From Plant to Chemistry: Sources of Antinociceptive Non-Opioid Active Principles for Medicinal Chemistry and Drug Design.

Pain is associated with many health problems and a reduced quality of life and has been a common reason for seeking medical attention. Several therapeutics are available on the market, although side effects, physical dependence, and abuse limit their use. As the process of pain transmission and modulation is regulated by different peripheral and central mechanisms and neurotransmitters, medicinal chemistry continues to study novel ligands and innovative approaches. Among them, natural products are known to be a rich source of lead compounds for drug discovery due to their chemical structural variety and different analgesic mechanisms. Numerous studies suggested that some chemicals from medicinal plants could be alternative options for pain relief and management. Previously, we conducted a literature search aimed at identifying natural products interacting either directly or indirectly with opioid receptors. In this review, instead, we have made an excursus including active ingredients derived from plants whose mechanism of action appears from the literature to be other than the modulation of the opioid system. These substances could, either by themselves or through synthetic and/or semi-synthetic derivatives, be investigated in order to improve their pharmacokinetic characteristics and could represent a valid alternative to the opioid approach to pain therapy. They could also be the basis for the study of new mechanisms of action in the approach to this complex and disabling pathology.

Discovery of first novel sigma/HDACi dual-ligands with a potent in vitro antiproliferative activity.

Designing and discovering compounds for dual-target inhibitors is challenging to synthesize new, safer, and more efficient drugs than single-target drugs, especially to treat multifactorial diseases such as cancer. The simultaneous regulation of multiple targets might represent an alternative synthetic approach to optimize patient compliance and tolerance, minimizing the risk of target-based drug resistance due to the modulation of a few targets. To this end, we conceived for the first time the design and synthesis of dual-ligands σR/HDACi to evaluate possible employment as innovative candidates to address this complex disease. Among all synthesized compounds screened for several tumoral cell lines, compound 6 (Kiσ1R = 38 ± 3.7; Kiσ2R = 2917 ± 769 and HDACs IC50 = 0.59 µM) is the most promising candidate as an antiproliferative agent with an IC50 of 0.9 µM on the HCT116 cell line and no significant toxicity to normal cells. Studies of molecular docking, which confirmed the affinity over σ1R and a pan-HDACs inhibitory behavior, support a possible balanced affinity and activity between both targets.

From Plant to Chemistry: Sources of Active Opioid Antinociceptive Principles for Medicinal Chemistry and Drug Design.

Pain continues to be an enormous global health challenge, with millions of new untreated or inadequately treated patients reported annually. With respect to current clinical applications, opioids remain the mainstay for the treatment of pain, although they are often associated with serious side effects. To optimize their tolerability profiles, medicinal chemistry continues to study novel ligands and innovative approaches. Among them, natural products are known to be a rich source of lead compounds for drug discovery, and they hold potential for pain management. Traditional medicine has had a long history in clinical practice due to the fact that nature provides a rich source of active principles. For instance, opium had been used for pain management until the 19th century when its individual components, such as morphine, were purified and identified. In this review article, we conducted a literature survey aimed at identifying natural products interacting either directly with opioid receptors or indirectly through other mechanisms controlling opioid receptor signaling, whose structures could be interesting from a drug design perspective.

New Insights into the Opioid Analgesic Profile of cis-(-)-N-Normetazocine-derived Ligands.

In this work, we report on the in vitro and in vivo pharmacological properties of LP1 analogs to complete the series of structural modifications aimed to generate compounds with improved analgesia. To do that, the phenyl ring in the N-substituent of our lead compound LP1 was replaced by an electron-rich or electron-deficient ring and linked through a propanamide or butyramide spacer at the basic nitrogen of the (-)-cis-N-normetazocine skeleton. In radioligand binding assays, compounds 3 and 7 were found to display nanomolar binding affinity for the µ opioid receptor (MOR) (Ki = 5.96 ± 0.08 nM and 1.49 ± 0.24 nM, respectively). In the mouse vas deferens (MVD) assay, compound 3 showed an antagonist effect against DAMGO ([D-Ala2, N-MePhe4, Gly-ol]-enkephalin), a highly selective MOR prototype agonist, whereas compound 7 produced naloxone reversible effect at MOR. Moreover, compound 7, as potent as LP1 and DAMGO at MOR, was able to reduce thermal and inflammatory pain assessed by the mouse tail-flick test and rat paw pressure thresholds (PPTs) measured by a Randall-Selitto test.

Design, synthesis and biological evaluation of novel aminopropylcarboxamide derivatives as sigma ligands.

In our continuing effort to develop novel sigma receptor (SR) ligands, we present the design, synthesis and binding studies of a small library of aminopropylcarboxamide derivatives, obtained from a deconstruction of the piperidine ring of previously synthesized piperidine-based compounds. The best results were achieved with benzofuran (5c, 5g) and quinoline (5a, 5e) derivatives. These compounds revealed the highest affinity for both receptor subtypes. In particular, the 3,4-dimethoxyphenyl derivatives 5e and 5g showed the highest selectivity profile for S2R, especially the quinoline derivative 5e exhibited a 35-fold higher affinity for S2R subtype. The cytotoxic activity of aforementioned compounds was evaluated against SKBR3 and MCF7 cell lines, widely used for breast cancer studies. Whereas the potency of 5g was similar that of Siramesine and Haloperidol in both cell lines, compounds 5a, 5c and 5e exhibited a potency at least comparable to that of Haloperidol in SKBR3 cells. A molecular modelling evaluation towards the S2R binding site, confirmed the strong interaction of compound 5e thus justifying its highest S2R affinity.

Mu and Delta Opioid Receptor Targeting Reduces Connexin 43-Based Heterocellular Coupling during Neuropathic Pain.

Chronic neuropathic pain emerges from either central or peripheral lesions inducing spontaneous or amplified responses to non-noxious stimuli. Despite different pharmacological approaches to treat such a chronic disease, neuropathic pain still represents an unmet clinical need, due to long-term therapeutic regimens and severe side effects that limit application of currently available drugs. A critical phenomenon involved in central sensitization is the exchange of signalling molecules and cytokines, between glia and neurons, driving the chronicization process. Herein, using a chronic constriction injury (CCI) model of neuropathic pain, we evaluated the efficacy of the mu (M-) and delta (D-) opioid receptor (-OR) targeting agent LP2 in modulating connexin-based heterocellular coupling and cytokine levels. We found that long-term efficacy of LP2 is consequent to MOR-DOR targeting resulting in the reduction of CCI-induced astrocyte-to-microglia heterocellular coupling mediated by connexin 43. We also found that single targeting of DOR reduces TNF and IL-6 levels in the chronic phase of the disease, but the peripheral and central discharge as the primary source of excitotoxic stimulation in the spinal cord requires a simultaneous MOR-DOR targeting to reduce CCI-induced neuropathic pain.

Novel N-normetazocine Derivatives with Opioid Agonist/Sigma-1 Receptor Antagonist Profile as Potential Analgesics in Inflammatory Pain.

Although opioids and nonsteroidal anti-inflammatory drugs (NSAIDs) are the most common drugs used in persistent pain treatment; they have shown many side effects. The development of new analgesics endowed with mu opioid receptor/delta opioid receptor (MOR/DOR) activity represents a promising alternative to MOR-selective compounds. Moreover, new mechanisms, such as sigma-1 receptor (σ1R) antagonism, could be an opioid adjuvant strategy. The in vitro σ1R and σ2R profiles of previous synthesized MOR/DOR agonists (-)-2R/S-LP2 (1), (-)-2R-LP2 (2), and (-)-2S-LP2 (3) were assayed. To investigate the pivotal role of N-normetazocine stereochemistry, we also synthesized the (+)-2R/S-LP2 (7), (+)-2R-LP2 (8), and (+)-2S-LP2 (9) compounds. (-)-2R/S-LP2 (1), (-)-2R-LP2 (2), and (-)-2S-LP2 (3) compounds have Ki values for σ1R ranging between 112.72 and 182.81 nM, showing a multitarget opioid/σ1R profile. Instead, (+)-2R/S-LP2 (7), (+)-2R-LP2 (8), and (+)-2S-LP2 (9) isomers displayed a nanomolar affinity for σ1R, with significative selectivity vs. σ2R and opioid receptors. All isomers were evaluated using an in vivo formalin test. (-)-2S-LP2, at 0.7 mg/kg i.p., showed a significative and naloxone-reversed analgesic effect. The σ1R selective compound (+)-2R/S-LP2 (7), at 5.0 mg/kg i.p., decreased the second phase of the formalin test, showing an antagonist σ1R profile. The multitarget or single target profile of assayed N-normetazocine derivatives could represent a promising pharmacological strategy to enhance opioid potency and/or increase the safety margin.

LP1 and LP2: Dual-Target MOPr/DOPr Ligands as Drug Candidates for Persistent Pain Relief.

Although persistent pain is estimated to affect about 20% of the adult population, current treatments have poor results. Polypharmacology, which is the administration of more than one drug targeting on two or more different sites of action, represents a prominent therapeutic approach for the clinical management of persistent pain. Thus, in the drug discovery process the "one-molecule-multiple targets" strategy nowadays is highly recognized. Indeed, multitarget ligands displaying a better antinociceptive activity with fewer side effects, combined with favorable pharmacokinetic and pharmacodynamic characteristics, have already been shown. Multitarget ligands possessing non-opioid/opioid and opioid/opioid mechanisms of action are considered as potential drug candidates for the management of various pain conditions. In particular, dual-target MOPr (mu opioid peptide receptor)/DOPr (delta opioid peptide receptor) ligands exhibit an improved antinociceptive profile associated with a reduced tolerance-inducing capability. The benzomorphan-based compounds LP1 and LP2 belong to this class of dual-target MOPr/DOPr ligands. In the present manuscript, the structure-activity relationships and the pharmacological fingerprint of LP1 and LP2 compounds as suitable drug candidates for persistent pain relief is described.

Intercellular communication and ion channels in neuropathic pain chronicization.

BACKGROUND: Neuropathic pain is caused by primary lesion or dysfunction of either peripheral or central nervous system. Due to its complex pathogenesis, often related to a number of comorbidities, such as cancer, neurodegenerative and neurovascular diseases, neuropathic pain still represents an unmet clinical need, lacking long-term effective treatment and complex case-by-case approach. AIM AND METHODS: We analyzed the recent literature on the role of selective voltage-sensitive sodium, calcium and potassium permeable channels and non-selective gap junctions (GJs) and hemichannels (HCs) in establishing and maintaining chronic neuropathic conditions. We finally focussed our review on the role of extracellular microenvironment modifications induced by resident glial cells and on the recent advances in cell-to-cell and cell-to-extracellular environment communication in chronic neuropathies. CONCLUSION: In this review, we provide an update on the current knowledge of neuropathy chronicization processes with a focus on both neuronal and glial ion channels, as well as on channel-mediated intercellular communication.

Synthesis and Structure-Activity Relationships of LP1 Derivatives: N-Methyl-N-phenylethylamino Analogues as Novel MOR Agonists.

The opioid pharmacological profile of cis-(-)-N-normetazocine derivatives is deeply affected by the nature of their N-substituents. Here, our efforts were focused on the synthesis and pharmacological evaluation of novel derivatives of the lead LP1, a multitarget opioid analgesic compound featuring an N-phenylpropanamido substituent. LP1 derivatives 5a-d and 6a-d were characterized by flexible groups at the N-substituent that allow them to reposition themselves relative to cis-(-)-N-normetazocine nucleus, thus producing different pharmacological profiles at the mu, delta and kappa opioid receptors (MOR, DOR and KOR) in in vitro and in vivo assays. Among the series, compound 5c, with the best in vitro and in vivo profile, resulted a MOR agonist which displays a KiMOR of 6.1 nM in a competitive binding assay, and an IC50 value of 11.5 nM and an Imax of 72% in measurement of cAMP accumulation in HEK293 cells stably expressing MOR, with a slight lower efficacy than LP1. Moreover, in a mouse model of acute thermal nociception, compound 5c, intraperitoneally administered, exhibits naloxone-reversed antinociceptive properties with an ED50 of 4.33 mg/kg. These results expand our understanding of the importance of N-substituent structural variations in the opioid receptor profile of cis-(-)-N-normetazocine derivatives and identify a new MOR agonist useful for the development of novel opioid analgesics for pain treatment.

Development of novel LP1-based analogues with enhanced delta opioid receptor profile.

Pain relief achieved by co-administration of drugs acting at different targets is more effective than that obtained with conventional MOR selective agonists usually associated to relevant side effects. It has been demonstrated that simultaneously targeting different opioid receptors is a more effective therapeutic strategy. Giving the promising role for DOR in pain management, novel LP1-based analogues with different N-substituents were designed and synthesized with the aim to improve DOR profile. For this purpose, we maintained the phenyl ring in the N-substituent of 6,7-benzomorphan scaffold linked to an ethyl spacer bearing a hydroxyl/methyl or methoxyl group at carbon 2 or including it in a 1,4-benzodioxane ring. LP1 analogues were tested by competition binding assays. Compounds 6 (KiMOR=2.47nM, KiDOR=9.6nM), 7 (KiMOR=0.5nM and KiDOR=0.8nM) and 9 (KiMOR=1.08nM, KiDOR=6.6nM) retained MOR affinity but displayed an improved DOR binding capacity as compared to LP1 (KiMOR=0.83nM, KiDOR=29.1nM). Moreover, GPI and MVD functional assays indicated that compounds 6 (IC50=49.2 and IC50=10.8nM), 7 (IC50=9.9 and IC50=11.8nM) and 9 (IC50=21.5 and IC50=4.4nM) showed a MOR/DOR agonist profile, unlike LP1 that was a MOR agonist/DOR antagonist (IC50=1.9 and IC50=1240nM). Measurements of their antinociceptive effect was evaluated by mice radiant tail flick test displaying for compounds 6, 7 and 9 ED50 values of 1.3, 1.0 and 0.9mg/kg, i.p., respectively. Moreover, the antinociceptive effect of compound 9 was longer lasting with respect to LP1. In conclusion the N-substituent nature of compounds 6, 7 and 9 shifts the DOR profile of LP1 from antagonism to agonism.

Evaluation of N-substituent structural variations in opioid receptor profile of LP1.

The benzomorphan scaffold has great potential as lead structure and the nature of the N-substituent is able to influence affinity, potency, and efficacy at all three opioid receptors. Building upon these considerations, we synthesized a new series of LP1 analogues by introducing naphthyl or heteroaromatic rings in propanamide side chain of its N-substituent (9-15). In vitro competition-binding assays in HEK293 cells stably expressing MOR, DOR or KOR showed that in compound 9 the 1-naphthyl ring led to the retention of MOR affinity (Ki(MOR)=38±4nM) displaying good selectivity versus DOR and KOR. In the electrically stimulated GPI, compound 9 was inactive as agonist but produced an antagonist potency value (pA2) of 8.6 in presence of MOR agonist DAMGO. Moreover, subcutaneously administered it antagonized the antinociceptive effects of morphine with an AD50=2.0mg/kg in mouse-tail flick test. Modeling studies on MOR revealed that compound 9 fit very well in the binding pocket but in a different way in respect to the agonist LP1. Probably the replacement of its N-substituent on the III, IV and V TM domains reflects an antagonist behavior. Therefore, compound 9 could represent a potential lead to further develop antagonists as valid therapeutic agents and useful pharmacological tools to study opioid receptor function.

An LP1 analogue, selective MOR agonist with a peculiar pharmacological profile, used to scrutiny the ligand binding domain.

The hypothesis that central analgesia with reduced side effects is obtainable by occupying an 'allosteric' site in the MOR ligand binding domain requires the development of new ligands with peculiar pharmacological profile to be used as tools. New benzomorphan derivatives, analogues of LP1, a multitarget MOR agonist/DOR antagonist, were designed to examine in depth MOR ligand binding domain. Compound 5, bearing a diphenylic N-substituent on the benzomorphan nucleus, showed an affinity (Kiµ=0.5±0.2nM) comparable to that of LP1 and a better selectivity versus DOR and KOR. It elicits antinociceptive effects in ex vivo (GPI) and in vivo. This new compound engages receptor amino acidic residues not reached by LP1 and by other established MOR ligands. Molecular modeling studies, conducted on 5 and on several reference compounds, allowed us to propose possible residues in the MOR ligand binding domain essential for their interactions with 'orthosteric' and 'allosteric' binding sites.

New bifunctional antioxidant/σ1 agonist ligands: Preliminary chemico-physical and biological evaluation.

We previously reported bifunctional sigma-1 (σ1) ligands endowed with antioxidant activity (1 and 2). In the present paper, pure enantiomers (R)-1 and (R)-2 along with the corresponding p-methoxy (6, 11), p-fluoro derivatives (7, 12) were synthesized. σ1 and σ2 affinities, antioxidant properties, and chemico-physical profiles were evaluated. Para derivatives, while maintaining strong σ1 affinity, displayed improved σ1 selectivity compared to the parent compounds 1 and 2. In vivo evaluation of compounds 1, 2, (R)-1, 7, and 12 showed σ1 agonist pharmacological profile. Chemico-physical studies revealed that amides 2, 11 and 12 were more stable than corresponding esters 1, 6 and 7 under our experimental conditions. Antioxidant properties were exhibited by fluoro derivatives 7 and 12 being able to increase total antioxidant capacity (TAC). Our results underline that p-substituents have an important role on σ1 selectivity, TAC, chemical and enzymatic stabilities. In particular, our data suggest that new very selective compounds 7 and 12 could be promising tools to investigate the disorders in which σ1 receptor dysfunction and oxidative stress are contemporarily involved.

The antagonistic effect of the sigma 1 receptor ligand (+)-MR200 on persistent pain induced by inflammation.

OBJECTIVE AND DESIGN: The sigma 1 (σ1) receptor, which is widely distributed in the CNS in areas that are known to be important for pain control, may play a role in persistent pain characterized by the hypersensitivity of nociceptive transmission. Here, we investigated the effect of σ1 blockade in an inflammatory pain model. TREATMENT AND METHODS: An intraplantar injection of carrageenan (2 %) was used to induce paw inflammation. The effects of the σ1 antagonist (+)-MR200, given subcutaneously at a dose of 0.1, 0.25, 0.5,1, 1.5, and 2 mg/kg prior to injection of carrageenan, on inflammatory pain and inflammation were assessed. Mechanical allodynia with von Frey filaments, thermal hyperalgesia with the plantar test and edema evaluation with a plethysmometer were measured. Intergroup comparisons were assessed by one- or two-way analysis of variance when appropriate, followed by post-hoc tests (Dunnett's test for one-way or Bonferroni for two-way ANOVA). RESULTS: (+)-MR200 dose-dependently prevented allodynia and hyperalgesia induced by carrageenan. Furthermore, it reduced paw edema with a significant inhibition percentage of 37.82 % at 3 h after carrageenan treatment. CONCLUSIONS: The blockade of the σ1 receptor with the selective antagonist (+)-MR200 may contribute to the suppression of the typical symptoms of inflammatory pain.

Designing drugs optimized for both blood-brain barrier permeation and intra-cerebral partition.

INTRODUCTION: With the increasing incidence and prevalence of neurological disorders globally, there is a paramount need for new pharmacotherapies. BBB effectively protects the brain but raises a profound challenge to drug permeation, with less than 2% of most drugs reaching the CNS. AREAS COVERED: This article reviews aspects of the most recent design strategies, providing insights into ideas and concepts in CNS drug discovery. An overview of the products available on the market is given and why clinical trials are continuously failing is discussed. EXPERT OPINION: Among the available CNS drugs, small molecules account for most successful CNS therapeutics due to their ability to penetrate the BBB through passive or carrier-mediated mechanisms. The development of new CNS drugs is very difficult. To date, there is a lack of effective drugs for alleviating or even reversing the progression of brain diseases. Particularly, the use of artificial intelligence strategies, together with more appropriate animal models, may enable the design of molecules with appropriate permeation, to elicit a biological response from the neurotherapeutic target.

Design, synthesis, in vitro evaluation, and molecular modeling studies of N-substituted benzomorphans, analogs of LP2, as novel MOR ligands.

6,7-Benzomorphans have been investigated in medicinal chemistry for developing new drugs. This nucleus could be considered a versatile scaffold. The physicochemical properties of benzomorphan N-substituent are crucial in achieving a definite pharmacological profile at opioid receptors. Thus, the dual-target MOR/DOR ligands LP1 and LP2 were obtained through N-substituent modifications. Specifically, LP2, bearing as N-substituent the (2R/S)-2-methoxy-2- phenylethyl group, is a dual-target MOR/DOR agonist and is successful in animal models of inflammatory and neuropathic pain. To obtain new opioid ligands, we focused on the design and synthesis of LP2 analogs. First, the 2-methoxyl group of LP2 was replaced by an ester or acid functional group. Then, spacers of different lengths were introduced at N-substituent. In-vitro, their affinity profile versus opioid receptors has been performed through competition binding assays. Molecular modeling studies were conducted to deeply analyze the binding mode and the interactions between the new ligands and all opioid receptors.

Synthesis, Computational Insights, and Evaluation of Novel Sigma Receptors Ligands.

The development of diazabicyclo[4.3.0]nonane and 2,7-diazaspiro[3.5]nonane derivatives as sigma receptors (SRs) ligands is reported. The compounds were evaluated in S1R and S2R binding assays, and modeling studies were carried out to analyze the binding mode. The most notable compounds, 4b (AD186, KiS1R = 2.7 nM, KiS2R = 27 nM), 5b (AB21, KiS1R = 13 nM, KiS2R = 102 nM), and 8f (AB10, KiS1R = 10 nM, KiS2R = 165 nM), have been screened for analgesic effects in vivo, and their functional profile was determined through in vivo and in vitro models. Compounds 5b and 8f reached the maximum antiallodynic effect at 20 mg/kg. The selective S1R agonist PRE-084 completely reversed their action, indicating that the effects are entirely dependent on the S1R antagonism. Conversely, compound 4b sharing the 2,7-diazaspiro[3.5]nonane core as 5b was completely devoid of antiallodynic effect. Interestingly, compound 4b fully reversed the antiallodynic effect of BD-1063, indicating that 4b induces an S1R agonistic in vivo effect. The functional profiles were confirmed by the phenytoin assay. Our study might establish the importance of 2,7-diazaspiro[3.5]nonane core for the development of S1R compounds with specific agonist or antagonist profile and the role of the diazabicyclo[4.3.0]nonane in the development of novel SR ligands.

Sigma-1 Receptor Inhibition Reduces Mechanical Allodynia and Modulate Neuroinflammation in Chronic Neuropathic Pain.

Neuropathic pain is one of the most debilitating forms of chronic pain, resulting from an injury or disease of the somatosensory nervous system, which induces abnormal painful sensations including allodynia and hyperalgesia. Available treatments are limited by severe side-effects and reduced efficacy in the chronic phase of the disease. Sigma-1 receptor (σ1R) has been identified as a chaperone protein, which modulate opioid receptors activities and the functioning of several ion channels, exerting a role in pain transmission. As such, it represents a druggable target to treat neuropathic pain. This study aims at investigating the therapeutic potential of the novel compound (+)-2R/S-LP2, a σ1R antagonist, in reducing painful behaviour and modulating the neuroinflammatory environment. We showed that repeated administration of the compound significantly inhibited mechanical allodynia in neuropathic rats, increasing the withdrawal threshold as compared to CCI-vehicle rats. Moreover, we found that (+)-2R/S-LP2-mediated effects resolve the neuroinflammatory microenvironment by reducing central gliosis and pro-inflammatory cytokines expression levels. This effect was coupled with a significant reduction of connexin 43 (Cx43) expression levels and gap junctions/hemichannels mediated microglia-to-astrocyte communication. These results suggest that inhibition of σ1R significantly attenuates neuropathic pain chronicization, thus representing a viable effective strategy.

Discovery of a novel series of potent carbonic anhydrase inhibitors with selective affinity for µ Opioid receptor for Safer and long-lasting analgesia.

In this study, we investigated the development of dual-targeted ligands that bind to both µ-opioid receptor (MOR) and carbonic anhydrase (CA) enzymes, using fentanyl structure as a template. We synthesized and evaluated 21 novel compounds with dual-targeted affinity identifying the lead candidate compound 8, showing selective affinity for MOR and potent inhibition of several cytosolic CA isoforms. By means of repeated treatment of 3 daily administrations for 17 days, fentanyl (0.1 mg/kg, subcutaneously) led to tolerance development, pain threshold alterations and withdrawal symptoms in CD-1 mice, as well as astrocyte and microglia activation in the dorsal horn of the lumbar spinal cord. In contrast, compound 8 (0.32 mg/kg s.c.) maintained stable during days its analgesic effect at the higher dose tested with fewer withdrawal symptoms, allodynia development and glial cells activation. Our results suggest that targeting both MOR and CA enzymes can lead to the development of new class of potent analgesic agents with fewer side effects and reduced tolerance development. Further studies are needed to explore the potential mechanisms underlying these effects and to further optimize the therapeutic potential of these compounds.