Duncan Lascelles

The 2023 Pain in Animals Workshop (PAW) on Sept 26th and 27th is a 2-day workshop that will focus on exploring and discussing three topics: 1. Updates on validated approaches to measuring pain 2. Opportunity areas (biopsychosocial) for additional outcome measure development 3. Analytic approaches in utilizing outcome measures in clinical trials

Osteoarthritis (OA), the most common form of arthritis, affects ~ 27M Americans and is increasing in incidence. It occurs due to degeneration of tissues comprising joints, and is associated with pain. OA pain is a major contributor to the burden of chronic pain in society. Current treatment options are limited to steroid injections, nonsteroidal anti-inflammatory drugs (NSAIDS), opioids and non-pharmacological approaches (exercise, weight loss). Unfortunately, each of these therapeutic approaches are problematic. Exercise, which helps weight management, is difficult for patients due to ongoing pain. NSAIDS can cause gastrointestinal irritation and bleeding and increase risk of heart attack or stroke, and opioids are associated with addiction and abuse (and can actually worsen chronic pain). Clearly, there is a critical need to identify new therapeutic targets and/or treatments for individuals suffering from OA pain. Here, we propose that a heretofore unrecognized neural pathway is a critical component of OA pain. This pathway involves ARTN, its receptor GFRα3, and ‘pain’ channels on nerves (transient receptor potential [TRP] channels). Activation of this pathway initiates and maintains OA pain. The central hypothesis (based on preliminary data in multiple species [mouse, dog, cat, human]) is that ARTN, released from synovium of the OA joint in response to injury, results in de novo increase in its receptor, GFRα3, in local and distant sensory nerves, producing local and widespread pain and hypersensitivity via Proto-oncogene tyrosine-protein kinase receptor (RET)-mediated upregulation of multiple downstream transient receptor potential (TRP) receptors. In this proposal, we will use multiple OA models and clinically relevant outcome measures, and leverage our unique access to dogs with naturally occurring OA, to achieve the following aims: Aim 1: To test the hypothesis that ARTN expression is increased in OA and is responsible for pain. Aim 2: To test the hypothesis that ARTN/GFRα3 signaling is responsible for behaviorally manifested OA pain both in early and late stage disease. Aim 3: To test the hypothesis that RET-dependent ARTN/GFRα3 signaling results in changes in multiple TRP channel expression and activation. Aim 4: To test/validate involvement of the above-described key molecules in a naturally occurring large animal model of OA (dog). Overall, this will be the first work investigating the role and mechanisms of ARTN/GFRα3/TRP channel in OA pain and sensitivity. Based on solid, clinically relevant preliminary data, and leveraging PI expertise from two different and complementary disciplines, successful completion of this proposed work has the potential to identify clinically relevant neural mechanisms leading to the development of novel, effective therapeutics for the treatment of OA-pain in humans."

More than 80% of patients undergoing treatment for head and neck cancer are prescribed opioid pain killers for management of radiation-associated pain, and unfortunately, 6 months after finishing radiotherapy, one-third of head and neck cancer patients will still be opioid-dependent. Chronic opioid use is associated with side effects such as constipation and decreased alertness; it can also lead to opioid abuse, misuse, addiction, and eventual overdose. Safer, more effective and less addictive pain relief strategies are desperately needed, and our proposed research addresses that need by investigating a novel radiation-activated pain signaling pathway that might serve as an excellent target that could be inhibited with new pain medications.

Endurance and marathon sled dog races have an increased risk of orthopedic injuries that are associated with shoulder, carpal, and pelvic limb lameness, swelling, and pain. These injuries are considered the leading cause for dropping (i.e., removing) sled dogs from races (~50.6% of cases) [1, 2]. Further, acute and repetitive traumatic injuries may lead to early removal from a racing event and the development of degenerative joint disease or osteoarthritis (OA) later in life. OA is a major cause of pain and disability in nearly 40% of the canine population and is associated with structural abnormalities and alterations in peripheral and central sensitization. The first case of OA in sled dogs was documented in 1969 based on the British Antarctic Survey findings. No studies since then have investigated the prevalence of OA, potential diagnostic markers, or investigated validated systems for measuring OA associated pain in working sled dogs. We expect the incidence of OA to be quite high due to the physical demands of these canine athletes. Currently, thoracic and pelvic limb joints and muscle groups are evaluated using physical and orthopedic exams and serum biochemical analysis before races and at checkpoints. These exams ensure that the dogs are in good condition but are often subjective and may vary based on the veterinarian’s experience level. Additionally, to avoid suppression of signs of illness or injury, no analgesic oral or topical agents (Eg: NSAID’s-specific doses, opioids, locals, etc.) are approved for the use in sled dogs during races. Failure to identify signs of early joint pain in sled dogs may increase the likelihood of dropping dogs due to orthopedic injuries during the race. Hence, there is an urgent need to develop objective assays to quantify joint pain that would be indicative of OA-associated pain in racing sled dogs. These assays conducted before a race will predict which dogs may be prone to developing orthopedic injuries, will allow the dogs to work longer, will decrease the likelihood of OA pain later in life, and improve the overall quality of life. Currently, no such assays exist that can predict the incidence or outcome of subclinical OA. Thus, the proposed goal of this project is to use an interdisciplinary team science approach to evaluate if artemin (ARTN), a novel neurotrophin, can be used as a potential prognostic biomarker for diagnosing subclinical OA pain.

Over half of dogs in the United States are overweight or obese, and many of these dogs show signs of osteoarthritis that can be corroborated by use of imaging technologies. Overweight dogs, especially those with osteoarthritis are not as likely to walk or run as other dogs, and their sleep may be disturbed by the discomfort they feel in their joints. The purpose of this clinical study is to evaluate the efficacy of an investigational food for aiding in the nutritional management of overweight adult dogs with osteoarthritis. It is expected that at the time of enrollment, all dogs will be overweight and exhibit lameness and pain attributable to chronic osteoarthritis. The study will employ collar-worn activity sensors to generate data that support new, quantitative claims. The primary endpoints of this study are to determine if sensor detected activities (walking, running, resting, sleep quality, and sleep time) are impacted by dietary intervention. The secondary endpoints are the clinical outcomes impacted by dietary intervention and the relationship of those outcomes to the sensor data. The tertiary endpoint of this study is to determine the feasibility of using sensor data to develop algorithms to detect gait changes associated with signs of osteoarthritis.

Millions of Americans suffer acute pain every year, and in the new era of the opioid epidemic, there is an urgent search for safe, effective, non-opioid analgesics. Studies in animals with naturally occurring pain conditions can contribute to the translational research paradigm, accelerating the discovery of safe and effective novel analgesics. It was this central idea that inspired us to plan, organize and hold a workshop on measuring chronic pain in companion animals in 2017 (www.PAW2017.com). The reason for focusing on measurement and animals with naturally occurring conditions was that, 1) measurement of pain in animals is critical for advancing translational research; and 2) naturally occurring conditions have so much in common with certain human pain conditions that the ‘model’ is as close to the human condition one can get. Additionally, the challenges faced with measuring and alleviating pain in animals are similar to those faced in non-verbal human populations. Our meeting in 2017 was extremely successful, with 300 delegates from academia, veterinary and human health, industry and regulatory authorities discussing and brainstorming about how to foster translational pain research by using a cross-species approach. Our proposed 2019 workshop is the logical next step in our theme of measuring pain - focusing on acute pain across multiple species in the context of improving translational research and improving pain control in animals. Our proposed 2019 workshop reflects the innovative approach the MAYDAY fund takes. This workshop is unique – there is no other forum within the pain field where academia, industry and regulatory authorities on the human and animal side come together to discuss and exchange ideas. Like many, we recognize that translational pain research needs innovation, however our innovative workshop platform will provide a fertile environment for game-changing ideas and approaches that will benefit both humans and animals suffering acute pain.

To use quantitative sensory testing (QST) to compare pain in dogs with chronic glaucoma following pharmacologic ciliary body ablation (CBA) and enucleation

Osteoarthritis (OA) affects nearly 100 million Americans, with being the most common form of arthritis and a leading cause of pain and disability. Prevalence and incidence of the disorder are predicted to increase as a result of increased lifespan and obesity. Symptomatic treatments are often ineffective, and/or associated with severe side effects and there is over-reliance on opioids. Recent insurance claim data indicate that >50% of OA were treated with opioids for OA pain, and many OA patients on opioids have become opioid-dependent. Despite the fact that OA is a leading cause of disability, and imposes an annual economic burden exceeding $60 billion, there is no way as yet to cure OA or prevent its progression. There is an urgent need to identify and validate therapeutic targets for pain (symptom) management and to decrease disease progression. Results from our group and others found that a striking feature of OA is the dramatic increase in vascular endothelial growth factor (VEGF) expression and in new blood vessel formation in the joints, both of which correlate with the severity of OA joint pain1. Also, genomic studies revealed that vegf, as an OA marker whose expression is strongly associated with symptomatic OA progression in humans. FDA approval of several inhibitors of the VEGF pathway (i.e., for cancer treatment) has enabled significant advances in the therapy of diseases related to pathological angiogenesis. However, there are multiple VEGF ligands with redundant and compensatory roles4 that may contribute to OA progression and pain. Thus, targeting individual ligands may be less efficacious than targeting multiple ligands. Specifically, VEGF ligands signal via two receptors, VEGFR-1 (known as Flt1) and VEGFR-2 (known as Flk1). We hypothesize that VEGFR-2/Flk1 is primarily responsible for cartilage tissue degeneration during OA progression, while activation of VEGFR-1/Flt1 appears to be the major driver of joint pain transmission. Thus, targeting both Flt1 and Flk1 should simultaneously elicit dual function: (i) immediate symptom alleviation and (ii) gradual cartilage tissue regeneration, and restoring joint function.

Our recent survey found that both veterinarians and members of the public believe that dog breeds differ in their sensitivity to pain. However, whether or not breed differences in pain sensitivity actually exist has never been investigated previously. These beliefs could negatively impact the recognition and treatment of pain in the dog population and result in unnecessary suffering, particularly for dog breeds that are viewed as less sensitive to pain. We believe that dogs have similar pain sensitivity thresholds, regardless of what breed they are, but that perceptions about breed-based differences in pain sensitivity affect ratings of pain and the clinical recognition and treatment of pain. To investigate this, we will use a novel and innovative three-pronged approach. First, we will review records from veterinary hospitals to identify differences in pain scores assigned to dogs of different breeds. Second, we will use a survey to evaluate whether conditions are rated as more or less painful, depending on the breed of dog. Third, we will use non-invasive sensory testing to directly measure pain sensitivity across different breeds, to determine whether breed differences in pain sensitivity actually exist. This work is critical because real differences in sensory thresholds between different breeds may exist; if so, this would lay the foundation for further work to understand genetic differences in pain sensitivity and further understanding of pain in dogs. However, if no differences exist, then the impact of the perception of breed differences must be understood so that we can ensure that dogs of every breed are getting appropriate pain management.

"SPECIFIC AIMS: This project will provide insights into mechanisms of multiple joint osteoarthritis (MJOA) and build the foundation for therapeutic development by developing and analyzing an integrated dataset in both humans and dogs including characterization of intestinal permeability (IP), the microbiome, inflammatory and OA-related biomarkers and development or worsening of MJOA. Osteoarthritis (OA) is highly prevalent, a frequent cause of disability, and lacks highly effective therapies in both humans1, 2 and pet dogs3. Multiple joint OA (MJOA), referring to OA in more than one joint site within an individual, is common in both species4-6. Performing a systematic review that included ~30,000 people, we found a high frequency of MJOA based on most definitions (25-74%)7, 8. MJOA has a significant detrimental effect on sufferers with increased OA burden being associated with increased mortality and disability, poorer health and function7, 9-11. Despite its frequency and significant negative impact, MJOA remains poorly understood7, 8. Given the similar manifestations and therapies available for humans and dogs with MJOA3, pet dogs represent an ideal naturally occurring model of MJOA for study and mutual benefit. It is critical to determine underlying mechanism(s) driving MJOA in order to develop more effective therapies for the millions of human and canine patients afflicted. There are few modifiable risk factors for OA in general, and none specifically for MJOA. Although systemic factors such as soluble biomarkers, genetics and metabolism, are related to the overall body burden of OA (i.e., MJOA) rather than OA at single sites of OA, this is not often taken into account. These and other risk factors may act mechanistically, at least in part, by effects on the gut microbiome12, the impact of which is related to intestinal permeability (IP). Increased IP leading to low-level systemic inflammation has been associated with obesity, diabetes, fatty liver, and inflammatory bowel conditions13, 14. Systemic markers indirectly associated with IP and reflecting increased exposure to microbial products that may promote OA include lipopolysaccharide (LPS) and LPS-binding protein (LBP). IP can also be directly assessed using the lactulose-mannitol dual sugar test (DST, humans), iohexol absorption (dogs) or in gut biopsies. Our group has demonstrated associations between LPS, LBP, disease severity, and clinical signs of knee OA15, 16, likely via LPS-driven low-grade inflammation. Fecal transplant from OA patients into germ-free mice with and without meniscal injury resulted in colonic mucosa bacterial invasion in mice transplanted with OA but not control microbiota, establishing a “gut microbiome-OA connection15.” To further elucidate the role of IP as a mechanism in MJOA, the proposed work will leverage two well-characterized observational human cohorts and an animal study: The JoCoOA, a longitudinal study of over 4000 Black and White men and women aged 45 and older and ongoing since 1991; The Johnston County Health Study (JoCoHS), an actively enrolling cohort (2019-, n~2000) with the same infrastructure as the JoCoOA with the addition of younger (35-70 years) and Hispanic individuals; naturally occurring MJOA in pet dogs17-22. Data from all three cohorts will be used to address the following 3 aims: 1. Determine cross-sectional associations between altered IP, systemic inflammation, and radiographic and symptomatic MJOA in humans and pet dogs by: a) assessing the frequency of increased IP both directly by the lactulose-mannitol DST and indirectly through determination of plasma LPS and LBP, along with assessment of inflammation and the microbiome, in 100 JoCoHS participants with and without MJOA, and b) determining the frequency of increased IP by directly analyzing intestinal biopsies using a 2-dimensional organoid approach and measuring iohexol absorption, and indirectly by measuring LPS and LBP, and characterizing inflammation and the microbiome in a cohort of pet dogs (n=?) with and without MJOA and not receiving c