013).

In a study published in PLoS One in June, first author Qi-Liang Mao-Ying and coworkers reported that administration of metformin before cisplatin in mice prevented mechanical allodynia, sensory deficits, and loss of peripheral nerve fibers. Metformin also reversed mechanical allodynia induced by paclitaxel.

“Since metformin is the most frequently prescribed drug and is very inexpensive, it could become of great use, provided the drug also works in humans to prevent chemotherapy-induced neuropathy,” said Heijnen.

Metformin impacts mitochondrial respiration, leading to the activation of adenosine monophosphate-activated protein kinase (AMPK), which potentially has a neuroprotective effect in the peripheral nervous system (for more on this topic, see PRF related webinar). Through the same mechanism, metformin itself may also have anti-cancer properties—studies suggest the drug decreases the lifetime risk of cancer in diabetics by 30 percent (Decensi et al., 2010).

Like FTY720, metformin could potentially decrease the dose-limiting side effects of chemotherapy and also act to enhance the anti-tumor activity of chemotherapeutic agents.

“Since metformin is already being tested in clinical trials for its potential anti-cancer effect, it should not be difficult to run additional trials focusing on prevention of CIPN,” said Heijnen.

Cautious optimism
“Having these drugs that have already been given to people minimizes the obstacles for FDA approval,” said Sara Ward, a pharmaceutical scientist at Temple University, Pennsylvania, US. Ward works on new approaches to treating CIPN and recently demonstrated the efficacy of the cannabis derivative cannabidiol in a mouse model of paclitaxel-induced CIPN (Ward et al., 2014).

Although FTY720 and metformin are already FDA approved and show potency in these preclinical studies, “One of the big questions that’s unanswered right now is, Are our animal models of CIPN going to be predictive of something that will work in the clinic?” said Ward.

For more on CIPN and the latest research toward new treatments, see PRF related news story.

Abdul-Kareem Ahmed is a medical student and freelance science writer in Providence, Rhode Island, US.

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New Possibilities for Pain Treatment

7th Annual Pain Therapeutics Summit showcases therapies in development
The annual Pain Therapeutics Summit, organized by Arrowhead Publishers and Conferences and chaired by William Schmidt, NorthStar Consulting, Davis, California, US, brings together investigators from industry and academia to discuss the progress of new pain therapies in preclinical and clinical development (see PRF related coverage of last year’s sessions here and here). For the most recent event, the organizers introduced a new format of two meetings, held September 25-26, 2013, in Boston, US, and November 7-8, 2013, in San Diego, US. The following report summarizes select presentations from the Boston meeting on novel pain targets and drug molecules.

Angiotensin II type 2 receptor antagonist gives first results
Inhibitors of the vasoconstrictive angiotensin II type 1 receptor (AT1) are used widely to treat high blood pressure and heart failure. But researchers have also discovered that another member of the same receptor family, the AT2 receptor, plays important roles in peripheral pain-sensing neurons (for recent results, see Anand et al., 2013). AT2 is not involved in blood pressure regulation, and AT2 receptor antagonists have been shown to be analgesic in a variety of rodent pain models, including a nerve injury model of neuropathic pain (Smith et al., 2013), a prostate cancer-induced bone pain model (Muralidharan et al., 2014), a model of antiretroviral drug-induced polyneuropathy (Smith et al., 2014), and an inflammatory pain model (Chakrabarty et al., 2013). Tom McCarthy of Spinifex Pharmaceuticals, Melbourne, Australia, presented results from a Phase 2 trial of the company’s AT2 inhibitor, EMA401, in patients with post-herpetic neuralgia (PHN; Australian New Zealand Clinical Trials Registry). (Since the meeting, the trial results have been published; see Rice et al., 2014, and the accompanying review by Finnerup and Baastrup, 2014).

The study included 183 patients with PHN whose self-reported average pain intensity at baseline was 6.3 on a 0-10 scale. Patients were randomized to EMA401, taken orally twice a day for four weeks, or to placebo. In the treatment group, average pain scores dropped 2.3 points by the last week of treatment, compared to 1.6 points in the placebo group, for a statistically significant improvement over placebo of 0.69 points. Among patients receiving the drug, 34 percent showed at least 50 percent improvement in pain scores, compared to 19 percent of patients in the placebo group. Overall, McCarthy said that EMA401’s efficacy looked comparable to gabapentin, which is approved by the US Food and Drug Administration to treat PHN. And interestingly, benefit from EMA401 was similar regardless of whether patients were taking other pain medications during the trial. Adverse events were similar between drug and placebo groups and mostly mild; those that appeared more frequently in the drug group included headache and allergic dermatitis.

Spinifex is now testing EMA401 in a Phase 2 open-label study in patients with chemotherapy-induced painful neuropathy (Australian New Zealand Clinical Trials Registry) and is planning clinical trials in a range of chronic pain conditions, including diabetic neuropathy and pain due to osteoarthritis.

Glia-targeted therapy tested in dogs
In another talk, Linda Watkins, University of Colorado, Boulder, US, presented data on a glia-targeted therapy from Xalud Therapeutics, a company she founded and where she is currently chief scientific officer. The company, based in Boulder and San Francisco, US, is developing a non-viral interleukin-10 (IL-10) gene therapy.

Studies in animals have shown that in pathological pain states, activated glia in the spinal cord release proinflammatory products that drive pain and undercut opioid analgesia, and also create dependence and other undesirable effects (see PRF related news stories on morphine-induced neuroinflammation and hyperalgesia). IL-10, an endogenous anti-inflammatory cytokine, broadly counteracts those effects—it is “nature’s own negative feedback on neuroinflammation,” Watkins said. To boost IL-10 levels, Xalud developed XT-101, a non-viral gene delivery product in which IL-10-encoding plasmid DNA is packaged in biodegradable polymer microparticles. Watkins presented animal data showing that a single intrathecal injection of XT-101 reverses mechanical hypersensitivity for up to three months in rodent models of injury- and chemotherapy-induced nerve damage.

Watkins reported that the company has also tested XT-101 in pet dogs with naturally occurring painful conditions. In one dog with intractable neuropathic pain, she showed that a single intrathecal dose of XT-101 produced dramatic, progressive improvement in pain and disability (based on the treating veterinarian’s assessment) that continued for at least nine weeks. And injection of XT-101 directly into the osteoarthritic elbow joint of a dog resulted in a loss of pain and return to normal activity after three weeks, according to the owner’s assessment. To date, 12 dogs have been treated, and Watkins says “every single one” has shown improvement in their pain. “The dog data are so compelling,” she said. “It’s not just rats. It’s not just animal models. It’s real disease, spontaneous pain, quality of life.”

Now, Watkins hopes to replicate that improvement in people. She said Xalud is working toward beginning a Phase 1/2a ascending-dose trial of XT-101 in early 2015 in patients with radiculopathy or peripheral nerve injury-induced pain.

Building a gene delivery platform
Darren Wolfe of PeriphaGen, Pittsburgh, US, spoke about that company’s gene delivery technology for introducing therapeutic genes into peripheral neurons. PeriphaGen (formerly a subsidiary of Diamyd Medical) is developing an engineered herpes simplex virus type 1 (HSV1) for several different potential products, each of which expresses a natural pain-modulating or neurotrophic gene product.

The viral vectors are designed to be administered locally by intradermal injection. HSV1 naturally infects peripheral sensory neurons, and the vectors likewise enter those neurons at their peripheral termini. Experiments in animals show that the neurons transport the vectors back to the nerve body in the ganglia and express the gene there—and, in the case of secreted protein products, release the protein into the dorsal horn. Gene expression continues for weeks. (For a recent review from Wolfe and colleagues, see Goss et al., 2014.)

The first vector tested in people was PGN-202 (formerly called NP2), which expresses preproenkephalin (PENK), a precursor of the endogenous peptide ligands of the δ (delta) opioid receptor. In a previous Phase 1 open-label trial, patients with severe intractable pain from terminal cancer experienced profound pain relief, which lasted throughout the 28-day study for patients who received the highest dose (Fink et al., 2011; But, Wolfe said, a subsequent placebo-controlled Phase 2 study in 30 patients ( failed to show statistically significant pain relief. He said PeriphaGen is interested in trying PGN-202 in other pain conditions. (For earlier coverage of preclinical and Phase 1 results presented by David Fink, University of Michigan, Ann Arbor, US, see PRF related news stories here and here.)

Wolfe also reported that two more vectors are moving toward clinical trials. PeriphaGen plans to test PGN-305, which expresses the GABA-producing enzyme glutamic acid decarboxylase (GAD), in patients with pain from diabetic neuropathy, and to test PGN-703, which expresses neurotrophin-3 (NT-3), as a prophylactic against chemotherapy-induced peripheral nerve damage. A vector expressing the endogenous opioid peptide endomorphin is also in preclinical development.

Targeting TRPV1
Selectively targeting primary afferent neurons was also the focus of a talk by Michael Iadarola, National Institutes of Health (NIH), Bethesda, Maryland, US. Iadarola and his team are working on agents that activate the transient receptor potential channel TRPV1. The channel is the target of multiple therapeutic strategies, including both agonists and antagonists (for a recent review, see Brederson et al., 2013). Paradoxically, while opening TRPV1 temporarily causes pain, keeping the channel open long enough can reduce pain. The prolonged activation of TRPV1 lets in a toxic flood of calcium ions that damage the axons of TRPV1-containing nociceptors or even kills the cells.

To wedge TRPV1 open, Iadarola is using resiniferatoxin (RTX), a highly selective, ultra-potent capsaicin analog and TRPV1 agonist from resin spurge plants. In previous studies in pet dogs with pain from bone cancer, intrathecal RTX produced dramatic improvement in comfort (Brown et al., 2005). The treatment has also been applied to arthritic pain in goats and nerve injury pain in horses. Iadarola and colleagues at the NIH are now performing an open-label Phase 1 trial ( in people with pain from advanced cancer. The trial is ongoing, and it is too early to draw firm conclusions, but so far, Iadarola said, it appears that a single intrathecal injection of RTX can alleviate pain rapidly and greatly boost patients’ quality of life, allowing them to reduce or stop opioid use and become much more active. He said a future clinical trial will test intraganglionic injection for osteosarcoma pain.

In parallel, Iadarola is working on administering RTX at peripheral sites, to treat pain by targeting peripheral nerve terminals. In a proof-of-concept study in pet dogs, Iadarola reported that he and collaborator Dorothy Cimino Brown, University of Pennsylvania School of Veterinary Medicine, Philadelphia, US, found that peripheral treatment can have a strong and lasting effect. Iadarola showed a video of two dogs—one healthy and one with osteoarthritis treated with an RTX injection in the affected joint—and both were so active that the audience had a hard time telling the two apart. Iadarola said the treated dog is still pain free after a year and a half.

As an alternative to RTX, Iadarola and colleagues are pursuing positive allosteric modulators of TRPV1—compounds that do not open the channel by themselves but that help keep it open when it is already activated. Such drugs should selectively target neurons at sites of tissue damage or inflammation where TRPV1 is active. In a study published in 2012, Iadarola and colleagues injected the hind paws of rats with capsaicin to activate TRPV1 and found that co-injecting the allosteric modulator MRS1477 ablated nearby nerve terminals and reduced the animals’ thermal hypersensitivity near the injection site for several days. MRS1477 alone had no effect (Lebovitz et al., 2012). The results hint that MRS1477 could target nerve endings at sites of ongoing pain and selectively inactivate them, although the strategy has not yet been tested in a clinically relevant pain model. Iadarola and his colleagues at the National Center for Advancing Translational Sciences (NCATS) are also screening molecular libraries for additional allosteric modulators of TRPV1.

Trigeminal treatments
For the first time, the Pain Therapeutics Summit included talks on migraine, and Nadia Rupniak of CoLucid Pharmaceuticals kicked off with a presentation on lasmiditan, a 5HT1F subtype-selective serotonin receptor agonist being developed as an acute therapy for migraine. CoLucid, a virtual biotech based in Durham, North Carolina, US, acquired lasmiditan from Eli Lilly and so far has taken the compound through successful Phase 2 testing.

The current frontline treatments for migraine, triptans, activate members of the 5HT1 family of serotonin receptors, particularly vasoconstrictor 5HT1B receptors expressed in vascular smooth muscle. Lasmiditan selectively activates 5HT1F receptors, which are not widely expressed in blood vessels but are expressed on neurons in the trigeminal ganglia and brain.

That non-vascular mechanism of action may be important: Not only do triptans often provide inadequate pain relief, Rupniak said, but—because they constrict blood vessels—the drugs are contraindicated for the many patients who have cardiovascular or cerebrovascular conditions, and their use is further limited in patients with risk factors for undiagnosed cardiovascular disease. Animal studies indicate that lasmiditan does not cause vasoconstriction and instead acts on 5HT1F receptors on trigeminal neurons to inhibit nociceptive neurotransmission (Nelson et al., 2010).

Lasmiditan has shown significant migraine pain relief in two Phase 2 studies in migraine (Ferrari et al., 2010; Färkkilä et al., 2012; and see review by Charles, 2012). Rupniak also reported that the drug has been generally well tolerated in the Phase 1 and 2 program, in which over 700 patients have taken the drug. Side effects have been more pronounced at higher doses, with the most being CNS related, such as dizziness, with no evidence of cardiovascular adverse events. Now, she said, CoLucid is seeking to move the drug into Phase 3 studies.

David Yeomans, Stanford University School of Medicine, California, US, described work on nasal oxytocin as a new therapy for head pain conditions that involve the trigeminal nerves. A peptide hormone, oxytocin has been used therapeutically for 60 years, administered intravenously to induce labor or intranasally to promote lactation. Nasal oxytocin is no longer approved in most countries for lactation, Yeomans said, because of a lack of efficacy. But there is abundant evidence that oxytocin does relieve pain (for recent results in rodent models, see Juif et al., 2013, and de Araujo et al., 2014). And with its long history of widespread clinical use, Yeomans said, “It’s about as safe as you can get.” In 2007, Yeomans founded a company, Trigemina, to repurpose nasal oxytocin to treat chronic migraine, trigeminal neuralgia, and other forms of intractable chronic head pain.

The trigeminal neurons that signal pain in the face and head express oxytocin receptors, especially during painful inflammation. As a result, Yeomans reported, oxytocin has proven to be highly analgesic in rat models of head pain and inflammation. Nasal administration seems to be uniquely suited to deliver the peptide to the trigeminal system: Oxytocin sprayed into the nose gets picked up by the nerve terminals, transported back to the trigeminal ganglia, and then becomes distributed through the entire trigeminal system—“suggesting we have the potential to affect pain pretty much anywhere in the head,” Yeomans said.

Yeomans’ clinical colleagues have tested that theory by giving nasal oxytocin off-label to patients with a variety of intractable head pain conditions, and in 22 of 27 cases, he said, patients experienced complete or near complete relief.

Based on that anecdotal evidence, Trigemina initiated clinical trials with a nasal oxytocin formulation, TI-001. The first two trials, in episodic migraine, yielded results that were good but not dramatic, Yeomans said. But based on the evidence that oxytocin receptors are upregulated in inflammatory conditions, he and his colleagues looked at chronic migraine, which is thought to involve neurogenic inflammation. A controlled trial in 40 chronic migraineurs bore out that idea: By two hours after treatment, oxytocin showed a significant effect on pain compared to placebo. (Results from the trial were first presented at the June 2013 International Headache Congress.) Four hours after treatment, 27 percent of patients on oxytocin rated their pain at 0, compared to none of the patients on placebo. And a post-hoc analysis produced the interesting observation that the drug’s effects were stronger in patients who had not taken a non-steroidal anti-inflammatory drug (NSAID) within 24 hours—which echoed preclinical evidence that NSAIDs block oxytocin receptor expression. A larger Phase 2 trial for chronic migraine, in which patients receive TI-001 for eight weeks, is underway (

Yeomans is also keen to test nasal oxytocin in patients with trigeminal neuralgia, a condition notorious for its excruciating pain and lack of effective treatment. In a trigeminal root compression model in rats, TI-001 brought the animals’ mechanical sensitivity back almost to baseline, and Yeomans said that, in a small number of patients with trigeminal neuralgia who were treated with TI-001 in an open trial, all experienced pain relief.

Latching onto lipids
Roger Sabbadini, Lpath, San Diego, California, US, described a drug development effort focused not on proteins, but on the pain-promoting lipid lysophosphatidic acid (LPA). Lpath creates monoclonal antibodies that specifically and tightly latch onto and neutralize bioactive lipids. Lpath already has two antibodies targeting sphingosine-1-phosphate in Phase 2 clinical trials for cancer and macular degeneration. Sabbadini described their work on an anti-LPA antibody, Lpathomab, as a potential treatment for neuropathic pain.

LPA promotes neuronal damage and pain by acting via specific G protein­-coupled receptors in dorsal root ganglia (DRG) sensory neurons. In addition, a recent study showed that LPA may produce pain by directly activating TRPV1 (see PRF related news story). Given the multiple receptors through which LPA acts, Sabbadini said that targeting LPA itself, rather than its receptors, is the better strategy for stopping painful signaling.

Sabbadini reported unpublished data from collaborators showing activity of the antibody in several pain models in animals. In the partial sciatic nerve ligation model of neuropathic pain in mice, intrathecal injection of the antibody achieved days-long reduction of thermal and mechanical pain hypersensitivity. In streptozotocin (STZ)-induced diabetic neuropathy in rats, where LPA levels are increased in small-diameter DRG neurons, a single intravenous injection of the antibody ameliorated mechanical allodynia; chronic administration brought the animals’ sensitivity almost to normal, and allodynia returned when the treatment was stopped. The antibody also showed near complete, durable relief of mechanical allodynia in the Zucker diabetic fatty rat model of type 2 diabetes. And in the collagen antibody-induced arthritis (CAIA) model of rheumatoid arthritis, subcutaneous injection of Lpathomab relieved mechanical allodynia and reversed upregulation of pain-associated protein markers in the spinal dorsal horn.

Lpath is now manufacturing the antibody and beginning toxicity studies, Sabbadini said, in anticipation of clinical trials.

Finally, an old target made new
Not all talks covered such novel targets: Gavril Pasternak, Memorial Sloan-Kettering Cancer Center, New York, US, made the case for a still untapped potential in the opioid receptors. Each opioid receptor gene produces not one receptor, but a plethora of different splice variants that may have different downstream signaling properties. If pharmacologists could identify and target just those variants involved in modulating pain, they might effect analgesia without side effects. Pasternak focused on a set of μ (mu) opioid receptor (MOR) variants that lack the N-terminal transmembrane (TM) domain, producing receptors with only six (rather than the usual seven) TM segments. (For results on other splice variants from Pasternak and others, see PRF related news story.)

Pasternak’s lab has found, in experiments using knockout mice in which the different splice products are selectively deleted, that the MOR agonist IBNtxA (3-iodobenzoyl-6β-naltrexamide) acts on the six-transmembrane-domain (6TM) receptors. Morphine, on the other hand, acts only at the full-length receptor. Interestingly, IBNtxA is 10 times more potent than morphine, Pasternak said, and has performed well in animal models of neuropathic and inflammatory pain. Yet unlike morphine, IBNtxA does not cause respiratory depression in rodents, he said, and causes less constipation than morphine. Moreover, IBNtxA does not create physical dependence and is not rewarding in mice (Majumdar et al., 2011). “This is an interesting pharmacological profile,” he said. Encouragingly, he and his lab have found evidence that some opioid drugs that are used clinically and are well tolerated, such as buprenorphine, act on 6TM receptors—suggesting that targeting the truncated receptors might work in clinical practice.

How IBNtxA targets 6TM receptors remains unclear, but cell culture studies from Pasternak’s lab hint that the compound binds heterodimers of 6TM and full-length receptors. To dig further into the pharmacology, Pasternak said his lab is now synthesizing compounds with increased activity and selectivity for 6TM receptors.

As the meeting wound down, the participants were left with a number of new therapeutic possibilities to ponder. Which of the new drugs or mechanisms do you find interesting, or which have caveats that need to be addressed? Share your thoughts by commenting below.