Melanocortin Receptors, Melanotropic Peptides and Penile Erection
2007
Abstract
Penile erection is a complex physiologic event resulting from the interactions of the nervous system on a highly specialized vascular organ. Activation of central nervous system melanocortinergic (MC) receptors with either endogenous or synthetic melanotropic ligands may initiate and/or facilitate spontaneous penile erection.
While the CNS contains principally the MC3 and MC4 receptor subtypes, there is conflicting data as to which receptor mediates erection. Although the MC4R is emerging as the principle effector of MC induced erection, the role of the MC3R is poorly understood. Manipulation of each receptor subtype with newly synthesized receptor specific agonists and antagonists, as well as knockout mice, has elucidated their individual contributions. Novel data from our laboratories suggests that antagonism of forebrain MC3R may enhance melanocortin-induced erections. Furthermore, melanocortin agents may interact with better-studied systems such as oxytocinergic pathways at the hypothalamic, brainstem or spinal level.
Current therapies for erectile dysfunction target end organ vascular tissue. Manipulation of MC receptors may provide an alternative, centrally mediated therapeutic approach for erectile and other sexual dysfunctions. The non-specific “superpotent” MC agonist, PT-141, which is the carboxylate derivative of Melanotan II, has reached phase 2 clinical trials. Through their centrally mediated activity, melanocortin agonists have potential to treat erectile dysfunction as well as possible applications to the unmet medical needs of decreased sexual motivation and loss of libido.
INTRODUCTION
The melanocortinergic (MC) system mediates a wide and complex array of physiological effects including skin pigmentation, salt regulation, food intake regulation, pain nerve regeneration, sexual behavior and penile erection. These vastly different effects occur through selective activation of five known receptor subtypes by unique peptides derived from alternate posttranslational modification of proopiomelanocortin (POMC) gene products including ACTH, ?-MSH, ?-MSH and ?-MSH. The recognition that differential manipulation of specific receptor subtypes could lead to specifically desired physiological outcomes has led to the development of a variety of synthetic compounds, many of which are being actively studied for potential therapeutic effects.
The proerectile effects of MC compounds have been recognized since the mid-20th century when studies by Ferrari and colleagues showed increased sexual excitement after intracerebral delivery of ?-MSH and ACTH in a variety of mammalian species. Interest in human applications of melanocortinergic agents toward penile erection did not occur until fortuitous events of the 80s. During initial testing of a novel synthetic agent intended for artificial tanning, melanotan II, after administration experienced an 8 hour-long erection, along with some nausea and vomiting. Gastrointestinal effects aside, the potential therapeutic effect of this agent for erectile dysfunction was recognized.
In the ensuing years, much effort has been spent toward understanding the relationship between the melanocortinergic system and penile erection. This paper focuses on general and MC specific neuroerectile pathways, receptor subtypes and a detailed discussion of actively studied melanocortin agonists and antagonists. Although several important investigative groups have established MC related neural pathways and implicated specific MC receptors, there continues to be areas of active debate as well as a significant potential for research peptide development as treatment for sexual dysfunctions.
Normal Erectile Physiology
Penile erection is the final endpoint of a complex coordination between the central nervous system, peripheral nervous system, endocrine system, voluntary and involuntary pelvic musculature and the highly specialized vascular tissue of the penis. Supraspinal centers in the brain integrate sensory input and hormonal cues as part of the initiation of sexual desire, arousal and libido. These centrally initiated pro-erectile signals are relayed to sympathetic and parasympathetic centers in the thoracolumbar and sacral spinal cord in order to regulate vascular tone in the penile tissues. Alternately, direct genital afferents to the lumbosacral spinal cord can initiate a reflexogenic erection independent of supraspinal input. Inhibition of sympathetic vasoconstriction coordinated with vasodilatory parasympathetic activation greatly increases blood flow through the paired cavernosal arteries. As the cavernous spaces within the corpora cavernosa expand, they compress the venous outflow pathways leading to marked increase in intracavernosal pressure with subsequent tissue expansion. Voluntary contraction of the bulbocavernosus muscle further increases intracavernous pressures to produce a rigid erection while periurethral and bulbospongiosus muscular contractions assist with seminal ejaculation.
The most important end-organ neurotransmitter modulating erection is now recognized as nitric oxide (NO) [11-13]. Release of NO from the terminals of non adrenergic non cholinergic parasympathetic nerve fibers results in activation of cavernosal smooth muscle cell guanalyl cyclase (GC). This leads to increased production of the cyclic nucleotide guanosine monophosphate (cGMP), which in turn leads to cellular relaxation through direct calcium regulating mechanisms. Smooth muscle contraction and penile detumescence is in turn regulated by phosphodiesterase type 5 enzyme degradation of cGMP, as well as sympathetic activation at the moment of ejaculation.
Erectile Neural Circuits and Melanocortins
Neural control of erection results from a complex interaction between the forebrain, midbrain, spinal cord and peripheral nervous system. Although MC agonists are known to induce penile erection, whether or not endogenous melanocortins are necessary for normal physiologic penile erection remains unknown. A broader knowledge of neural erectile pathways, including the non-melanocortinergic pathways may lead to a greater understanding of areas where the melanocortinergic system may exert influence.
Forebrain and Hindbrain
Erections occur in many different physiological and behavioral contexts and in response to a variety of stimuli, some of which clearly are relevant to reproduction while others seemingly have little to do with reproduction. Examples of the former may include erections during copulation and genital stimulated erections while examples of the latter include erections observed in-utero, nocturnal erections and “non-contact erections”. The supraspinal CNS is primarily responsible for sexual motivation as related to penile erection. Multiple areas of the forebrain and hindbrain may be responsible for erections in each of these contexts. In particular, based on their projections to the lumbosacral spinal cord, the following forebrain regions have been identified as potential initiators or facilitators of penile erection]
The MPOA of the hypothalamus is well recognized for its role in male sexual behavior, likely through the integration and redistribution of information to other hypothalamic and brainstem nuclei. Electrical stimulation of this area as well as the PVN elicits complex sexual responses and erection in male monkeys and rats. Neurons of the PVN are activated by dopamine and send oxytocinergic and vasopressinergic projections to the lumbosacral spinal cord. Lesions of this area decrease non-contact erections while having little effect on copulatory erections. Brainstem NPGI neurons send serotonergic signals to the lumbosacral spinal cord. Lesions of this area remove inhibition of both reflex erections and copulatory erections. PVN projections to the NPGI may be responsible for physiological release of this tonic inhibition of erection. Lesions of the MeA facilitate reflexive erections, depress non-contact erections, and have no effect on copulatory erections.
Complex circuits involving many neurotransmitters, including oxytocin and dopamine, have been described with potential effects related to erection, most of which are beyond the scope of this info...
Within this complex neural network, the melanocortinergic system has multiple potential sites for regulation. In male rats exposed to sexual situations, ?-MSH is secreted in the MPOA. The arcuate nucleus (Arc) of the hypothalamus is a primary source of POMC secreting neurons in the CNS with projections to the lateral hypothalamus, dorsal medial nucleus and the PVN. POMC expression has been documented in regions of the PVN known to send oxytocinergic (OT) projections to the spinal cord, implicating possible regulatory interactions between the OT and MC systems. Male rats show increased expression of the immediate-early gene, Fos, in magnocellular oxytocin neurons in the PVN when exposed to either intromission or direct ?-MSH intracerebral exposure. Central administration of an MC4R antagonist attenuated the increased Fos expression in these PVN neurons and inhibited copulatory behaviors.
Indirect evidence that hindbrain melanocortin signaling may contribute to supraspinal erection may be provided by the localization of POMC neurons in the nucleus tractus solitareus (NTS). An independent caudal brainstem melanocortin receptor trigger for sympathetically stimulated metabolic responses has been reported.
Spinal Cord
The spinal cord contains neurons which project to the penis and are linked with penile erection. These include thoracolumbar sympathetic, sacral parasympathetic and sacral pudendal pathways. Sensory afferents from the penis project primarily to the lumbosacral spinal cord while some corpus cavernosal afferents have been traced to the thoracolumbar spinal cord.
The spinal cord coordinates ascending and descending inputs affecting penile erection utilizing a variety of neurotransmitters. A well-documented pro-erectile pathway involves the aforementioned OT neurons projections from the paraventricular nucleus to the sacral parasympathetic nuclei expressing the OT receptors. These sacral nuclei directly innervate the corpus cavernosum.
More recently, proerectile functions of spinal melanocortin receptors have been proposed. Spinal MC4R mRNA has been demonstrated in multiple studies. Intrathecal injection of the melanocortin agonist, Melanotan II, to the lumbar spinal cord dose-dependently increased spontaneous erections in males. This effect was abolished by intrathecal co-administration of the melanocortin antagonist, SHU-9119. When SHU-9119 was given intracereroventricularly (ICV), it did not block MT-II spinally induced erections. These results suggest that MC agonists act on independent spinal loci for initiation of erection. Notably, these results are in contrast with a study of intrathecal administration of ?-MSH, which failed to affect intracavernosal pressure in anesthetized rats. However, the lack of effect of ?-MSH may be attributable to its relatively lower affinity for the MC4R and/or its rapid metabolism.
While Melanotan 2 (MT-II) clearly induces erections at the supraspinal level, both inductive and facilitative effects. In acutely spinally transected rats (T8 level) as well those with bilateral transaction of pelvic or dorsal penile nerves, systemic Melanotan II facilitated erections induced by cavernous nerve stimulation as measured by increased inter-cavernous pressures. However, the facilitator effect of MT-II was abolished by removal of the lumbar paravertebral sympathetic chain. These results suggest that the facilitatory effects of MT-II act principally to modulate the sympathetic efferents to the pelvis, with little effect on the parasympathetics. The mechanism by which sympathetic modulation promotes increased cavernosal pressures is unclear, highlighting the need for further studies of spinal melanocortin action.
Peripheral
Many studies demonstrate pro-erectile effects of melanocortinergic agents after systemic delivery. However, MC agonists have yet to demonstrate modulation of erection through direct action on the cavernosum. One study used in-situ hybridization to localize MC4R mRNA to stretch activated mechanoreceptors and sensory afferent nerves of the penis. However, in anesthetized male rats, MC agonists injected intracavernosally neither increased intracavernous pressure nor augmented neurostimulated erectile responses. Direct application of an MC4R agonist failed to produce relaxation of cavernosal strips in organ bath experiments or alter calcium currents of isolated cavernosal smooth muscle cells in vitro.
MC Receptor Agonists
A variety of research modalities have been used to elucidate the action of MC compounds on penile erection. MC compound affinity and activity properties are determined by cell culture and membrane receptor assays. In general, MC agonists bind strongly to subsets of the five G-protein coupled MC receptors and cause increased intracellular production of cAMP while MC antagonists bind strongly but do not stimulate cAMP production. Notably MCRs 1, 3, 4 and 5 have high constitutive (ligand-independent) activity enabling antagonists to decrease basal levels of cAMP production.
Models for study of MC compounds on Penile Erection
Endogenous MC Receptor Agonists
ACTH, ?-MSH, ?-MSH and ?-MSH are the known endogenous agonists of the MC system. Each hormone is a product of posttranslational modification of the POMC gene transcript and contains the sequence of His-Phe-Arg-Trp, considered to be the “core” of agonist activity. Only ACTH and ?-MSH have shown the ability to generate sexual stimulation and penile erection in various animal species including rats, rabbits, cats, dogs and monkeys. These pro-erectile effects appear to be androgen-dependent as castration abolishes the aforementioned response. Notably, many of the synthetic MC agonists contain the “core” sequence present in ACTH and ?-MSH, particularly the agents MT-II and PT-141.
Melanotan II - Pharmacology
MT-II is a synthetic cyclic heptapeptide that was initially designed as an artificial tanning agent. Its structure is based on an earlier linear peptide, Melanotan I, however cyclization was introduced to prevent degradation and allow both N and C terminal truncation of the peptide. The pro-erectile activity of Melanotan II was reported as a significant unexpected reaction during a phase-I trial of human tanning. MT-II contains a seven amino acid sequence with homology to receptor binding portions of ?-MSH and ACTH. The compound is thought to cross the blood brain barrier and has high affinity for the MC1R, MC3R and MC4R. MT-II has a similar affinity for MC4R compared with MC3R and may be considered “superpotent” because of its relatively high affinity for MC4R compared with the endogenous peptides ?-MSH and ACTH (10-100 fold difference).
Melanotan studies
The pro-erectile activity of MT-II appears to be both forebrain and spinally mediated, with little, if any, peripheral effect. Dose dependent increases in spontaneous erections in awake Long-Evans rats were noted with administration of MT-II intracerebrally, intrathecally and intravenously. Increases in yawning and grooming behaviors paralleled erectile activity with intracerebral administration but not spinal administration. As discussed previously, when the non-selective MCR antagonist SHU-9119 was given spinally, it blocked spinal MT-II induced erections, however intrathecal SHU-9119 failed to block intracerebral MT-II induced erections. This indicates potentially independent sites of melanocortin action along the CNS axis with intracerebral sites activating multiple downstream pathways including those independent of melanocortinergic activation.
Human studies
A double blind placebo-controlled crossover study by Wessells et al. demonstrated the safety and pro-erectile activity of subcutaneous MT-II. In the absence of erotic stimulation, 10 men with psychogenic (non-organic) erectile dysfunction received subcutaneous doses ranging from 0.025 to 0.157 mg/kg, while erections were monitored by RigiScan over a 6-hour period. Eight of the 10 men developed clinically apparent erections with greater than 80% rigidity of an average duration of 38 minutes compared with 3 minutes for placebo controls. The time to onset ranged from 15 to 270 minutes. Side effects were dose dependent included nausea, stretching, yawning and decreased appetite. At the preferred dose of 0.025 mg/kg side effects were mild.
The above study documented erectogenic effects of MT-II in men with presumed normal underlying physiology. A subsequent study of MT-II was carried out on men with organic ED. In a similar double blind, placebo-controlled crossover study, 10 men received 2 subcutaneous doses of 0.025 mg/kg MT-II and 2 doses of vehicle. MT-II initiated subjectively reported erections following 63% of the drug injection verses 5% of the placebo injections. The mean rigidity score of the responders was 6.9 on a scale of 0 to 10. Mean duration of tip rigidity greater than 80% was 45 minutes with Melanotan II compared to two minutes for placebo. There was increased subjective reporting of sexual desire after MT-II administration compared with placebo, although the question used to assess desire was not designed specifically to measure desire in men not engaging in sexual intercourse.
PT141 (Bremelanotide) -Pharmacology
PT-141 bremelanotide is currently the most studied melanocortinergic compound with regard to therapeutic potential for treatment of erectile dysfunction. PT-141 is a synthetic heptapeptide. PT-141 is a deaminated derivative and likely metabolite of MT-II. Bremelanotide research peptide has strong binding to MC receptors 3 and 4, with a higher affinity for MC4R over MC3R. Application of PT-141 to HEK-293 cells expressing MC4R increases cAMP production, indicating that this compound, like MT-II, acts as an agonist.
PT141 studies
Studies with adult male Sprague-Dawley rats indicate pro-erectile responses through multiple modes of delivery. Intranasal injection of 50?g/kg PT-141 produced a significant increase in spontaneous erections compared with saline controls in rats observed over a 30-minute period. As well, 100% of the drug treated rats had at least one erection. In this study the pro-erectile effect of PT-141 was attributed to hypothalamic stimulation of MC3R and/or MC4R. Two hours after PT-141 (50?g/kg IN) administration, immunostaining for FOS, a measure of neural activation, showed increased expression in the paraventricular nucleus compared with rats administered saline.
Human pt 141 studies
Preliminary trials in humans have established both safety and efficacy of PT-141. A phase 1 randomized double-blind placebo controlled trial involved 24 healthy male subjects without erectile dysfunction. Intranasal doses of 4 to 20mg were delivered to patients in the absence of visual sexual stimulation (VSS). Safety and tolerability were monitored revealing no significant hemodynamic changes or side effects, including priapism. Serum concentration of peptide was dose dependent and peaked at 30 minutes in the maximum dose group. Serum half-life was measured at 120 minutes. Rigi-Scan monitoring of erectile response revealed a significantly increased duration of rigid erections of 140 minutes compared to 22 minutes in the placebo group. Time to onset of erection ranged from 34 to 63 minutes. Erections were considered rigid if they were more than 60% of base rigidity.
Based on the above results, phase II studies were initiated in patients with mild to moderate ED who showed positive erectile response to PDE-5 inhibitors. RigiScan monitoring in the presence of VSS detected a 3-fold increase in erectile activity with PT-141 (20mg intranasal) administration. The duration of base rigidity was significantly increased utilizing both a 60% and 80% cut-off versus placebo. Timing of erections corresponded well to visual stimulation indicating a potential facilitator mechanism of drug action.
In a first Phase IIB at home study, PT-141 induced dose dependent improvements in erectile function as assessed by the International Index of Erectile Function (IIEF). Of the patients who completed at least 3 at home attempts (n = 203), the mean IIEF erectile function domain score increased in a dose dependent fashion (p < 0.05 for 10, 15, and 20 mg). Normal erectile function (EF>26) was achieved by 10, 30, 36, 53, and 50% of patients in the placebo, 5, 10, 15, and 20 mg groups respectively. Improved erections as defined by a global assessment question were reported by 17, 49, 67, 66, and 66% of patients in the placebo, 5, 10, 15, and 20 mg groups respectively. There were no episodes of syncope or hypotension. The only serious adverse event reported in this study occurred in one patient who reported a prolonged erection that was painless and required no treatment. Gastrointestinal side-effects were the primary reasons for discontinuation in the higher two higher dose groups.
In summary, PT-141 is a potent initiator of erection with minimal side effects, a rapid onset of action and a sufficiently long duration of action. Notably, the recent Phase II studies confirm that the erectile responses are augmented by sexual stimulation. With its central mechanism of action, PT-141 may act independent or synergistically with PDE-5 inhibitors and provide a useful alternative therapy for erectile dysfunction, both from organic and psychogenic origin. A randomized prospective placebocontrolled study compared treatement of ED with sildenafil alone verses sildenafil with 7.5mg intranasal PT-141. Co-administration of the two agents resulted in significantly prolonged time of increased base rigidity (>60%) compared with sildenafil alone during a 2.5 hour monitoring session. The combination of drugs was well tolerated with no significantly increased side-effects over either sildenafil or PT-141 alone. The ability of the peptide to safely and effectively induce high quality erections allowed Palatin Technologies to initiate a second Phase IIB study in 2006 and propose Phase III studies in 2007.
THIQ is a synthetic small molecule hMC4R agonist with moderate bioavailability (14%), rapid absorption (Tmax = 1hr) and a short T1/2 (0.5hrs). THIQ has a >600 selectivity for the MC4R compared with MC3R. Studies evaluated the effects of THIQ delivered both systemically and intracerebrally in rodents. Systemic administration (1mg/kg) potentiated electrically stimulated erections as well as decreasing mounting and intromission latency mating behaviors in wild type mice. In an ex copula model using male rats, systemic THIQ dose dependently increased total numbers of erections. This effect was blocked by central administration of the non-specific antagonist, AgRP, as well as the MC4R specific antagonist, MBP-10. ICV delivery of 20?g of THIQ increased reflexive penile erections. There are no studies of THIQ in humans to date. Interestingly, and in contradistinction to MT-II, THIQ has not been reported to initiate spontaneous erections in rodents.
SHU-9119 is a classical inhibitor of both the MC3R and the MC4R. This synthetic cyclic lactam ?-MSH analogue is closely related in structure to MT-II. SHU-9119 actually has agonist properties at MC1R and MC5R, but for the purposes of discussing erection, this compound is considered primarily an antagonist because of the lack of these receptors in the CNS. In rabbits this highly potent compound readily blocked MT-II induced erections when administered systemically. In rats, SHU-9119 blocked erections and grooming/yawning behaviors stimulated by MT-II both at supraspinal and spinal locations. This compound has not been studied in humans.
Erectile Dysfunction and Current Therapies
Erectile dysfunction (ED) is defined as the inability to produce or maintain a penile erection with rigidity sufficient for intercourse. Risk factors include advanced age, diabetes mellitus, hypertension, obesity, dyslipidemia, pharmacologic side effects and cardiovascular disease. The prevalence increases with age and may affect up to one third of men over the age of 50, representing a significant source of morbidity in an aging population.
Currently medical therapies for ED are limited to direct manipulation of cavernosal smooth muscle relaxation. Selective pharmacological inhibition of phosphodiesterase-5 enzyme in penile smooth muscle cells prevents breakdown of cGMP leading to higher intracellular levels of this molecule resulting in increased smooth muscle relaxation and erection. Available PDE-5 inhibitors include sildenafil, vardenafil and tadalafil.
The aforementioned therapies each have drawbacks leading to the search for alternative treatment methods. The PDE-5 inhibitors have been the most successful pharmacotherapy class, however up to 50% of diabetic men with ED remain refractory to such agents. Importantly, while each of the above therapies addresses the mechanical issue of rigidity necessary for penetration and intercourse, none of the above therapies is known to affect sexual desire/libido, an important component to the overall treatment of ED. Thus, there is an unmet medical need to study alternative pathways and agents, such as the melanocortinergic compounds, which may fill in the gaps left by current forms of treatment.
SUMMARY
Complex interactions between the supraspinal, spinal and peripheral nervous system lead to the highly specific and regulated vasculogenic event of penile erection. Of the many neurotransmitters involved, melanocortins appear to play a significant role in regulation of erection, particularly at the supraspinal and spinal levels. MC agents may regulate physiologic erection, and could also have as yet unexplored effects on sexual motivation and libido. Much knowledge has been gained of MC receptor sites and MC receptor subtypes involved in erection, particularly through the utilization of novel compounds which activate and/or inhibit specific MC receptors. However, further detailed studies are necessary, particularly if new therapeutic agents are to be developed. The two superpotent synthetic MC agonists, MT-II and PT-141, have been tested in human subjects, with PT-141 showing promise in early clinical trials for treatment of ED.
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