INTRODUCTION:The opioid crisis burdens the health care system and poses major research challenges. One approach toward safer analgesics involves targeting opioid receptors in areas of tissue injury/inflammation. Increased proton concentrations have been successfully exploited for developing novel ligands with limited side effects. This review investigates the impact of other inflammatory components (free radicals) on opioid receptors, focusing on redox-sensitive thiols and disulfides. AREAS COVERED:This narrative review is based on a systematic literature search up to 3 March 2025. Of the identified 938 articles, 29 articles met the authors' inclusion criteria. Risk of bias was assessed according to NINDS recommendations, and the review followed PRISMA guidelines. As the included studies indicate that disulfides are an essential structural component, it is conceivable that free radicals affect opioid receptor activity. EXPERT OPINION:Current drug design has largely overlooked that function of G-protein coupled receptors (GPCRs) can differ between healthy and pathological microenvironments. The success of pH-dependent opioid ligands illustrates the therapeutic potential of exploiting such conditions. Redox changes may represent another regulatory component, but functional and in vivo evidence is still lacking. The consideration of microenvironmental factors may enable the development of safer, peripherally acting analgesics and refine GPCR-targeted drug design.

BACKGROUND:Diagnosing cLBP is complex due to its heterogeneity, lack of definitive biomarkers, and the subjective nature of pain. This study aims to analyze cLBP patients comprehensively, characterizing demographic and clinical profiles of patients and evaluating the types and effectiveness of previous treatments. METHODS:From January 2022 to April 2024, we recruited 1262 participants aged between 18 and 72 years from the general population by advertisements and word-of-mouth propaganda. Collected data included a detailed medical history, standardized questionnaires (e.g., Von Korff) and a clinical examination. RESULTS:Our population included 471 (38%) with chronic back pain, 335 (27%) without back pain, and an additional group in between of 327 (26%) with intermittent back pain. The majority of participants experienced multifocal back pain, which was further subdivided based on the predominant pain localization. In patients with localized cLBP (median: 5 years, IQR: 1-10 years), the pain persisted for a shorter time than in the group with predominant back pain other than the lower back (median: 10 years, IQR: 5-15 years). The majority of participants did either use no pain medication at all (cLBP: 32%; iLBP: 37%; no-BP(2): 55%) or on-demand medication (cLBP: 56%; iLBP: 56%; no-BP(2): 39%). CONCLUSION:Our data show considerable heterogeneity underlying the widespread diagnosis "chronic back pain." Quantitative differentiation is difficult due to the low pain intensity on the day of the examination, and adequate treatment recommendations are challenging. To better understand chronic back pain, there is a strong need for a subclassification. TRIAL REGISTRATION:German Clinical Trial Register: DRKS-ID: DRKS00027907.

G-protein-coupled receptors are integral membrane proteins that transduce chemical signals from the extracellular matrix into the cell. Traditional drug design has considered ligand-receptor interactions only under normal conditions. However, studies on opioids indicate that such interactions are very different in diseased tissues. In such microenvironments, protons play an important role in structural and functional alterations of both ligands and receptors. The pertinent literature strongly suggests that future drug design should take these aspects into account in order to reduce adverse side effects while preserving desired effects of novel compounds.

Revolutionizing pain medication: Preventing addiction and side effects Stephen Vanner, president of pHarm Therapeutics Inc., and Christoph Stein, inventor and co-founder, discuss a new class of analgesics for pain caused by tissue injury and inflammation. Severe pain resulting from major tissue injury and inflammation, such as arthritis, trauma, surgery, cancer, migraines, sickle cell crisis, and inflammatory bowel disease, can often only be treated effectively with conventional opioid medication. However, the risks of addiction and serious side effects, such as life-threatening respiratory arrest, have led to an opioid crisis in North America and worldwide. The 2018 USA Federal Drug Administration roadmap to combat this crisis states that ‘transitioning from the current market, dominated by conventional opioids, to one in which opioids have abuse-deterrent properties, holds significant promise for a meaningful public health benefit.’ However, so far, competitive approaches, including biased opioid agonists, have failed.

In our previous studies, a new opioid (NFEPP) was developed to only selectively bind to the μ-opoid receptor (MOR) in inflamed tissue and thus avoid the severe side effects of fentanyl. We know that NFEPP has a reduced binding affinity to MOR in healthy tissue. Inspired by the modelling and simulations performed by Sutcliffe et al., we present our own results of coarse-grained molecular dynamics simulations of fentanyl and NFEPP with regards to their interaction with the μ-opioid receptor embedded within the lipid cell membrane. For technical reasons, we have slightly modified Sutcliffe’s parametrisation of opioids. The pH-dependent opioid simulations are of interest because while fentanyl is protonated at the physiological pH, NFEPP is deprotonated due to its lower pKa value than that of fentanyl. Here, we analyse for the first time whether pH changes have an effect on the dynamical behaviour of NFEPP when it is inside the cell membrane. Besides these changes, our analysis shows a possible alternative interaction of NFEPP at pH 7.4 outside the binding region of the MOR. The interaction potential of NFEPP with MOR is also depicted by analysing the provided statistical molecular dynamics simulations with the aid of an eigenvector analysis of a transition rate matrix. In our modelling, we see differences in the XY-diffusion profiles of NFEPP compared with fentanyl in the cell membrane.

N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide is a newly-designed pain killer selectively activating G-protein-coupled mu-opioid receptors (MOR) in acidic injured tissues, and therefore devoid of central side effects which are typically elicited at normal pH values in healthy tissues. However, the neuronal mechanisms underlying NFEPP's antinociceptive effects were not examined in detail so far. Voltage-dependent Ca2+ channels (VDCCs) in nociceptive neurons play a major role in the generation and inhibition of pain. In this study, we focused on the effects of NFEPP on calcium currents in rat dorsal root ganglion (DRG) neurons. The inhibitory role of the G-protein subunits Gi/o and Gβγ on VDCCs was investigated using the blockers pertussis toxin and gallein, respectively. GTPγS binding, calcium signals and MOR phosphorylation were also investigated. All experiments were performed at acidic and normal pH values using NFEPP in comparison to the conventional opioid agonist fentanyl. At low pH, NFEPP produced more efficient G-protein activation in transfected HEK293 cells and significantly reduced VDCCs in depolarized DRG neurons. The latter effect was mediated by Gβγ subunits, and NFEPP-mediated MOR phosphorylation was pH-dependent. Fentanyl's responses were not affected by pH changes. Our data indicate that NFEPP-induced MOR signaling is more effective at low pH and that the inhibition of calcium channels in DRG neurons underlies NFEPP's antinociceptive actions.

AbstractTargeting the acidified inflammatory microenvironment with pH-sensitive opioids is a novel approach for managing visceral pain while mitigating side effects. The analgesic efficacy of pH-dependent opioids has not been studied during the evolution of inflammation, where fluctuating tissue pH and repeated therapeutic dosing could influence analgesia and side effects. Whether pH-dependent opioids can inhibit human nociceptors during extracellular acidification is unexplored. We studied the analgesic efficacy and side-effect profile of a pH-sensitive fentanyl analog, (±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide (NFEPP), during the evolution of colitis induced in mice with dextran sulphate sodium. Colitis was characterized by granulocyte infiltration, histological damage, and acidification of the mucosa and submucosa at sites of immune cell infiltration. Changes in nociception were determined by measuring visceromotor responses to noxious colorectal distension in conscious mice. Repeated doses of NFEPP inhibited nociception throughout the course of disease, with maximal efficacy at the peak of inflammation. Fentanyl was antinociceptive regardless of the stage of inflammation. Fentanyl inhibited gastrointestinal transit, blocked defaecation, and induced hypoxemia, whereas NFEPP had no such side effects. In proof-of-principle experiments, NFEPP inhibited mechanically provoked activation of human colonic nociceptors under acidic conditions mimicking the inflamed state. Thus, NFEPP provides analgesia throughout the evolution of colitis with maximal activity at peak inflammation. The actions of NFEPP are restricted to acidified layers of the colon, without common side effects in normal tissues.N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide could provide safe and effective analgesia during acute colitis, such as flares of ulcerative colitis.

We previously reported the successful design, synthesis and testing of the prototype opioid painkiller NFEPP that does not elicit adverse side effects. The design process of NFEPP was based on mathematical modelling of extracellular interactions between G-protein coupled receptors (GPCRs) and ligands, recognizing that GPCRs function differently under pathological versus healthy conditions. We now present an additional and novel stochastic model of GPCR function that includes intracellular dissociation of G-protein subunits and modulation of plasma membrane calcium channels and their dependence on parameters of inflamed and healthy tissue (pH, radicals). The model is validated against in vitro experimental data for the ligands NFEPP and fentanyl at different pH values and radical concentrations. We observe markedly reduced binding affinity and calcium channel inhibition for NFEPP at normal pH compared to lower pH, in contrast to the effect of fentanyl. For increasing radical concentrations, we find enhanced constitutive G-protein activation but reduced ligand binding affinity. Assessing the different effects, the results suggest that, compared to radicals, low pH is a more important determinant of overall GPCR function in an inflamed environment. Future drug design efforts should take this into account.

This presentation will review basic and clinical concepts on pain and mechanisms underlying opioid analgesia. It will describe the structure, function, and cellular signaling of opioid receptors; endogenous and exogenous opioid receptor ligands; as well as central and peripheral sites of opioid actions. The presentation will also discuss novel opioid-based therapeutic strategies, developed from recently gained knowledge on opioid receptor structures and signaling, pathological pain situations, and in vivo studies, aimed at the reduction of side effects such as respiratory depression, constipation, addiction, sedation and tolerance. Lastly, clinical problems associated with opioid use will be addressed.

BackgroundPatients with chronic inflammatory arthritis (e.g. rheumatoid arthritis; RA) or inflammatory exacerbations of chronic degenerative joint diseases (e.g. osteoarthritis; OA) suffer from recurrent pain, restricted function and reduction of daily activities. The current standard of intraarticular (i.a.) therapy is the injection of steroids, which can increase risk of infection, cartilage degenerations, and other well-known systemic side effects. A novel approach without such complications could be the activation of peripheral opioid receptors, e.g. by i.a. application of small, systemically inactive doses of morphine.ObjectivesThe aim of this randomized placebo-and active drug controlled double blind trial was to investigate reduction of pain in chronic knee arthritis patients following i.a. injections of morphine, a standard steroid (triamcinolone), or placebo. The primary hypothesis was that i.a. morphine results in significantly lower pain scores than placebo. The primary outcome parameter was reduction of the Visual Analogue Scale (VAS) pain at day 7.MethodsAdult patients with chronic knee arthritis because of osteoarthritis (OA) or inflammatory arthritis (IA, rheumatoid arthritis, psoriatic arthritis, spondyloarthritis, oligoarthritis or monarthritis) and a high level of pain (VAS pain ≥ 4 out of 10) at baseline received a single dose of either morphine 3 mg i.a., or triamcinolone 40 mg i.a., or placebo (NaCl 0.9%) i.a., Patients were monitored closely throughout the entire study period with a total of 4 visits over weeks and documented pain in the morning and evening in a patient´s diary. Safety data was collected during the whole study period. P-values were calculated using two-sided T-tests.Results114 patients were screened, 93 were treated and 89 (96%) completed day 7. Of these n= 61 (66%) were diagnosed with OA and n= 32 (34%) with IA 48 (52%) patients were female, mean age was 58.5 (SD 14 years) and mean disease duration 6.7 years (median 2 years, range <1 year – 42 yearss, IQR <1 – 10 years). The mean VAS pain improvement at day 7 for morphine, triamcinolone and placebo was -22.8, -37.7, and -19.8 respectively. The differences were not significant (p=0.69) for placebo vs. morphine, but significant for placebo vs. triamcinolone and for triamcinolone vs. morphine (p=0.013 and p=0.006). Mean improvements of the everyday pain documentation are shown in Figure 1. During the study period, there were no serious adverse events and 45 adverse events, most of them were mild.Figure 1.Mean VAS pain over one week in patients with chronic knee arthritis treated with morphine, triamcinolone or placebo as a single intraarticular injection.ConclusionIn this randomized, placebo and active controlled double blind trial a single dose of 3 mg i.a. administered morphine did not lead to significant improvements in comparison to placebo and was inferior to triamcinolone at day 7. The same was true during the first 7 days as shown in the pain documentation in patient diaries. These data does not support the use of i.a. morphine for pain reduction in patients with chronic arthritis.Disclosure of InterestsHildrun Haibel Speakers bureau: AbbVie, MSD, Janssen, Roche and Pfizer, Consultant of: Roche, Boehringer, Janssen, MSD, Novartis, and Sobi, Grant/research support from: BMBF Neuroimpa 01EC1403F, Joachim Sieper Speakers bureau: Abbvie, Janssen, Lilly, Merck,Novartis, UCB, Consultant of: Abbvie, Lilly, Merck, Novartis, UCB, Denis Poddubnyy Speakers bureau: AbbVie, Bristol Myers Squibb, Eli Lilly, MSD, Novartis, Pfizer, and UCB, Consultant of: AbbVie, Biocad, Eli Lilly, Gilead, GlaxoSmithKline, MSD, Novartis, Pfizer, Samsung Bioepis, and UCB, Grant/research support from: AbbVie, Eli Lilly, MSD, Novartis, and Pfizer, Valeria Rios Rodriguez: None declared, Fabian Proft Speakers bureau: Abbvie, BMS, MSD, Novartis, Pfizer, Roche and UCB, Consultant of: Abbvie, BMS, MSD, Novartis, Pfizer, Roche and UCB, Grant/research support from: Novartis, Judith Rademacher: None declared, Sabrina Igel: None declared, Peter Martus: None declared, Christoph Stein Grant/research support from: BMBF Neuroimpa 01EC1403F.

The sensory ion channel transient receptor potential vanilloid 1 (TRPV1) is mainly expressed in small to medium sized dorsal root ganglion neurons, which are involved in the transfer of acute noxious thermal and chemical stimuli. The Ankyrin-rich membrane spanning protein (ARMS) interaction with TRPV1 is modulated by protein kinase A (PKA) mediating sensitization. Here, we hypothesize that PKA phosphorylation sites of ARMS are crucial for the modulation of TRPV1 function, and that the phosphorylation of ARMS is facilitated by the A-kinase anchoring protein 79 (AKAP79). We used transfected HEK293 cells, immunoprecipitation, calcium flux, and patch clamp experiments to investigate potential PKA phosphorylation sites in ARMS and in ARMS-related peptides. Additionally, experiments were done to discriminate between PKA and protein kinase D (PKD) phosphorylation. We found different interaction ratios for TRPV1 and ARMS mutants lacking PKA phosphorylation sites. The degree of TRPV1 sensitization by ARMS mutants is independent on PKA phosphorylation. AKAP79 was also involved in the TRPV1/ARMS/PKA signaling complex. These data show that ARMS is a PKA substrate via AKAP79 in the TRPV1 signaling complex and that all four proteins interact physically, regulating TRPV1 sensitization in transfected HEK293 cells. To assess the physiological and/or therapeutic significance of these findings, similar investigations need to be performed in native neurons and/or in vivo.

Abstract Background and Purpose NFEPP is a newly-designed pain killer selectively activating G-protein coupled mu opioid receptors in injured tissues, and therefore devoid of central side effects. However, the cellular mechanisms underlying NFEPP’s antinociceptive effects were not examined in sufficient detail so far. Here we investigated the effects of NFEPP on G-protein activation, on voltage gated calcium channels and on mu opioid receptor phosphorylation. Experimental Approach HEK293 cells stably transfected with mu opioid receptors were used to study [35S]-GTPγS binding and mu opioid receptor phosphorylation. Voltage dependent calcium currents and intracellular calcium signals were examined in rat sensory neurons. All experiments were performed at acidic and physiological pH values using NFEPP compared to the conventional mu opioid receptor agonist fentanyl. To investigate the role of G protein subunits, we used pertussis toxin and gallein. Key Results At low pH, NFEPP produced more efficient G-protein activation and reduction of calcium currents in depolarized sensory neurons. The latter was mediated by G protein βγ subunits and NFEPP-mediated MOR phosphorylation was pH-dependent. Fentanyl-induced signaling was not affected by pH changes. Conclusion and Implications Our study shows that, at low pH, MOR signaling induced by NFEPP is more effective and neuronal calcium channels are directly modulated by G protein βγ subunits dissociated from G protein αi/o subunits. Apparently, the enhanced efficacy of NFEPP is dependent on extra- rather than intracellular effects on opioid receptor function.

OBJECTIVE:The effectiveness of µ-opioid receptor (MOPr) agonists for treatment of visceral pain is compromised by constipation, respiratory depression, sedation and addiction. We investigated whether a fentanyl analogue, (±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide (NFEPP), which preferentially activates MOPr in acidified diseased tissues, would inhibit pain in a preclinical model of inflammatory bowel disease (IBD) without side effects in healthy tissues.DESIGN:Antinociceptive actions of NFEPP and fentanyl were compared in control mice and mice with dextran sodium sulfate colitis by measuring visceromotor responses to colorectal distension. Patch clamp and extracellular recordings were used to assess nociceptor activation. Defecation, respiration and locomotion were assessed. Colonic migrating motor complexes were assessed by spatiotemporal mapping of isolated tissue. NFEPP-induced MOPr signalling and trafficking were studied in human embryonic kidney 293 cells.RESULTS:NFEPP inhibited visceromotor responses to colorectal distension in mice with colitis but not in control mice, consistent with acidification of the inflamed colon. Fentanyl inhibited responses in both groups. NFEPP inhibited the excitability of dorsal root ganglion neurons and suppressed mechanical sensitivity of colonic afferent fibres in acidified but not physiological conditions. Whereas fentanyl decreased defecation and caused respiratory depression and hyperactivity in mice with colitis, NFEPP was devoid of these effects. NFEPP did not affect colonic migrating motor complexes at physiological pH. NFEPP preferentially activated MOPr in acidified extracellular conditions to inhibit cAMP formation, recruit β-arrestins and evoke MOPr endocytosis.CONCLUSION:In a preclinical IBD model, NFEPP preferentially activates MOPr in acidified microenvironments of inflamed tissues to induce antinociception without causing respiratory depression, constipation and hyperactivity.

Abstract Background Opioid drugs are used to treat pain in inflammatory bowel disease (IBD) but their side effects can cause serious morbidity. Therefore, we tested a novel opioid analgesic, ±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenylpropionamide (NFEPP) which selectively activates peripheral µ-opioid receptors at acidic pH, as occurs in inflamed tissue. Aims Evaluate whether NFEPP causes analgesia in the inflamed colon of DSS-colitis mice using both in vitro and in vivo techniques. Methods To measure the visceral motor reflex (VMR) in response to colorectal distention, EMG electrodes connected to a telemetric transmitter were implanted in mice (c57BL/6), after 10 days recovery acute dextran sodium sulfate (DSS) colitis was induced (5 days 2.5% DSS, 2 days water). VMR was measured 30 min after s.c. injection of vehicle or 0.2 mg/kg of NFEPP or fentanyl. Motility was assessed by fecal pellet count 1 hour after NFEPP. Colonic tissue pH was evaluated using the SNARF-4F-5 carboxylic acid probe. Excitability of mouse dorsal root ganglia (DRG) neurons was measured by recording the rheobase (minimum input current to fire an action potential) after superfusion of NFEPP (300 nM, 10 min) or vehicle at pH 6.5 or 7.4. Colonic afferent nerve responses to probing with a von Frey filament (1 gm) were examined before and after exposure to NFEPP (300 nM, 5 min superfusion) at pH 6.5 and 7.4 respectively. The data was analyzed with Welch’s t-test, 1- or 2-way ANOVA with post hoc Dunnett or Bonferroni or Tukey’s test. Results NFEPP significantly inhibited the VMR in response to distension in mice with colitis compared to vehicle (decreased response by 65%, P<0.001). NFEPP had no effect in control mice. Conversely, fentanyl caused a similar decreased response in both groups (DSS 79% and control 67%, P<0.001). Pelleting was not affected by NFEPP injection in either group compared to vehicle. The pH measurement revealed a more acidic environment in DSS colonic tissue (ΔpH0.37±0.14, P<0.05) compared to controls. In patch-clamp studies, NFEPP decreased DRG excitability at pH 6.5 compared to the baseline and vehicle (increased rheobase 53.84%, P<0.01 and 36.36%, P<0.05 respectively) but had no effect at pH 7.4. In colonic afferent nerve recordings, NFEPP significantly attenuated afferent responses (28.9% P<0.01) to probing at pH 6.5 but also had no effect at pH 7.4. Conclusions This pH-selective opioid agonist significantly inhibits pain at the site of inflammation where the tissue pH is acidic but has no effect in tissues where the pH is in the physiological range. Thus, NFEPP could be an effective opioid analgesic in IBD while being devoid of any unwanted side effects. Funding Agencies CCC

HIV remains a major burden to the health care system and neuropathic pain is the most common neurological complication of HIV infection. Because current treatment strategies often lack satisfying pain relief, cannabinoids (CBs) are discussed as a new option. We investigated cannabidivarin (CBDV) as treatment for HIV‐associated neuropathic pain. We conducted a randomized, double‐blind, placebo‐controlled crossover study. Patients underwent two successive treatment phases (4 weeks each) and were treated with CBDV (400 mg/day) or placebo in a randomized order. A 3‐week washout phase was designed to eliminate potential carry‐over effects. Patients were followed up for 3 weeks after the end of the second treatment phase. The primary end point was pain intensity on an 11‐point numeric rating scale, recorded in a diary. Secondary end points were additional pain medication, pain characteristics, and quality of life. We included 32 patients. The mean pain intensity under CBDV was 0.62 points higher compared with placebo (P = 0.16, 95% confidence interval −0.27 to 1.51). CBDV did not influence the amount of additional pain medication, pain characteristics, or quality of life. The incidence of adverse events was similar during both treatments. No suspected unexpected adverse reactions occurred during either treatment. CBDV was safe but failed to reduce neuropathic pain in patients with HIV. This may be explained by a lack of CB receptor activation, as indicated by preclinical experiments. Although a larger patient number might be desirable, we would not expect a change in the conclusions because the present differences are far from statistical significance. Therefore, we would currently not consider CBDV as a clinically meaningful treatment option for neuropathic pain.

The newly designed fentanyl derivative [( ±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide] (NFEPP) was recently shown to produce analgesia selectively via peripheral mu-opioid receptors (MOR) at acidic pH in rat inflamed tissues. Here, we examined the pH-dependency of NFEPP binding to brain MOR and its effects on bone cancer-induced pain in mice. The IC50 of NFEPP to displace bound [3H]-DAMGO was significantly higher compared to fentanyl at pH 7.4, but no differences were observed at pH 5.5 or 6.5. Intravenous NFEPP (30–100 nmol/kg) or fentanyl (17–30 nmol/kg) inhibited heat hyperalgesia in mice inoculated with B16-F10 melanoma cells. The peripherally-restricted opioid receptor antagonist naloxone-methiodide reversed the effect of NFEPP (100 nmol/kg), but not of fentanyl (30 nmol/kg). The antihyperalgesic effect of NFEPP was abolished by a selective MOR- (cyprodime), but not delta- (naltrindole) or kappa- (nor-binaltorphimine) receptor antagonists. Ten-fold higher doses of NFEPP than fentanyl induced maximal antinociception in mice without tumors, which was reversed by the non-restricted antagonist naloxone, but not by naloxone-methiodide. NFEPP also reduced heat hyperalgesia produced by fibrosarcoma- (NCTC 2472) or prostate cancer-derived (RM1) cells. These data demonstrate the increased affinity of NFEPP for murine MOR at low pH, and its ability to inhibit bone cancer-induced hyperalgesia through peripheral MOR. In mice, central opioid receptors may be activated by ten-fold higher doses of NFEPP.