Andy Kuo

In vivo profiling of seven common opioids for antinociception, constipation and respiratory depression: no two opioids have the same profile

For patients experiencing inadequate analgesia and intolerable opioid‐related side effects on one strong opioid analgesic, pain relief with acceptable tolerability is often achieved by rotation to a second strong opioid. These observations suggest subtle pharmacodynamic differences between opioids in vivo. This study in rats was designed to assess differences between opioids in their in vivo profiles.

Validating Eaton's Hypothesis: Cubane as a Benzene Bioisostere

Pharmaceutical and agrochemical discovery programs are under considerable pressure to meet increasing global demand and thus require constant innovation. Classical hydrocarbon scaffolds have long assisted in bringing new molecules to the market place, but an obvious omission is that of the Platonic solid cubane. Eaton, however, suggested that this molecule has the potential to act as a benzene bioisostere. Herein, we report the validation of Eatons hypothesis with cubane derivatives of five molecules that are used clinically or as agrochemicals. Two cubane analogues showed increased bioactivity compared to their benzene counterparts whereas two further analogues displayed equal bioactivity, and the fifth one demonstrated only partial efficacy. Ramifications from this study are best realized by reflecting on the number of bioactive molecules that contain a benzene ring. Substitution with the cubane scaffold where possible could revitalize these systems, and thus expedite much needed lead candidate identification.

Theoretical and practical applications of the intracerebroventricular route for CSF sampling and drug administration in CNS drug discovery research: A mini review

Clinically, central nervous system (CNS) disorders account for more hospitalisations and prolonged care than almost all other diseases combined. In the preclinical setting, the intracerebroventricular (ICV) route for cerebrospinal fluid (CSF) sampling or dose administration in rodent models of human CNS disorders has potential to provide key insight on the pathobiology of these conditions. Low level neuroinflammation is present in >40% of patients with severe depression or schizophrenia and so comparative assessment of CSF composition between patients and rodent models of CNS disorders is potentially invaluable for hypothesis generation and for assessing rodent model validity. As molecules in the CSF have relatively low protein binding and are freely exchanged into the extracellular fluid of the brain parenchyma, supraspinal drug administration into the CSF can produce therapeutic drug concentrations in the brain. Direct administration of investigational agents into the CSF of the lateral ventricle of the brain enables intrinsic efficacy and adverse effect profiles to be evaluated without the confounding effects of drug metabolism, due to the low capacity of the CNS to metabolise exogenous compounds. It is our view that the ICV route for CSF sampling and for administration of novel drugs in development is under-utilised in preclinical research on CNS disorders. This is due to the high degree of technical skill and low margin for error associated with correct ICV guide cannula implantation in the rat. However, these technical challenges can be overcome by using standardised procedures and attention to detail during surgery and in the post-operative period.

The Walker 256 Breast Cancer Cell- Induced Bone Pain Model in Rats

The majority of patients with terminal breast cancer show signs of bone metastasis, the most common cause of pain in cancer. Clinically available drug treatment options for the relief of cancer-associated bone pain are limited due to either inadequate pain relief and/or dose-limiting side-effects. One of the major hurdles in understanding the mechanism by which breast cancer causes pain after metastasis to the bones is the lack of suitable preclinical models. Until the late twentieth century, all animal models of cancer induced bone pain involved systemic injection of cancer cells into animals, which caused severe deterioration of animal health due to widespread metastasis. In this mini-review we have discussed details of a recently developed and highly efficient preclinical model of breast cancer induced bone pain: Walker 256 cancer cell- induced bone pain in rats. The model involves direct localized injection of cancer cells into a single tibia in rats, which avoids widespread metastasis of cancer cells and hence animals maintain good health throughout the experimental period. This model closely mimics the human pathophysiology of breast cancer induced bone pain and has great potential to aid in the process of drug discovery for treating this intractable pain condition.

Comparison of Burrowing and Stimuli-Evoked Pain Behaviors as End-Points in Rat Models of Inflammatory Pain and Peripheral Neuropathic Pain

Establishment and validation of ethologically-relevant, non-evoked behavioral end-points as surrogate measures of spontaneous pain in rodent pain models has been proposed as a means to improve preclinical to clinical research translation in the pain field. Here, we compared the utility of burrowing behavior with hypersensitivity to applied mechanical stimuli for pain assessment in rat models of chronic inflammatory and peripheral neuropathic pain. Briefly, groups of male Sprague-Dawley rats were habituated to the burrowing environment and trained over a 5-day period. Rats that burrowed ≤ 450 g of gravel on any 2 days of the individual training phase were excluded from the study. The remaining rats received either a unilateral intraplantar injection of Freunds complete adjuvant (FCA) or saline, or underwent unilateral chronic constriction injury (CCI) of the sciatic nerve- or sham-surgery. Baseline burrowing behavior and evoked pain behaviors were assessed prior to model induction, and twice-weekly until study completion on day 14. For FCA- and CCI-rats, but not the corresponding groups of sham-rats, evoked mechanical hypersensitivity developed in a temporal manner in the ipsilateral hindpaws. Although burrowing behavior also decreased in a temporal manner for both FCA-and CCI- rats, there was considerable inter-animal variability. By contrast, mechanical hyperalgesia and mechanical allodynia in the ipsilateral hindpaws of FCA- and CCI-rats respectively, exhibited minimal inter-animal variability. Our data collectively show that burrowing behavior is altered in rodent models of chronic inflammatory pain and peripheral neuropathic pain. However, large group sizes are needed to ensure studies are adequately powered due to considerable inter-animal variability.

Progress in understanding mechanisms of opioid-induced gastrointestinal adverse effects and respiratory depression

ABSTRACT Opioids evoke analgesia through activation of opioid receptors (predominantly the &mgr; opioid receptor) in the central nervous system. Opioid receptors are abundant in multiple regions of the central nervous system and the peripheral nervous system including enteric neurons. Opioid‐related adverse effects such as constipation, nausea, and vomiting pose challenges for compliance and continuation of the therapy for chronic pain management. In the post‐operative setting opioid‐induced depression of respiration can be fatal. These critical limitations warrant a better understanding of their underpinning cellular and molecular mechanisms to inform the design of novel opioid analgesic molecules that are devoid of these unwanted side‐effects. Research efforts on opioid receptor signalling in the past decade suggest that differential signalling pathways and downstream molecules preferentially mediate distinct pharmacological effects. Additionally, interaction among opioid receptors and, between opioid receptor and non‐opioid receptors to form signalling complexes shows that opioid‐induced receptor signalling is potentially more complicated than previously thought. This complexity provides an opportunity to identify and probe relationships between selective signalling pathway specificity and in vivo production of opioid‐related adverse effects. In this review, we focus on current knowledge of the mechanisms thought to transduce opioid‐induced gastrointestinal adverse effects (constipation, nausea, vomiting) and respiratory depression. HighlightsReview of currently understood mechanisms of opioid‐related gastrointestinal and respiratory side‐effects.The cellular mechanisms underpinning these opioid related side‐effects are poorly understood.New insight on these mechanisms is opening new frontiers for discovery of novel opioids with improved side effects profile.

Comparative analgesic efficacy of pregabalin administered according to either a prevention protocol or an intervention protocol in rats with cisplatin-induced peripheral neuropathy

Chemotherapy‐induced peripheral neuropathy (CIPN) is a type of peripheral neuropathic pain that may be dose‐limiting in patients administered potentially curative cancer chemotherapy dosing regimens. In cancer survivors, persistent CIPN adversely affects patient quality of life and so adjuvant drugs (anticonvulsants eg pregabalin or antidepressants eg amitriptyline) are recommended for the relief of CIPN. However, most studies in rodent models of CIPN involve administration of single bolus doses of adjuvant drugs to assess pain‐relieving efficacy. Hence this study was designed to assess the efficacy of pregabalin administered to CIPN‐rats according to either a prevention or an intervention protocol. Groups of male Sprague‐Dawley rats received four single intraperitoneal bolus doses of cisplatin at 3 mg/kg at once‐weekly intervals to induce CIPN. For the prevention protocol, oral pregabalin (or vehicle) was administered to CIPN‐rats once‐daily for 21 consecutive days from day 0 to day 20 inclusive. For the intervention protocol, oral pregabalin was administered once‐daily for 21 consecutive days from day 28 to day 48, inclusive. Mechanical allodynia and mechanical hyperalgesia in the bilateral hindpaws were assessed just prior to each dose of cisplatin and at least once weekly until study completion (day 27, prevention protocol; or day 48, intervention protocol). Mechanical allodynia and mechanical hyperalgesia were also determined at the time of peak effect at about 2 hours post pregabalin/vehicle administration, once weekly until study completion. For the prevention protocol in CIPN‐rats, pregabalin alleviated mechanical hyperalgesia but not mechanical allodynia. For the intervention protocol, pregabalin alleviated both mechanical allodynia and mechanical hyperalgesia in the hindpaws.

Optimization and in vivo profiling of a refined rat model of Walker 256 breast cancer cell-induced bone pain using behavioral, radiological, histological, immunohistochemical and pharmacological methods

In the majority of patients with advanced breast cancer, there is metastatic spread to bones resulting in pain. Clinically available drug treatments for alleviation of breast cancer-induced bone pain (BCIBP) often produce inadequate pain relief due to dose-limiting side-effects. A major impediment to the discovery of novel well-tolerated analgesic agents for the relief of pain due to bony metastases is the fact that most cancer-induced bone pain models in rodents relied on the systemic injection of cancer cells, causing widespread formation of cancer metastases and poor general animal health. Herein, we have established an optimized, clinically relevant Wistar Han female rat model of breast cancer induced bone pain which was characterized using behavioral assessments, radiology, histology, immunohistochemistry and pharmacological methods. In this model that is based on unilateral intra-tibial injection (ITI) of Walker 256 carcinoma cells, animals maintained good health for at least 66 days post-ITI. The temporal development of hindpaw hypersensitivity depended on the initial number of Walker 256 cells inoculated in the tibiae. Hindpaw hypersensitivity resolved after approximately 25 days, in the continued presence of bone tumors as evidenced by ex vivo histology, micro-computed tomography scans and immunohistochemical assessments of tibiae. A possible role for the endogenous opioid system as an internal factor mediating the self-resolving nature of BCIBP was identified based upon the observation that naloxone, a non-selective opioid antagonist, caused the re-emergence of hindpaw hypersensitivity. Bolus dose injections of morphine, gabapentin, amitriptyline and meloxicam all alleviated hindpaw hypersensitivity in a dose-dependent manner. This is a first systematic pharmacological profiling of this model by testing standard analgesic drugs from four important diverse classes, which are used to treat cancer induced bone pain in the clinical setting. Our refined rat model more closely mimics the pathophysiology of this condition in humans and hence is well-suited for probing the mechanisms underpinning breast cancer induced bone pain. In addition, the model may be suitable for efficacy profiling of new molecules from drug discovery programs with potential to be developed as novel agents for alleviation of intractable pain associated with disseminated breast cancer induced bony metastases.

Establishment and Characterization of a Novel Rat Model of Mechanical Low Back Pain Using Behavioral, Pharmacologic and Histologic Methods

Chronic low back pain (LBP), the leading cause of disability globally, is notoriously difficult to treat. Most rodent models of LBP mimic lumbar radicular pain rather than mechanical LBP. Here, we describe establishment of a new rat model of mechanical LBP that is devoid of a neuropathic component. Groups of adult male Sprague Dawley rats were anesthetized and their lumbar L4/L5 and L5/L6 intervertebral disks (IVDs) were punctured (0.5 mm outer diameter, 2mm-deep) 5 (LPB-5X), or 10 (LBP-10X) times per disk. Sham-rats underwent similar surgery, but without disk puncture. Baseline noxious pressure hyperalgesia of lumbar axial deep tissues, mechanical allodynia in the hindpaws and gait were assessed prior to surgery and once-weekly until study completion on day 49. The model was also characterized using pharmacologic and histologic methods. Good animal health was maintained for ≥ 49 days post-surgery. For LBP- but not sham-rats, there was temporal development of noxious pressure hyperalgesia in lumbar axial deep tissues at days 14–49 post-surgery. Importantly, there were no between-group differences in von Frey paw withdrawal thresholds or gait parameters until study completion. On day 49, significant histologic changes were observed in the L4/L5 and L5/L6 IVDs for LBP-10X rats, but not sham-rats. In LBP-10X rats, single bolus doses of morphine produced dose-dependent relief of primary and secondary mechanical hyperalgesia in lumbar axial deep tissues at L4/L5 and L1, respectively. In conclusion, our new rat model has considerable potential for providing novel insight on the pathobiology of mechanical LBP and for analgesic efficacy assessment of novel compounds.

The somatostatin receptor-4 agonist J-2156 alleviates mechanical hypersensitivity in a rat model of breast cancer induced bone pain

In the majority of patients with breast cancer in the advanced stages, skeletal metastases are common, which may cause excruciating pain. Currently available drug treatments for relief of breast cancer-induced bone pain (BCIBP) include non-steroidal anti-inflammatory drugs and strong opioid analgesics along with inhibitors of osteoclast activity such as bisphosphonates and monoclonal antibodies such as denosumab. However, these medications often lack efficacy and/or they may produce serious dose-limiting side effects. In the present study, we show that J-2156, a somatostatin receptor type 4 (SST4 receptor) selective agonist, reverses pain-like behaviors in a rat model of BCIBP induced by unilateral intra-tibial injection of Walker 256 breast cancer cells. Following intraperitoneal administration, the ED50 of J-2156 for the relief of mechanical allodynia and mechanical hyperalgesia in the ipsilateral hindpaws was 3.7 and 8.0 mg/kg, respectively. Importantly, the vast majority of somatosensory neurons in the dorsal root ganglia including small diameter C-fibers and medium-large diameter fibers, that play a crucial role in cancer pain hypersensitivities, expressed the SST4 receptor. J-2156 mediated pain relief in BCIBP-rats was confirmed by observations of a reduction in the levels of phosphorylated extracellular signal-regulated kinase (pERK), a protein essential for central sensitization and persistent pain, in the spinal dorsal horn. Our results demonstrate the potential of the SST4 receptor as a pharmacological target for relief of BCIBP and we anticipate the present work to be a starting point for further mechanism-based studies.