Peptide Derived Ligands for the Discovery of Safer Opioid Analgesics

Abstract

Drugs targeting the μ-opioid receptor (MOR) remain the most efficacious analgesics for the treatment of pain, but activation of MOR with current opioid analgesics also produces harmful side effects, notably physical dependence, addiction, and respiratory depression. Opioid peptides have been accepted as promising candidates for the development of safer and more efficacious analgesics. To develop peptide-based opioid analgesics, strategies such as modification of endogenous opioid peptides, development of multifunctional opioid peptides, G-protein biased opioid peptides, and peripherally restricted opioid peptides have been reported. This review seeks to provide an overview of the opioid peptides that produce potent antinociception with much reduced side effects in animal models and highlight the potential advantages of peptides as safer opioid analgesics.

Introduction

The endogenous opioid system primarily modulates pain through the activation of the three classical opioid receptors, the μ- (MOR), δ- (DOR), and κ- (KOR) opioid receptors. While activating either receptor subtype produces analgesia, each receptor differentially mediates the adverse effects. Activation of MOR is linked to physical dependence, euphoria, and respiratory depression; activation of DOR causes convulsant effect and activation of KOR is associated with dysphoric and aversive effects. As a natural response to pain, the endogenous opioid peptides such as β-endorphin, enkephalins, and dynorphins are released from the enzymatic cleavage of their respective precursors proopiomelanocortin, proenkephalin, and prodynorphin.¹ The endogenous opioid peptides and their related receptors are broadly expressed across the pain pathways. By binding to and activating the three opioid receptors, these endogenous opioid peptides act together to block pain signaling.²

Endomorphins, EM-1 (Tyr¹-Pro²-Trp³-Phe⁴, Fig. 1) and EM-2 (Tyr¹-Pro²-Phe³-Phe⁴, Fig. 1) are another set of endogenous opioid peptides that have been isolated from bovine brains³ and human brain cortex tissues.⁴ However, whether EM-1 and EM-2 are endogenous in nature is not certain because the sequences are not found in the human genome and no precursors have been identified. EMs selectively bind to and activate the MOR.⁵ Among the endogenous opioid peptides, EM-1 and EM-2 have the highest binding affinity (K_i, μ = 0.36 ± 0.08 nM and 0.69 ± 0.16 nM, respectively) and selectivity for the MOR⁴ (δ/μ = 4,183 and 13,381, respectively, and κ/μ = 15,077 and 7,594, respectively). Intrathecal (i.t.) or intracerebroventricular (i.c.v.) administration of EMs produce efficacious antinociception in rodents, which is blocked by naloxone and MOR-selective antagonists β-funaltrexamine.^6–7 Spinal administration of EM-1 produces analgesia with 6-time higher potency than morphine,⁸ and both EM-1 and EM-2 are without cardiorespiratory effects.⁹

Figure 1.

Structures of endomorphin analogs

The endogenous opioid system relieves pain without producing adverse effects; this differs dramatically from the small molecule opioid pain medications, which produce potent analgesia while exhibiting notorious side effects such as physical dependence, tolerance, addiction, respiratory depression, and constipation. However, endogenous opioid peptides cannot be directly utilized as drugs because they are rapidly inactivated by enzymatic degradation in vivo.¹⁰ To improve their pharmacokinetic property, various synthetic strategies, e.g., cyclization,¹¹ D-amino acid¹² or N-methylated amino acid¹³ replacements, stapling,¹⁴ lipidation,¹⁵ and glycosylation¹⁶ have been developed to modify the structures of the opioid peptides. These efforts have led to the synthesis and identification of a diverse array of close and distal analogs that exhibit improved stability and bioavailability compared with the parent endogenous opioid peptides.

Like small molecule opioid ligands, opioid peptides have also been designed to produce the distinct functional activities, such as G-protein biased agonism,¹⁷ dual- or multi-functional opioid activitis,¹⁸ and peripherally restricted opioid activities¹⁹ for fewer side effects. Compared to small molecule opioid ligands, opioid peptide ligands are more hydrophilic, larger in size, and less permeable to cell membrane. These physicochemical characteristics provide peptides with some unique advantages over small molecules in the context of safer opioid analgesics.

First, opioid peptides bind to the opioid receptors at more extended areas compared with small molecules. Like small molecules, opioid peptides reach to and bind at the bottom of the orthosteric binding pocket; additionally, they interact with the receptor at the extracellular loops as well.²⁰ Simultaneous engagement of the orthosteric binding pocket and the extracellular vestibule of GPCRs has shown to cause additional conformational restriction of the receptor, leading to functional selectivity (bias) through increased specificity of signal transducer binding.^21–23 Secondly, opioid peptides activate opioid receptors in a distinct receptor activation pattern that involves both plasma membrane and endosomal activations, while small molecule drugs, in addition to activating receptors at plasma membrane and endosomes, readily penetrate the cell membrane to drive the internal activation into the Golgi apparatus.²⁴ The spatiotemporal specificity affects signal duration and downstream pathway selection, contributing to distinct downstream physiological effects.^25–27 This suggests that the membrane permeability of ligand may have an impact on the therapeutic and side-effect profiles of opioid analgesics. In addition, opioid peptides have an analgesic duration profile shorter than small molecules.^28–29 However, opioid peptides generally show fewer side effects than classic short-acting pain treatment such as fentanyl.

A great amount of research efforts has been conducted on the design, synthesis, and identification of peptide-based opioid ligands.^30–33 For brevity, this review will primarily focus on opioid peptide ligands that have been reported to demonstrate potent antinociception with attenuated adverse effects in vivo. For a broader review of safer opioid ligands please refer to Varga et al. 2023,³⁴ for a review of opioid peptidomimetics please refer to Lee 2022,³⁵ and for a broader review of non-morphinan based opioid ligands please refer to Smith et al. 2022.³⁶

Endomorphin Analogs

Since the discovery of EMs, various structural modification strategies have been utilized to improve their proteolytic stability/bioavailability. Cyclization of EMs has resulted in some EM analogs that produce potent antinociception with substantially reduced abuse potential and absence of respiratory depression in rodent models. Cyclic EM analogs are often achieved by replacing the Pro² with a D-Lys² and cyclizing through its side chain and the carboxylic terminal. CYT-1010 (Fig. 1), a cyclic EM-1 analog, maintains selective and potent binding at the MOR.³⁷ CYT-1010 is under phase II clinical trials for pain management. In the phase I clinical trials, administration of CYT-1010 (0.1 mg/kg, i.v.) produces antinociception in the cold pressor test for pain without depressing breath at 0.15 mg/kg (i.v.) in human males.³⁸

Adding Glu⁵ and Gly-NH₂⁶ to EM-1 while cyclizing through the side chains of the D-Lys² and Glu⁵ leads to ZH853 (Fig. 1). In addition to producing potent and safer antinociception for treating acute pain in mice,³⁹ ZH853 also provides effective pain relief in chronic pain models,⁴⁰ shortens recovery time from inflammatory and postoperative pain and prevents the development of latent sensitization.⁴¹

Incorporating unnatural amino acids into peptides improves their stability against enzymatic degradation. Two distal EM analogs, MEL-N1606 and MEL-0614 (Fig. 1), generated from replacing multiple residues with unnatural amino acids show equal or greater antinociception but significant reduction of side-effect profile relative to morphine.⁴² MEL-N1906 has three replacements of Tyr¹ with 2’,6’-dimethyl tyrosine (Dmt), Pro² with N-Me-D-Ala, and Phe⁴ with (2-furyl)-α-methylene-β-amino propanoic acid, while MEL-0614 incorporates a β-Pro². MEL-0614 also inhibits the expression of pro-inflammatory cytokines, which may account for their ability to modulate inflammatory pain.⁴³

Conjugating an oligoarginine peptide to EM at C-terminus also improves its stability and bioavailability, as oligoarginine can enhance the delivery of peptides into cell and brain.^44–45 Although attaching an oligoarginine to EM does not improve the binding affinity to MOR, its half-life in mouse brain and serum increases 3–5-fold over that of EM-1, indicating enhanced stability in serum and permeability to blood-brain barrier (BBB).⁴⁶ Attaching an oligoarginine vector to cyclic EM analogs results in “Analog 5” (Dmt-c[Cys-Trp-p-Cl-Phe-Cys]-Gly-(D-Arg)₂, Fig. 1) and “Analog 6” (Dmt-c[Cys-Phe(p-Cl)-Phe-Cys]-Gly-(D-Arg)₂, Fig. 1). Analogs 5 and 6 produce potent antinociception in mice yet significantly less tolerance after chronic administration, significantly less constipation and locomotive impairment, and significantly less (Analog 5) or no (Analog 6) conditioned place preference compared to morphine.⁴⁷

EM-2 analogs have been synthesized by replacing one or two of the residues with unnatural amino acids; these analogs showed selective agonism at MOR with varying degrees of bias toward G-protein. While none of them have been tested in animal models for analgesia or adverse effects, they may serve as a tool to understand the biological and pharmacological significance of biased agonism at MOR for safer opioid analgesia.⁴⁸

Bifunctional and Multifunctional Opioid Peptides

Simultaneously interacting with two or more opioid receptor types, as well as activation of an opioid receptor and a non-opioid receptor can lead to efficacious pain relief with reduced side effects. This has been demonstrated by co-administration of DAMGO and DPDPE,⁴⁹ chronic co-administration of morphine and naltrindole,⁵⁰ and the bifunctional MOR/KOR ligands,⁵¹ MOR/NOP dual agonists,⁵² bifunctional MOR/σ₁ ligands,⁵³ and others MOR/non-opioid ligands.^54–55 Though the mechanism is not currently known, evidence suggests these receptors may interact with and modulate each other.^56–57 In the endogenous opioid system, while an individual endogenous opioid peptide preferentially activates one receptor over the other, as a whole they activate all opioid receptors.⁵⁸ This mixed/balanced effects of the endogenous opioid peptides may account for their powerful analgesia without producing side effects.

MOR /DOR Dual Functional Peptides

Dual functional peptide ligands activating MOR yet antagonizing DOR have shown efficacious analgesia with significantly improved side effect profiles compared to morphine.⁵⁹ The first reported mixed MOR agonist/DOR antagonist peptide is TIPP-NH₂ (Tyr-Tic-Phe-Phe-NH₂) (Fig. 2), an EM-1 analog with a Pro²-to-Tic² replacement.⁶⁰ Replacement of Tyr¹ with 3,5-dimethyltyrocine (Dmt) yielded DIPP-NH₂; further introduction of a reduced peptide bond at Dmt-Tic yielded DIPP-NH₂[Ψ] (Dmt-TicΨ[CH₂NH]Phe-Phe-NH₂) (Fig. 2).⁶¹ However, these peptides with mixed function of MOR agonist/DOR antagonist are unstable in vivo, limiting their application to tool compounds.⁶²

Figure 2.

Structures of bifunctional and multifunctional opioid peptides.

Cyclized bifunctional MOR agonist/DOR antagonist peptides have a significantly improved in vivo stability profile. CycloAnt (Tyr-[D-Lys-Dap(Ant)-Thr-Gly]) (Fig. 2), a MOR agonist/DOR antagonist, was identified from deconvolution of a mixture-based cyclic peptide library containing 24,624 individual peptides using a phenotypic screening in mice.⁶³ CycloAnt produces dose- and time-dependent antinociception with an ED₅₀ of 0.7 mg/kg after intraperitoneal (i.p.) administration. It does not produce respiratory depression at doses up to 15 times the analgesic ED₅₀. Chronic i.p. administration of CycloAnt at 3 mg/kg does not result in opioid-induced hyperalgesia, and significantly reduces signs of naloxone-precipitated withdrawal. The potent antinociception and low liability profile suggests it has a preferred pharmacodynamic and pharmacokinetic profile for systemic use as a potentially safer pain modulator.⁶³ Other cyclic peptide-based MOR agonists/DOR antagonists include JOM-13⁶⁴ (Tyr-c[D-Cys-Phe-D-Pen]OH) (Fig. 2) and VRP26⁶⁵ (Dmt-c(SEtS)[DCys-Aic-DPen]-Ser(Glc)-NH₂) (Fig. 2), which are cyclized by disulfide or dithioether bonds. VRP26 also incorporates a glycosylated serine Ser(Glc)⁵ and a tetrahydroquinoline motif (Aic)⁴. VRP26 produces a maximal antinociceptive effect at 10 mg/kg i.p. with an ED₅₀ of 4.77 mg/kg. Mice given VRP26 at analgesic dose for 7 days do not develop significant tolerance, conditioned place preference, or naltrexone-precipitated withdrawal symptoms.⁶⁴

Interestingly, dual functional MOR/DOR agonists have also been reported to produce potent antinociception with fewer opioid-associated side effects, demonstrated by MMP-2200, a derivative of Leu-enkephalin incorporating a Gly²-to-Thr² replacement and a glycosylated Ser(Glc)⁶(Fig. 2). Compared to morphine, MMP-2200 produces significantly less naloxone precipitated withdrawal symptoms or antinociceptive tolerance, and does not show conditioned place preference.^66–67 Therefore, peptides simultaneously modulating MOR and DOR hold great promise for the development of safe analgesics.

Multifunctional Opioid Peptides

Multifunctional ligands interacting with all three opioid receptors also produce potent and safer analgesia. CYX-6 (Fig. 2) (H-Dmt-Pro-Tmp-Tmp-NH₂, where Tmp is a 2’,4’,6’-trimethylphenylalanine), with a mixed function of MOR agonist and DOR/KOR antagonist, shows potent antinociception with reduced gastrointestinal and respiratory side effects after i.c.v. administration.⁶⁸

Structural modification of a naturally occurring peptide CJ-15,208, cyclo[Phe-D-Pro-Phe-Trp] has yielded several multifunctional opioid peptides. CJ-15,208 binds primarily at the KOR, with an IC₅₀ value of 47 nM, 260 nM, and 2600 nM, to the KOR, MOR, and DOR respectively.⁶⁹ Both [L-Trp⁴]-CJ-15,208 and [D-Trp⁴]-CJ-15,208 isomers display KOR- and MOR- mediated antinociception.⁷⁰ Modification of CJ-15,208 resulted in Cyclo[Pro-Sar-Phe-D-Phe] (Fig. 2), which is a dual KOR/MOR agonist and produces potent antinociception without MOR- and KOR-mediated side effects such as locomotive impairment, ambulation, respiratory depression, conditioned place preference or aversion.⁷¹ Investigation of the stereoisomeric effects of the Phe^1,3 residues of CJ-15,208 led to the discovery of analogs with diverse in vivo functional profiles mediated by multiple opioid receptors, with [D-Phe^1,3]CJ-15,208 (Fig. 2) primarily activating KOR and DOR, and [D-Phe³, D-Trp⁴]CJ-15,208 (Fig. 2) activating all three opioid receptors. These stereoisomers are orally active.⁷²

G Protein-Biased Opioid Peptides

Activated opioid receptors are known to couple to two distinct downstream signaling pathways, mediated by G-protein and the β-arrestin, respectively. Preferential activation of the G-protein signaling pathway may lead to enhanced analgesia with reduced adverse effects.^73–74

LOR17 (c[Phe-Gly-β-Ala-D-Trp]) (Fig. 3), a KOR-selective, G protein-biased agonist, provides significant antinociception in the mouse models of acute, inflammatory, and cancer pain without the KOR-associated side effects.⁷⁵ Naturally occurring peptides Rubiscolin-5 (Tyr-Pro-Leu-Asp-Leu) (Fig. 3) and rubiscolin-6 (Tyr-Pro-Leu-Asp-Leu-Phe) (Fig. 3), which are derived from the large subunit of the enzyme spinach rubisco, are two DOR-selective, G protein-biased agonists. Both peptides show significant antinociceptive effects following i.c.v. or oral (p.o.) administration.⁷⁶ Currently rubiscolin-6 (Rubixyl^®) is under clinical evaluation for its potential to repair skin damage.⁷⁷

Figure 3.

Structures of biased opioid peptides.

Derived from the naturally occurring peptide Bilaid C(Tyr-D-Val-Val-D-Phe), bilorphin (Dmt-D-Val-Val-D-Phe-NH₂) (Fig. 3) is a potent and selective G protein-biased MOR agonist. Bilorphin only produces antinociception in mice after i.t. administration. Coupling a L-Ser(β-Lac)⁵ at bilorphin’s C-terminus resulted in the glycosylated analog bilactorphin (Fig. 3), which shows efficacious analgesia after subcutaneous (s.c.) (ED₅₀ = 35 μmol/kg) as well as p.o. administration. Though bilactorphin does not possess the G protein-biased profile, glycosylation provided an effective way to improve the BBB permeability of a peptide.⁷⁸

Peripherally Restricted Opioid Peptides

Opioid receptors are widely expressed throughout the central nervous system (CNS) and peripheral nervous system (PNS). Under inflammatory condition, the expression levels of opioid receptors in the peripheral neuron are upregulated; inflammatory milieu may also improve opioid receptor function by more efficiently coupling to G-protein and inhibiting cAMP production.^79–80 Restriction of opioid ligands to PNS can produce peripheral opioid analgesia whiling eliminating the adverse effects associated with activating the receptors in the CNS.⁸¹ Peptides generally have limited BBB accessibility; they are promising candidates for targeting opioid receptors in PNS.

Peripherally Restricted Peptide KOR agonists

KOR agonists are promising candidates for developing safer analgesics as they produce antinociception without the side effects of respiratory depression, tolerance, and addiction. In addition to pain, KOR agonists also can modulate inflammation, mood, and reward. However, activation of KOR in CNS can result in sedation, dysphoria, and hallucinations, limiting the clinical application of CNS-acting KOR agonists. Peripherally restricted KOR agonists have therefore received a lot of interest for the development of safer analgesics.

Several peripherally restricted KOR agonists are reported to provide potent analgesic activity without dysphoria or euphoria. These KOR ligands possess all D-amino acids and are analogs of the tetrapeptides f-f-i-r-NH₂ (Fig. 4) and f-f-D-Nle-r-NH₂ (Fig. 4) that were initially identified from screening a mixture-based tetrapeptide library.⁸² Capping the C-terminus of f-f-D-Nle-r-NH₂ with 4-(aminomethyl)pyridine generated CR665 (Fig. 4). Replacing the D-Arg⁴ with N^ε-dimethyl-D-lysine at CR665 led to the discovery of TP-2021 (formerly JT09, Fig. 4).⁸³ CR665 and TP-2021 are both under clinical trial for their therapeutic application in pain management.^84–85 Structural modification of f-f-i-r-NH₂ has resulted in an FDA approved drug Difelikefalin (Fig. 4), which has the D-Leu³-to-D-Ile³ and D-Lys⁴-to-D-Arg⁴ replacements and a C-terminal capping with 4-aminopiperidinine-4-carboxylic acid. Difelikefalin is approved for the treatment of moderate-to-severe pruritus in hemodialysis patients;⁸⁶ no symptoms of physical dependence were observed 2 weeks after discontinuation.⁸⁷

Figure 4.

Structures of peripherally restricted opioid peptides.

Screening a synthetic peptide library constituted of marine cone snail venom derived peptides led to the discovery of Conorphin-T (NCCRRQICC), which exhibits moderate, selective agonism for KOR. Extensive structural modification of Conorphin-T resulted in Analogue 39 (Bz-PrrQ[CHA]CC-NH₂) (Fig. 4), which selectively binds to and produces potent agonism at KOR, and has improved plasma stability compared to its parent peptide.⁸⁸ Analogue 39 also demonstrated analgesia in a mouse model of colonic visceral hypersensitivity after administration to the mucosal surface of the colon, suggesting it may be useful in the treatment of irritable bowel syndrome.

Employing the molecular grafting approach to insert the dynorphin A 1–13 sequence into the scaffold of a plant-derived cyclotide generated a KOR agonist Helianorphin-19 (c-CYGGFLRRCIRPKLK) (Fig. 4). Helianorphin-19 produces potent KOR-regulated antinociception in the chronic visceral hypersensitivity mouse model after intracolonic administration, but neither alters mouse motor coordination in the rotarod test nor exhibits sensitivity for central pain in the jump-flinch test comparing to centrally active KOR agonist U50,488, suggesting a peripherally restricted KOR agonist may be clinically useful for the treatment of chronic abdominal pain.⁸⁹

[T20K]kalata B1 (cyclo-GLPVCGETCVGGTCNTPGCTCSWPVCTRN), an orally bioavailable peptide found in ipeca root powder, is a candidate drug for the treatment of multiple sclerosis. T20K has recently been identified as a positive allosteric modulator of KOR, which increases the efficacy of dynorphin A1−13 and the potency and efficacy of U50,488 in functional cAMP assays.⁹⁰ The discovery of T20K highlights the potential to design novel cyclotide-based allosteric modulators of KOR for developing safer treatments for pain as well as multiple sclerosis.

Other peripherally restricted opioid peptides

In addition to the peripherally restricted KOR agonist, DN-9 (Tyr-D-Ala-Gly-NMe-Phe-Gly-Pro-Gln-Arg-Phe-NH₂) (Fig. 4) is reportedly a multifunctional MOR/KOR/neuropeptide FF receptor agonist that displays effective analgesia following s.c. administration in the mouse acute, inflammatory, and neuropathic pain models.⁹¹ A D-Ala²-to-D-Lys² together with a Gly⁵-to-Asp⁵ replacement and subsequently cyclization through D-Lys² and Asp⁵ led to the discovery of the amide cyclic DN-9 analog c[D-Lys², Asp⁵]-DN-9 (Fig. 4). c[D-Lys², Asp⁵]-DN-9 not only provides more potent analgesia than DN-9 after s.c. and p.o. administration in mice, but it can also be orally administrated.⁹² Though DN-9 and the c[D-Lys², Asp⁵]-DN-9 are peripherally restricted, the cyclic disulfide analog of DN-9, OFP011 (c[D-Cys², Cys⁵]-DN-9)⁹³, and the hydrocarbon-stapled analogs⁹⁴ are brain-permeable. The CNS-acting DN-9 analogs also show reduced side effects, suggesting their multifunctional activity is the main drive for low side-effect profile.

Future Perspectives for Opioid Peptides

The X-ray crystal structures of the MOR, DOR, and KOR were first reported in 2012, all of which are in antagonist-bound inactive states.^95–98 The recent advances in cryo-electron microscopy (cryo-EM), particularly the single-particle cryo-EM technique, which provides comparable resolution to X-ray crystallography, have triggered an explosion of the structural determination of the opioid receptors. Crystal and cryo-EM structures of the opioid receptors have been reported not only in the agonist-bound active states^99–101 but also in complex with G protein.^102–106 The high-resolution cryo-EM structures of the entire family of opioid receptors in complex with the Gi protein and bound to their respective, highly selective endogenous or exogenous opioid peptides have recently been elucidated.²⁰ These cryo-EM structures have offered molecular insights into the interactions between opioid peptides and their receptors. These new insights provide a molecular framework for the rational design and creation of new peptide or peptide-inspired ligands that can interact with their receptors in a similar way as do the endogenous opioid peptides.

Though many of the opioid peptides discussed in this review were discovered through traditional SAR studies, structure-based design has shown promises in designing opioid ligands through fine-tuning the opioid receptor-ligand interactions to improve their pharmacological profiles. Many successes have been achieved to discover safer small-molecule opioid ligands using structure-based design.^107–108 Fentanyl derivatives were recently designed to target the MOR at both the orthosteric binding region and the Na⁺ binding pocket, which has been hypothesized to act as an efficacy and functional selectivity switch for opioid receptors, leading to biased and/or partial agonism. These derivatives demonstrated potent and effective analgesia with significantly fewer adverse effects than morphine.¹⁰⁹

While small molecule ligands can be designed to interact with the Na⁺ binding subpocket and extracellular vestibule subpocket, opioid peptides with a relative larger molecular size may target both the orthosteric site and an additional subpocket more effectively to offer enhanced functional selectivity. Recently a computational approach was developed to design conjugates of De Novo cyclic peptide (DNCP) and β-naloxamine (β-NalA), a KOR partial agonist with a morphinan structure. The cryo-EM structure of human KOR bound to DNCP-β-NalA and G protein heterotrimer showed that the conjugate occupies the orthosteric binding site by β-NalA as well as the extracellular region with the DNCP moiety surrounded by ECL2, ECL3, TM6, and TM7. DNCP-β-NalA produces a potent KOR-mediated antinociception and anti-inflammatory effects, with reduced KOR-mediated side-effects in male mice.¹¹⁰ With the crystal and cryo-EM structures of the peptide-bound receptors and the advancements in algorithms for molecular modeling and dynamic simulations, we anticipate that the rational design and creation of new peptide or peptide-inspired safer opioid analgesics will be significantly facilitated.

Conclusion

Pain not only affects patients with impaired physical functioning and reduced quality of life, but also increases by 2–3 times anxiety, mood, and mental disorders in patients. While opioids remain the most effective analgesics, it is more crucial than ever to develop safer and efficacious opioid analgesics. In addition to their higher target affinity and lower toxicity relative to small molecules, opioid peptides may provide extra advantages over small molecule opioids due to their relatively larger size and higher hydrophilicity. Their larger size may help them engage additional subpockets of opioid receptors, leading to more selective downstream signaling, and partial and/or biased agonism, while their higher hydrophilicity may restrict them to the peripheral nervous system and/or prevent them from quickly reaching to and activating the Golgi opioid receptors after activation in membrane.

In the past, peptides were generally viewed as suboptimal drug candidates due to their poor bioavailability and metabolic instability. Recent advances in peptide synthesis such as cyclization, incorporation of D-amino acids or unnatural amino acids have led to metabolic stable opioid peptides.¹¹¹ Furthermore, advances in formulation and drug delivery systems may provide solutions to overcome this barrier.^112–113 Recent intranasal delivery of the opioid peptide antagonist, CTOP demonstrated improved brain delivery and enhanced antagonism in morphine-treated mice.¹¹⁴ This intranasal drug delivery technique may also improve the delivery of safer peptide opioid agonists to the brain.

The ligands highlighted in this review demonstrate that opioid peptides can produce potent analgesia with attenuated adverse effects through a variety of mechanisms. Multifunctional, biased, and peripherally restricted opioid peptides, as well as endomorphin analogs have all been proven to be promising methods for the development of safer opioid analgesics. With the high-resolution structures of each receptor type in complex with endogenous opioid peptides, design of opioid peptides that act biasedly, selectively or with mixed actions at opioid receptor types is achievable. Recent research on opioid receptor signaling, peptide structural modification, and drug formulation and delivery have deemed opioid peptides more advantageous than ever before.^115–116