HK40005263B - Magnesium-containing oxytocin formulations and methods of use - Google Patents

Description

Magnesium-containing oxytocin formulations and methods of use

Cross Reference to Related Applications

This application claims priority to U.S. provisional patent application No. 62/321,654, filed 2016, 4, 12, the disclosure of which is incorporated herein by reference in its entirety.

Technical Field

The present invention relates to methods and compositions comprising oxytocin peptides and magnesium ions for the treatment of autism spectrum disorders (autism spectrum disorders), related disorders and symptoms of such disorders.

Background

Oxytocin is a naturally occurring nine amino acid neuropeptide produced primarily in the paraventricular and supraoptic nuclei of the mammalian hypothalamus. It is released through distributed neural pathways into the central nervous system and through the posterior pituitary into the peripheral circulation. Currently, the United states approves the synthesis of oxytocin by intramuscular injection or intravenous infusionTo produce or improve uterine contractions to promote vaginal delivery and controlAnd (5) treating postpartum hemorrhage. From 1960 to 1997, intranasal oxytocinHas been approved in the united states for stimulating milk secretion to promote breast feeding. Although it is used forThe nasal spray of (a) should be withdrawn from the us market at the request of the manufacturer, but intranasal oxytocin is still sold in countries outside the us, such as switzerland, portugal or brazil. The use of oxytocin peptides for the treatment of autism spectrum disorders has recently been demonstrated. See WO2004/030524 A2 and WO2008/042452 A1, the disclosures of which are incorporated herein by reference.

Autism spectrum disorders are becoming more prevalent in the human population and are typically identified by certain behaviors and characteristics, such as impairment in communication skills and/or social interactions, lack of eye contact, and/or inability to form and/or maintain social relationships. Children and adults diagnosed with an autism spectrum disorder may exhibit one or more of the above behaviors and characteristics to varying degrees. Symptoms often observed in individuals with autism spectrum disorders are persistent deficits in social communication and social interaction, social anxiety, and limited repetitive behaviors, interests, and activities. Other behaviors and characteristics also observed in patients with autism spectrum disorder include aversion to physical contact, generalized anxiety, monotonous voice or inability to adjust the volume of voice, failure to develop companion relationships, lack of shared enjoyment and interest, and lack of social or emotional reciprocity. Other disorders that present with social and communication deficits may include social anxiety disorders, obsessive compulsive disorders, social (pragmatic) communication disorders, and neurodevelopmental disorders, including but not limited to attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, frangle-X syndrome, rett syndrome, or Williams syndrome, which present symptoms similar to those present in autism spectrum disorders. People with autism spectrum disorders often have communication deficiencies such as improper responses in conversations, misreading non-verbal interactions, or difficulty in establishing friendships that suit their age. Furthermore, people with autism spectrum disorders may be overly reliant on practice, highly anxious and sensitive to changes in their environment, or have strong concerns about inappropriate items (e.g., inanimate objects and/or a narrow interest in a particular subject). Moreover, the symptoms of people with autism spectrum disorders vary widely and are continuous, with some individuals exhibiting mild symptoms and others exhibiting very severe symptoms. The core obstacles in social communication and social interaction, or limited repetitive behaviors, interests and activities, of patients with autism spectrum disorders and related disorders do not have available drug therapies and there is still a pressing need for such therapies.

Oxytocin has been shown to improve the core symptoms of autism, in particular social and communication deficits and associated anxiety symptoms. Human clinical trials have demonstrated the efficacy of intranasal oxytocin in the treatment of autism spectrum disorders, related disorders, and symptoms of such disorders. See, e.g., yatawara et al, mol. Psychiatry2015,1-9; gorka et al, neuropsychopharmacology2015,40 (2): 278-286; angnostou et al, mol. Autism2012,3 (1): 16; guastella et al, psychoneuroendocrinology2009,34 (6): 917-923. However, these trials have shown that patients with autism spectrum disorders and related disorders have a large variability in response to the treatment with oxytocin. Therefore, there is a need for an oxytocin peptide formulation that is capable of providing more significant efficacy in the treatment of autism spectrum disorders and related disorders.

Summary of The Invention

Methods and compositions comprising oxytocin peptide and magnesium ions for treating autism spectrum disorders, related disorders, and symptoms of such disorders are provided, including co-administration of oxytocin peptide and magnesium ions by craniofacial mucosal administration (e.g., intranasal administration). The methods and magnesium-containing oxytocin peptide formulations (formulations) described herein provide enhanced efficacy in treating autism spectrum disorders compared to oxytocin alone.

In one aspect, the invention provides a method for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect. The oxytocin peptide and the magnesium ion may be co-administered simultaneously or sequentially. In some embodiments, the oxytocin peptide is administered simultaneously with the magnesium ion in the same unit dose or in separate unit doses or formulations. In some embodiments, the oxytocin peptide and the magnesium ion are administered sequentially. For example, oxytocin peptide is administered a period of time after administration of magnesium ions. In some embodiments, the subject is a human.

The oxytocin peptide and the magnesium ion may be administered to a subject in need thereof by the same route or different routes. In some embodiments, the oxytocin peptide is administered via craniofacial mucosal administration (e.g., nasal, buccal, sublingual, or ocular administration). In one embodiment, both the oxytocin peptide and the magnesium ion are administered intranasally in the same formulation.

In some aspects, interleukin-6 (IL-6) is used as a biomarker of potential efficacy of administration of oxytocin peptide in a subject according to the methods described herein for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with autism spectrum disorder, a social and communication deficit, or anxiety; and for selecting a subject for applying the method. In some embodiments, the method comprises measuring the level of IL-6 in the subject and administering to the subject having an elevated level of IL-6 an effective dose of an oxytocin peptide and a magnesium ion.

In one aspect, a method for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect, further comprises administering to the subject an effective dose of interleukin-6 (IL-6), wherein administration of IL-6 causes an increase in oxytocin receptor expression.

In some embodiments, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1). In some embodiments, the effective dose of oxytocin peptide is about 0.5 μ g to about 2000 μ g, preferably about 8 μ g to about 1000 μ g, more preferably about 15 μ g to about 120 μ g. In some embodiments, an effective dose of magnesium ions administered is from about 50 μ g to about 68mg, preferably from about 50 μ g to about 34mg, more preferably from about 1mg to about 3mg. In some embodiments, the methods comprise administering an amount of a magnesium salt (e.g., magnesium citrate and/or magnesium chloride) to provide from about 50 μ g to about 68mg of magnesium, or from about 50 μ g to about 34mg of magnesium, or from about 1mg to about 3mg of magnesium. In some embodiments, the method comprises administering an amount of magnesium citrate or magnesium chloride to provide from about 50 μ g to about 68mg of magnesium, or from about 50 μ g to about 34mg of magnesium, or from about 1mg to about 3mg of magnesium. In some embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 0.5 μ g to about 2000 μ g, or about 15 μ g to about 120 μ g (e.g., about 60 μ g or about 66 μ g) of oxytocin peptide administered as an aqueous solution containing about 0.11% to about 2.8% (preferably about 1.1% to about 1.6%, e.g., about 1.36%) (w/v) magnesium.

In some embodiments, the present invention provides methods of reducing one or more symptoms associated with an autism spectrum disorder. Symptoms treatable by this method include any social or communication deficit treatable by oxytocin peptides, such as eye contact, social anxiety, generalized anxiety, accuracy in determining complex social cues, mental awareness (empathy), and a deficit in communication capacity including expression of language functions.

In some embodiments, the invention provides methods for treating a disorder exhibiting one or more symptoms associated with an autism spectrum disorder. In some embodiments, the disorder is a social anxiety disorder, obsessive compulsive disorder, a social (pragmatic) communication disorder, and a neurodevelopmental disorder, including but not limited to attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, fragile-X syndrome, rett syndrome, or Williams syndrome, which exhibit symptoms similar to those exhibited in autism spectrum disorders.

In one embodiment, the present invention provides a method for treating an autism spectrum disorder, comprising administering (e.g., intranasally) an effective dose of oxytocin peptide and magnesium ion to a subject in need thereof, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect. In one embodiment, the invention provides a method for treating an autism spectrum disorder comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein the effective dose of oxytocin peptide and magnesium ion are administered intranasally in a liquid formulation and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _.

In one embodiment, the invention provides a method for treating Prader-Willi syndrome comprising administering (e.g., intranasally) an effective dose of oxytocin peptide and magnesium ions to a subject in need thereof, wherein co-administration of the oxytocin peptide and magnesium ions produces a synergistic or potentiating effect. In one embodiment, the invention provides a method for treating Prader-Willi syndrome comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions, wherein the effective dose of the oxytocin peptide and magnesium ions are administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _.

In one embodiment, provided is a method for treating social and communication deficits comprising administering (e.g., intranasally) an effective dose of oxytocin peptide and magnesium ion to a subject in need thereof, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or enhanced effect. In one embodiment, the present invention provides a method for treating social and communication deficits comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein the effective dose of oxytocin peptide and magnesium ion are administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _.

In one embodiment, provided is a method for treating anxiety comprising administering (e.g., intranasally) effective doses of oxytocin peptide and magnesium ion to a subject in need thereof, wherein co-administration of the oxytocin peptide and the magnesium ion produces a synergistic or potentiating effect. In one embodiment, the invention provides a method for treating anxiety comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions, wherein the effective dose of the oxytocin peptide and magnesium ions are administered intranasally in a liquid formulation and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _.

In some of these embodiments, the effective dose of oxytocin peptide is about 0.5 μ g to about 2000 μ g. In some of these embodiments, the effective dose of magnesium ion is from about 50 μ g to about 68mg. In some of these embodiments, the effective dose of the oxytocin peptide and the magnesium ion comprises about 15 μ g to about 120 μ g of the oxytocin peptide — administered as an aqueous solution containing about 1.1% to about 1.6% (w/v) magnesium. In some of these embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 66 μ g of oxytocin peptide — administered as an aqueous solution containing about 1.36% magnesium. In some of these embodiments, the weight ratio between the dose of oxytocin peptide administered and the dose of magnesium ion administered is between about 1. In some of these embodiments, the molar ratio between the dose of oxytocin peptide administered and the dose of magnesium ion administered is between about 1. In some of these embodiments, the liquid formulation is administered in a volume of between about 50 μ L and about 200 μ L. In some of these embodiments, the liquid formulation is administered from 1 to 4 units, about 50 μ L/unit (e.g., spray (spray) or jet (puff)) using a metered nasal device. In some of these embodiments, the oxytocin peptide is human oxytocin (SEQ. ID NO: 1).

In some of these embodiments, the liquid formulation is contained in an intranasal administration device. In some of these embodiments, the intranasal administration device is a nasal pump device. In some of these embodiments, the nasal pump device includes a container bottle attached to the pump actuator. In some of these embodiments, the pump actuator is metered to deliver a specified volume of about 50 μ Ι _. In some of these embodiments, the nasal pump device comprises a container bottle attached to the aerosolizer. In some of these embodiments, the nasal pump device comprises one or more of: (ii) a filter for preventing backflow, (ii) a metal-free fluid path, and (iii) a gamma-radiation stable plastic material.

Further provided are magnesium-containing oxytocin peptide formulations as described herein for use in a method of treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety in a subject in need thereof. Also provided is the use of a magnesium-containing oxytocin peptide formulation as described herein for the manufacture of a medicament for the treatment of autism spectrum disorders, disorders exhibiting one or more symptoms associated with autism spectrum disorders, social and communication deficits, or anxiety.

Also provided is a kit comprising a magnesium-containing oxytocin peptide formulation as described herein, contained in an intranasal administration device such as a nasal pump device and suitable packaging. The kit may further comprise instructions for administering the magnesium-containing oxytocin peptide formulation in a subject in need thereof for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety.

Brief Description of Drawings

Figure 1 shows the effect of saline, oxytocin, a combination of magnesium citrate and oxytocin, and magnesium citrate on social behaviour in a rat model.

Figure 2 shows the effect of saline, oxytocin, a combination of magnesium citrate and oxytocin, and magnesium citrate on anxiety in a rat model.

Figures 3A and 3B show the effect of magnesium citrate, oxytocin, and a combination of magnesium citrate and oxytocin on anxiety in an elevated plus maze (elevated plus size) rat model.

Detailed description of the invention

The present invention provides, inter alia, methods for treating autism spectrum disorder, a disorder showing one or more symptoms associated with autism spectrum disorder, a social and communication deficit, or anxiety in a subject in need thereof by craniofacial administration (e.g., intranasal administration) of an oxytocin peptide and magnesium ions, or a magnesium-containing oxytocin peptide formulation described herein. The oxytocin peptide and the magnesium ion are administered in an effective dose to produce a synergistic or enhanced effect compared to the oxytocin peptide administered alone.

Definition of

As used herein, "oxytocin peptide" refers to a substance that has a biological activity associated with native oxytocin. The oxytocin peptide may be a naturally occurring endogenous peptide, a fragment, analog or derivative thereof. The oxytocin peptide may also be a non-endogenous peptide, fragment, analog or derivative thereof. In one aspect, the oxytocin peptide is human oxytocin. In other aspects, the oxytocin peptide may be an analog or derivative of human oxytocin.

As used herein, "analog" or "derivative" refers to any peptide similar to naturally occurring oxytocin in which one or more amino acids within the peptide have been substituted, deleted, or inserted. The term also refers to any peptide in which one or more amino acids (e.g., one, two, or three amino acids) have been modified (e.g., by chemical modification). In general, the term encompasses all peptides that exhibit oxytocin activity but may, if desired, have different potency (potency) or pharmacological profile (profile).

As used herein, unless otherwise specified, the terms "treatment" or "treating" refer to a method of achieving a beneficial or desired result, such as a clinical result. Beneficial or desired clinical results for autism spectrum disorders and related disorders include, but are not limited to, symptom relief and/or reduction in the degree of symptoms, e.g., reduction in the degree of social and/or communication deficits and/or repetitive behavior and/or anxiety. Social and communication deficits may include, but are not limited to, impairment in communication skills and/or social interactions, lack of eye contact, and/or inability to form and/or maintain social relationships.

"synergistic effect" (synergy), "synergy" or "synergistic effect" refers to the combined action of two or more compounds in such a way that one compound supplements or enhances the action of the other compound to produce an effect greater than would be expected or desired by the addition of a given metered amount of the two or more compounds, if administered separately. A "synergistic effect" may be said to be achieved when two or more agents used in combination produce an overall effect (e.g., improving social and communication deficits and/or reducing anxiety) that is greater than any of their individual effects by adding the effects of the individual agents in equal amounts as would be expected or expected. A "synergistic effect" is also considered to be achieved when the use of two or more agents in combination results in a faster onset and/or longer lasting effect following administration of the individual agents used alone in equal amounts.

"craniofacial mucosal administration" refers to delivery to the mucosal surfaces of the nose, nasal passages, nasal cavities; mucosal surfaces of the oral cavity, including the gums, floor of the mouth, lips, tongue, sublingual oral surfaces (including the frenulum of the tongue and floor of the mouth), and mucosal surfaces of or around the eye, including the mucosa of the conjunctiva, lacrimal gland, nasolacrimal duct, and upper or lower eyelids and the eye.

"intranasal administration" or "intranasally administered" refers to delivery to the nose, nasal passage, or nasal cavity by spray, drops, powder, gel, film, inhalant, or other means.

The "lower region of the nasal cavity" generally refers to the portion of the nasal cavity in the middle and lower nasal turbinate projections, and is the region of the nasal cavity that is significantly innervated by the trigeminal nerve. The "upper region of the nasal cavity" is defined by the upper third in which the olfactory innervation is located and the lamina cribrosa region.

As used herein, "subject" or "patient" refers to a mammal, including but not limited to a human. Mammals include, but are not limited to, domestic animals (e.g., cows), sport animals, pets (e.g., guinea pigs, cats, dogs, rabbits, and horses), primates, mice, and rats. In one embodiment, the subject is a human.

It should be noted that, as used herein, the singular forms "a," "an," and "the" include plural referents unless otherwise specified. In addition, as used herein, the term "comprise" and its cognates are used in their inclusive sense; that is, to the extent that the term "includes" and its corresponding cognates.

Where a range of values is provided, it is intended that each intervening value, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. For example, if a range of 1 μ g to 8 μ g is stated, it is intended that 2 μ g, 3 μ g, 4 μ g, 5 μ g, 6 μ g, and 7 μ g are also expressly disclosed, as well as ranges of values greater than or equal to 1 μ g and ranges of values less than or equal to 8 μ g. If a range of 10-14% is stated, it is intended that 10%, 11%, 12%, 13%, and 14% are also expressly disclosed. Moreover, each smaller range of stated ranges between any stated value or intervening value and any other stated or intervening value in that stated range is encompassed within the disclosure. The upper and lower limits of these smaller ranges may independently be included or excluded in the range, and each range where either, neither or both limits are included in the smaller ranges is also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where it is stated that a range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

Oxytocin peptides

Oxytocin is one of the first peptide hormones that was isolated and sequenced. Native oxytocin is a nine amino acid cyclic peptide hormone with two cysteine residues forming a disulfide bridge between positions 1 and 6. The amino acid sequence of human oxytocin is Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ ID NO: 1).

There are methods described for the production of oxytocin, see for example U.S. Pat. nos. 2,938,891 and 3,076,797; furthermore, oxytocin is commercially available. A variety of peptide analogs and derivatives are available, while others are contemplated for use in the present invention and can be produced and tested for biological activity according to known methods. Oxytocin analogs may include, but are not limited to, 4-threonine-1-hydroxy-diaminooxytocin, 4-serine-8-isoleucine-oxytocin, 9-deamidated oxytocin, 7-D-proline-oxytocin and deaminated analogs thereof, (2, 4-di-isoleucine) -oxytocin, deaminated oxytocin analogs, 1-deaminated-1-monocarba (monocarba) -E12-Tyr (OMe)]-OT (dCOMOT), 4-threonine-7-glycine-oxytocin (TG-OT), oxytocin, deamino-6-carba-oxotoxin (dC 60), L-371,257 anda related series of compounds containing an ortho-trifluoro (trigluro) ethoxyphenylacetyl nucleus is disclosed, for example, in L-374,943. Other exemplary oxytocin analogs include 4-threonine-1-hydroxy-desamino oxytocin, 9-deamidated oxytocin, oxytocin analogs containing a glycine residue in place of a glycinamide residue, (2, 4-dileucine) -oxytocin, analogs of oxytocin having natriuretic (natrietic) and diuretic (diuretic) activity, desamino oxytocin analogs; long-acting oxytocin analogue, 1-deamino-1-monocarba-E12- [ Tyr (OMe)]-OT (dCOMOT), carbetocin, (1-butyric acid-2- (O-methyl-L-tyrosine) -1-carbaoxytocin, deamination-1 monocarba- (2-O-methyltyrosine) -oxytocin [ d (COMOT)])、[Thr4-Gly7]Oxytocin (TG-OT), oxytocin, ile-conopressin, deamino-6-carba-oxitoxin (dC 60), d [ Lys (8) (5/6C-fluorescein)]VT, d [ Thr (4), lys (8) (5/6C-fluorescein)]VT、[HO(1)][ Lys (8) (5/6C-fluorescein)]VT、[HO(1)][ Thr (4), lys (8) (5/6C fluorescein)]VT, d [ Om (8) (5/6C-fluorescein)]VT, d [ Thr (4), om (8) (5/6C-fluorescein)]VT、[HO(1)][ Om (8) (5/6C-fluorescein)]VT、[HO(1)][ Thr (4), om (8) (5/6C-fluorescein)]VT, and 1-deamino-oxytocin in which the disulfide bridge between residues 1 or 6 is replaced by a thioether, and deamino (desamino) -oxytocin analogs in which the disulfide bridge is replaced by a diselenide, ditellurium, telluriumselenide, tellurithion or selenothioide bond (e.g., peptide analogs of oxytocin described in PCT patent application WO2011/120,071 (incorporated herein by reference)). The peptide used in the present invention may be a peptide obtainable by partial substitution, addition, or deletion of amino acids in a naturally occurring or natural peptide sequence. Peptides may be chemically modified, for example, by carboxy-terminal (-NH) ₂ ) Amidation, use of D amino acids in peptides, incorporation of small non-peptide moieties, and modification of the amino acids themselves (e.g., alkylation or esterification of side chain R-groups). Such analogs, derivatives and fragments should substantially retain the desired biological activity of the native oxytocin peptide. In some embodiments, the oxytocin analog is 4-serine-8-isoleucine-oxytocin or 9-deamidated oxytocin. In some embodiments, the oxytocin analog is carbetocin. The present disclosure also includes other known oxytocin analogs, e.g., PCT patent applications WO2012/042371 andpeptide oxytocin receptor agonists described in et al.j.med.chem.2014,57 5306-5317 (the entire contents of which are incorporated herein by reference). In some embodiments, the oxytocin analog is selected from the group consisting ofCompounds of compounds No. 1 to 65 described in tables 1 to 3 in et al.j.med.chem.2014,57 5306 to 5317. In some embodiments, the oxytocin analog is selected from compound No. 31 ([ 2-ThiMeGly 7)]dOT), compound No. 47 (carba-6- [ Phe2, buGly 7)]dOT), compound No. 55 (kappa-6- [3-MeBzlGly 7)]dOT) and Compound No. 57 (carba-1- [4-FBzlGly 7)]dOT, also known as merotocin).

In some embodiments, the oxytocin or oxytocin analog is isotopically labeled with one or more atoms replaced with isotopes having different atomic masses. Examples of isotopes that can be incorporated into the disclosed compounds include hydrogen (e.g., ² h and ³ h) Carbon (e.g., ¹³ c and ¹⁴ c) Nitrogen (e.g., ¹⁵ n), oxygen (e.g., ¹⁸ o and ¹⁷ o), phosphorus (e.g., ³¹ p and ³² p), fluorine (for example, ¹⁸ f) The amount of chlorine (e.g., ³⁶ cl) and sulfur (e.g. sulfur) ³⁵ S) isotopes. Isotopically-labeled compounds can be administered to a subject or other subject and subsequently detected in accordance with conventional techniques to generate useful diagnostic and/or therapeutic management data. Further, the isotopically labeled compound can be administered to a subject or other subject in need thereof, resulting in a therapeutically beneficial absorption, distribution, metabolism, and/or elimination profile. All isotopic variations of oxytocin peptides (e.g. human oxytocin or an analogue or derivative thereof) are contemplated, whether radioactive or non-radioactive.

In some embodiments, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1).

An "international unit" (IU, UI or IE) is an internationally accepted unit of activity for the quantification of vitamins, hormones and vaccines. It defines the amount of substance that gives the unit of activity as determined using a defined bioassay to standardize preparations from a variety of source materials. Similarly, USP units are defined dosage units created by the united states pharmacopeia in cooperation with the food and drug administration in order to ensure the characteristics, strength, quality, purity and consistency of pharmaceutical products. Typically, USP units are equal to international units due to coordination efforts. Conventionally, for oxytocin, 1 activity unit is usually defined as approximately equal to 2 micrograms of synthetic oxytocin peptide; or 1mg equals 500 units (Stedman's Medical Dictionary). Thus, as used herein, an "IU" or "international unit" of oxytocin peptide is an amount of oxytocin peptide that has the same biological activity as about 2 micrograms of synthetic peptide or produces the same level of biological effect (e.g., contractile response in rat uterine strips) as about 2 micrograms of synthetic peptide. Analogs with weaker activity will require more material to achieve the same level of biological effect. Assays for drug efficacy are well known to those skilled in the art and may include in vitro or in vivo assays using synthetic oxytocin as a reference. Atke and Vilhardt Acta Endocrinol.1987:115 (1): 155-60; engstrom et al, eur.J.Pharmacol.1998:355 (2-3): 203-10.

Magnesium-containing oxytocin peptide formulations

In the methods of the invention (including administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ions) for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, social and communication deficits, or anxiety, the oxytocin peptide and magnesium ions may be administered in the form of a magnesium-containing oxytocin peptide formulation or composition. In one aspect, a magnesium-containing oxytocin peptide formulation or composition includes an oxytocin peptide and magnesium ions in amounts that produce a synergistic or enhanced effect when used in treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety.

The relative proportions of oxytocin peptide and magnesium ion in magnesium-containing oxytocin peptide formulations are important in achieving the best synergistic or potentiating effect. The optimal amounts of oxytocin peptide and magnesium ion may depend on the particular disorder or condition, the type of synergistic or potentiating effect desired, and other factors such as the route of administration. For example, the amount of magnesium may be important to achieve a faster onset of action; the amount of oxytocin may be important to achieve a longer lasting effect, and the relative ratio between oxytocin and magnesium may be important to achieve maximum improvement in social function, reduction in social and communication deficits, and/or reduction in anxiety.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 0.01mg/mL and about 16mg/mL of oxytocin peptide. In some embodiments, the amount of oxytocin peptide in the liquid formulation is greater than about (lower limit) 0.01, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 1, or 2mg/mL. In some embodiments, the amount of oxytocin peptide in the liquid formulation is less than about (upper limit) 16, 12, 10, 8, 6, 4,2, 1.6, 1.2, 1, 0.8, 0.6, 0.4, 0.3, 0.2 or 0.1mg/mL. That is, the amount of oxytocin peptide in the liquid formulation is anywhere in the range of about 0.01 to 16mg/mL, with the lower limit being less than the upper limit. In some embodiments of the present invention, the substrate is, the magnesium-containing oxytocin peptide formulation or composition includes between about 0.01mg/mL and about 12mg/mL, between about 0.05mg/mL and about 16mg/mL, between about 0.1mg/mL and about 12mg/mL, between about 0.1mg/mL and about 8mg/mL, between about 0.1mg/mL and about 4mg/mL, between about 0.1mg/mL and about 2mg/mL, between about 0.1mg/mL and about 1.6mg/mL, between about 0.1mg/mL and about 1.2mg/mL, between about 0.1mg/mL and about 1mg/mL, between about 0.1mg/mL and about 0.8mg/mL, between about 0.1mg/mL and about 0.4mg/mL, between about 0.1mg/mL and about 0.3mg/mL, between about 0.2mg/mL and about 16mg/mL, between about 0.2mg/mL and about 12mg/mL, between about 0.2mg/mL, between about 10mg/mL between about 0.2mg/mL and about 8mg/mL, between about 0.2mg/mL and about 6mg/mL, between about 0.2mg/mL and about 4mg/mL, between about 0.2mg/mL and about 2mg/mL, between about 0.2mg/mL and about 1.6mg/mL, between about 0.2mg/mL and about 1.2mg/mL, between about 0.2mg/mL and about 1mg/mL, between about 0.2mg/mL and about 0.8mg/mL, between about 0.2mg/mL and about 0.6mg/mL, between about 0.2mg/mL and about 0.4mg/mL, between about 0.2mg/mL and about 0.3mg/mL, between about 0.3mg/mL and about 16mg/mL, between about 0.3mg/mL and about 12mg/mL, between about 0.3mg/mL and about 10mg/mL, between about 0.3mg/mL, between about 4mg/mL, between about 4.3 mg/mL, oxytocin peptide between about 0.3mg/mL and about 3mg/mL, between about 0.3mg/mL and about 1mg/mL, between about 0.3mg/mL and about 0.5mg/mL, between about 0.5mg/mL and about 16mg/mL, between about 0.5mg/mL and about 10mg/mL, between about 0.5mg/mL and about 5mg/mL, between about 0.5mg/mL and about 1mg/mL, between about 1mg/mL and about 16mg/mL, between about 1mg/mL and about 10mg/mL, or between about 1mg/mL and about 5mg/mL. In preferred embodiments, the magnesium-containing oxytocin peptide formulation or composition comprises between about 0.1mg/mL and about 2mg/mL, between about 0.15mg/mL and about 1.5mg/mL, or between about 0.2mg/mL and about 1.2mg/mL of oxytocin peptide. In one embodiment, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1).

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 5IU/mL and about 8000IU/mL of oxytocin peptide. In some embodiments, the amount of oxytocin peptide in the liquid formulation is greater than about (lower limit) 5, 25, 50, 75, 100, 150, 200, 250, 500, 750, or 1000IU/mL. In some embodiments, the amount of oxytocin peptide in the liquid formulation is less than about (upper limit) 8000, 6000, 5000, 4000, 3000, 2000, 1000, 800, 600, 500, 400, 300, 200, 150, 100, or 50IU/mL. That is, the amount of oxytocin peptide in the liquid formulation is anywhere in the range of about 5 to 8000IU/mL, with the lower limit being less than the upper limit. In some embodiments of the present invention, the substrate is, the magnesium-containing oxytocin peptide formulation or composition includes between about 500IU/mL and about 6000IU/mL, between about 25IU/mL and about 8000IU/mL, between about 50IU/mL and about 6000IU/mL, between about 50IU/mL and about 4000IU/mL, between about 50IU/mL and about 2000IU/mL, between about 50IU/mL and about 1000IU/mL, between about 50IU/mL and about 800IU/mL, between about 50IU/mL and about 600IU/mL, between about 50IU/mL and about 500IU/mL, between about 50IU/mL and about 400IU/mL, between about 50IU/mL and about 200IU/mL, between about 50IU/mL and about 150IU/mL, between about 100IU/mL and about 8000IU/mL, between about 100IU/mL and about 6000IU/mL, between about 100IU/mL and about 5000IU/mL, between about 5000IU/mL between about 100IU/mL and about 4000IU/mL, between about 100IU/mL and about 3000IU/mL, between about 100IU/mL and about 2000IU/mL, between about 100IU/mL and about 1000IU/mL, between about 100IU/mL and about 800IU/mL, between about 100IU/mL and about 600IU/mL, between about 100IU/mL and about 500IU/mL, between about 100IU/mL and about 400IU/mL, between about 100IU/mL and about 300IU/mL, between about 100IU/mL and about 200IU/mL, between about 100IU/mL and about 150IU/mL, between about 150IU/mL and about 8000IU/mL, between about 150IU/mL and about 6000IU/mL, between about 150IU/mL and about 5000IU/mL, between about 150IU/mL and about 4000IU/mL, between about 150IU/mL and about 2000IU/mL, oxytocin peptide between about 150IU/mL and about 1500IU/mL, between about 150IU/mL and about 500IU/mL, between about 150IU/mL and about 250IU/mL, between about 250IU/mL and about 8000IU/mL, between about 250IU/mL and about 5000IU/mL, between about 250IU/mL and about 2500IU/mL, between about 250IU/mL and about 500IU/mL, between about 500IU/mL and about 8000IU/mL, between about 500IU/mL and about 5000IU/mL, or between about 500IU/mL and about 2500 IU/mL. In preferred embodiments, the magnesium-containing oxytocin peptide formulation or composition includes between about 50IU/mL and about 1000IU/mL, between about 75IU/mL and about 750IU/mL, or between about 100IU/mL and about 600IU/mL of oxytocin peptide. In one embodiment, the oxytocin peptide is human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1).

The amount of magnesium present in the formulation may also be in weight percent (w/v) (magnesium or Mg per 100mL of solution ²⁺ In grams), in Mg/mL (magnesium or Mg per mL solution) ²⁺ In milligrams), or in molar concentration ("M" -is defined as magnesium or Mg per liter of solution ²⁺ The number of moles of (a); or "mM" -is defined as magnesium or Mg per liter of solution ²⁺ Millimoles) of the base oil.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a magnesium-containing oxytocin peptide formulation or composition comprising between about 1Mg/mL and about 30Mg/mL of magnesium or magnesium ions (Mg) ²⁺ ) The liquid preparation of (1). In some embodiments, the composition comprises between about 11mg/mL and about 15mg/mL of magnesium or magnesium ions. In some embodiments, the amount of magnesium or magnesium ions in the liquid formulation is greater than about (lower limit) 1,2, 3, 4,5, 6,7, 8, 9, 10, 11, or 12mg/mL. In some embodiments, magnesium or magnesiumThe amount of ions in the liquid formulation is less than about (upper limit) 30, 25, 20, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, or 5mg/mL. That is, the amount of magnesium or magnesium ions in the liquid formulation is anywhere in the range of about 1 to 30mg/mL, with the lower limit being less than the upper limit. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 0.01Mg/mL and about 16Mg/mL (preferably between about 0.1Mg/mL and about 2Mg/mL, more preferably between about 0.15Mg/mL and about 1.5Mg/mL, or about 0.33 Mg/mL) of an oxytocin peptide and between about 1Mg/mL and about 30Mg/mL (or between about 3Mg/mL and about 30Mg/mL, between about 4Mg/mL and about 30Mg/mL, between about 5Mg/mL and about 30Mg/mL, between about 8Mg/mL and about 30Mg/mL, between about 10Mg/mL and about 30Mg/mL, preferably between about 11Mg/mL and about 15Mg/mL, or about 13Mg/mL, or about 12 Mg/mL) of magnesium or Mg ²⁺ . In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a magnesium-containing oxytocin peptide comprising between about 50mM and about 1500mM magnesium or magnesium ions (Mg) ²⁺ ) The liquid preparation of (1). In some embodiments, the amount of magnesium or magnesium ions in the liquid formulation is greater than about (lower limit) 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, or 600mM. In some embodiments, the amount of magnesium or magnesium ions in the liquid formulation is less than about (upper limit) 1500, 1200, 1000, 750, 700, 650, 600, 550, 500, 450, 400, 350, 300, or 250mM. That is, the amount of magnesium or magnesium ions in the liquid formulation is anywhere in the range of about 50 to 1500mM, with the lower limit less than the upper limit. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 5IU/mL and about 8000IU/mL (preferably between about 50IU/mL and about 1000IU/mL, more preferably between about 75IU/mL and about 750IU/mL, or about 150 IU/mL) of oxytocin peptide and between about 1Mg/mL and about 30Mg/mL (preferably between about 11Mg/mL and about 15Mg/mL, or about 13Mg/mL, or about 12 Mg/mL) of magnesium or Mg ²⁺ . In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 5IU/mL and about 8000IU/mL (preferably between about 50IU/mL and about 1000IU/mL, more preferably between about 75IU/mL and about 750IU/mL, or about 150 IU/mL) of oxytocin peptide and between about 50mM and about 1200mM (or between about 100mM and about 1200mM, about 150mM and about 1200 m/mL)M, between about 200mM and about 1200mM, between about 300mM and about 1200mM, between about 400mM and about 1200mM, preferably between about 400mM and about 600mM, or about 500 mM) magnesium or Mg ²⁺ 。

Any magnesium salt (e.g., a water soluble magnesium salt) can be used to provide magnesium ions in the magnesium-containing oxytocin peptide formulation. The magnesium salt for the magnesium-containing oxytocin peptide formulation may be selected based on a number of factors, such as the amount of free magnesium ions that may be delivered upon administration of the formulation, the solubility of the magnesium salt in the medium of the liquid formulation, the acidity/basicity of the counter ion, and/or the dissociation constant of the salt. For example, in liquid formulations, the magnesium salt needs to be sufficiently soluble in the liquid medium to deliver magnesium ions at the concentration required to produce a synergistic or potentiating effect with the oxytocin peptide. Other factors such as compatibility with other materials in the formulation and the ability of the counterion to perform other functions in the formulation may also be considered in selecting the magnesium salt. For example, magnesium citrate can be sufficiently soluble in aqueous solution to provide a desired amount of magnesium or a desired concentration of magnesium ions; citrate is pharmaceutically acceptable; citrate may be part of the buffer; and magnesium citrate may add a palatable taste to the formulation. The magnesium ions in the magnesium-containing oxytocin peptide formulation may be provided by the use of one or more magnesium salts. The magnesium salt in the magnesium-containing oxytocin peptide formulation may be a magnesium salt used initially in the preparation of the magnesium-containing oxytocin peptide formulation or a magnesium salt formed in situ during the preparation of the magnesium-containing oxytocin peptide formulation. For example, magnesium chloride may be used initially in preparing the formulation; and the magnesium citrate may be formed in situ upon addition of citric acid to the formulation. In such an example, the magnesium ions in the magnesium-containing oxytocin peptide formulation are provided by magnesium chloride and magnesium citrate.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition includes one or more magnesium salts selected from the group consisting of magnesium citrate, magnesium chloride, magnesium sulfate, magnesium acetate, magnesium lactate, magnesium stearate, magnesium oxide, magnesium carbonate, magnesium glycinate, magnesium malate, magnesium taurate, magnesium gluconate, magnesium succinate, and magnesium pyrophosphate. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a composition comprising an amount of magnesium salt (e.g., magnesium citrate or magnesium chloride)Thereby providing between about 1mg/mL and about 30mg/mL of magnesium. In some embodiments, the composition includes an amount of magnesium salt to provide between about 1Mg/mL and about 30Mg/mL magnesium ions (Mg) ²⁺ ). In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising an amount of one or more magnesium salts (e.g., magnesium citrate and/or magnesium chloride) providing between about 1Mg/mL and about 30Mg/mL of magnesium or magnesium ions (Mg) ²⁺ ). In some embodiments, the composition includes an amount of one or more magnesium salts to provide between about 11mg/mL and about 15mg/mL of magnesium or magnesium ions. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 0.01Mg/mL and about 16Mg/mL (preferably between about 0.1Mg/mL and about 2Mg/mL, more preferably between about 0.15Mg/mL and about 1.5Mg/mL, or about 0.33 Mg/mL) of an oxytocin peptide and an amount of a magnesium salt (e.g., magnesium citrate or magnesium chloride) to provide between about 1Mg/mL and about 30Mg/mL (or between about 3Mg/mL and about 30Mg/mL, between about 4Mg/mL and about 30Mg/mL, between about 5Mg/mL and about 30Mg/mL, between about 8Mg/mL and about 30Mg/mL, between about 10Mg/mL and about 30Mg/mL, preferably between about 11Mg/mL and about 15Mg/mL, or about 13Mg/mL, or about 12 Mg/mL) of magnesium or Mg ²⁺ . In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 5IU/mL and about 8000IU/mL (preferably between about 50IU/mL and about 1000IU/mL, more preferably between about 75IU/mL and about 750IU/mL, or about 150 IU/mL) of an oxytocin peptide and an amount of one or more magnesium salts (e.g., magnesium citrate and/or magnesium chloride) to provide between about 1Mg/mL and about 30Mg/mL (preferably between about 11Mg/mL and about 15Mg/mL, or about 13Mg/mL, or about 12 Mg/mL) of magnesium or Mg ²⁺ . In some embodiments, the magnesium-containing oxytocin peptide formulation or composition is a liquid formulation comprising between about 5IU/mL and about 8000IU/mL (preferably between about 50IU/mL and about 1000IU/mL, more preferably between about 75IU/mL and about 750IU/mL, or about 150 IU/mL) of an oxytocin peptide and an amount of one or more magnesium salts (e.g., magnesium citrate and/or magnesium chloride) to provide between about 50mM and about 1200mM (or between about 100mM and about 1200mM, between about 150mM and about 1200 mM)200mM and about 1200mM, between about 300mM and about 1200mM, between about 400mM and about 1200mM, preferably between about 400mM and about 600mM, or about 500 mM) magnesium or Mg ²⁺ 。

The relative amounts of oxytocin peptide and magnesium ion in the magnesium-containing oxytocin peptide formulations or compositions described herein can be defined by weight ratio or molar ratio. The weight ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions in the formulation or composition is called the "OT/Mg (w) ratio". For example, in a magnesium-containing oxytocin peptide formulation or composition having an OT/Mg (w) ratio of about 1. The molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions in the formulation or composition is referred to as the "OT/Mg (m) ratio". For example, in a magnesium-containing oxytocin peptide formulation or composition having an OT/Mg (m) ratio of about 1.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition has an OT/Mg (w) ratio between about 1. In some embodiments, the OT/Mg (w) ratio in the formulation or composition is less than about (upper limit) 1. In some embodiments, the OT/Mg (w) ratio in the formulation or composition is greater than about (lower limit) 1. That is, the OT/Mg (w) ratio in the formulation or composition is anywhere in the range of about 1 to 1. In some embodiments, the formulation or composition has a OT/Mg (w) ratio between about 1. In some preferred embodiments, the formulation or composition has a OT/Mg (w) ratio of about 1. In some embodiments, the formulation or composition has an OT/Mg (w) ratio of: between about 1 and about 1 Between about 1 and about 1. In one embodiment, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1).

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition has an OT/Mg (m) ratio of between about 1. In some embodiments, the OT/Mg molar ratio in the formulation or composition is less than about (upper limit) 1. In some embodiments, the OT/Mg molar ratio in the formulation or composition is greater than about (lower limit) 1. That is, the OT/Mg (w) ratio in the formulation or composition is anywhere in the range of about 1. In some embodiments, the formulation or composition has a OT/Mg (m) ratio between about 1. In some preferred embodiments, the formulation or composition has a OT/Mg (m) ratio of about 1. In some embodiments, the formulation or composition has an OT/Mg (m) ratio of: between about 1 and about 200 Between about 1 and about 1200 Between about 1 and about 1, between about 1 and about 4000. In one embodiment, the oxytocin peptide is human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1).

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition comprising an oxytocin peptide and magnesium ions further comprises one or more pharmaceutically acceptable carriers (thus constituting a pharmaceutical composition) and optionally other ingredients such as excipients, vehicles (vehicle), emulsifiers, stabilizers, preservatives, buffers, and/or other additives that may enhance stability, delivery, absorption, half-life, efficacy, pharmacokinetics, and/or pharmacodynamics, reduce adverse side effects, or provide other advantages of pharmaceutical use. Exemplary excipients include solubilizers, surfactants, and chelating agents. For example, the formulation may include methyl- β -cyclodextrin (Me- β -CD), edetate disodium, arginine, sorbitol, naCl, sodium Methyl Paraben (MP), sodium Propyl Paraben (PP), chlorobutanol (CB), benzyl alcohol, zinc chloride, ethanol, didecanoyl L- α -phosphatidylcholine (DDPC), polysorbate, lactose, citrate, tartrate, acetate, and/or phosphate.

Liquid carriers include, but are not limited to, water, saline, aqueous dextrose, and glycols (particularly (when isotonic) for use in solution). The carrier may also be selected from various oils, including those of petroleum, animal, vegetable or synthetic origin (e.g., peanut oil, olive oil, soybean oil, mineral oil, sesame oil, and the like). Suitable pharmaceutical excipients include, but are not limited to, starch, cellulose, talc, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, magnesium stearate, sodium stearate, glycerol monostearate, sodium chloride, dried skim milk powder, glycerol, propylene glycol, water, ethanol and the like. The composition may be subjected to conventional pharmaceutical processes such as sterilization and may contain conventional pharmaceutical additives such as preservatives, stabilizers, reducing agents, antioxidants, chelating agents, wetting agents, emulsifiers, dispersants, gelling agents, salts for adjusting osmotic pressure, buffers and the like. The liquid carrier may be hypotonic or isotonic with body fluids and may have a pH in the range of 3.5-8.5. The use of additives in the preparation of peptide and/or protein based compositions, in particular pharmaceutical compositions, is well known in the art. In some embodiments, the composition has a pH of about 2 to about 7. In some embodiments, the composition has a pH of about 4 to about 7. In a preferred embodiment, the pH of the formulation/composition is about 4.5.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition may further comprise one or more mucosal delivery enhancers selected from the group consisting of (a) - (K): (A) a solubilizer; (B) a charge modifier; (C) a pH controlling agent; (D) a degrading enzyme inhibitor; (E) a mucolytic or mucoclearing agent; (F) cilium stabilizers (ciliostatic agents); (G) a membrane permeation enhancer; (H) Epithelial junction physiological modulators, such as Nitric Oxide (NO) stimulators, chitosan, and chitosan derivatives; (I) a vasodilator; (J) a selective transport enhancer; and (K) a stable delivery vehicle, carrier, support or complexing substance (to which the oxytocin peptide is operatively combined, associated, contained, encapsulated or bound to stabilize the active agent for enhanced mucosal delivery). The membrane penetration enhancer in group (G) may be (i) a surfactant, (ii) a bile salt, (iii) a phospholipid or fatty acid additive, a mixed micelle, liposome, or carrier, (iv) an alcohol, (v) an enamine, (iv) an NO donor compound, (vii) a long-chain amphipathic molecule, (viii) a small hydrophobic penetration enhancer; (ix) sodium or salicylic acid derivatives; (x) Acetoacetic acid glyceride, (xi) cyclodextrin or β -cyclodextrin derivative, (xii) medium chain fatty acid, (xiii) chelating agent, (xiv) amino acid or salt thereof, (xv) N-acetyl amino acid or salt thereof, (xvi) enzyme that degrades into a selected membrane component, (xvii) inhibitor of fatty acid synthesis, (xviii) inhibitor of cholesterol synthesis; or (xiv) any combination of the membrane permeation enhancers of (i) - (xviii). In various embodiments of the invention, the oxytocin peptide may be combined with one, two, three, four or more of the mucosal delivery enhancers described in (a) - (K). These mucosal delivery enhancers may be mixed alone, or with the oxytocin peptide, or otherwise combined with them in a pharmaceutically acceptable formulation or delivery vehicle. The magnesium-containing oxytocin peptide formulations or compositions described herein may provide increased bioavailability of oxytocin peptides upon delivery thereof to a mucosal surface (e.g., in the nasal cavity) of a mammalian subject.

The list of carriers and additives discussed herein is not complete and one skilled in the art can select carriers and excipients from the GRAS (generally recognized as safe) list of chemicals allowed in pharmaceutical preparation as well as those currently allowed by the U.S. food and drug administration in topical and parenteral formulations, and those allowed in the future. (see also Wang et al, (1980) j.parent. Drug asn, 34-462, (1988) j.parent. Sci.and tech, 42.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition comprising an oxytocin peptide and magnesium ions, wherein the oxytocin peptide and magnesium ions are in amounts to produce a synergistic or potentiating effect when used in the treatment of an autism spectrum disorder, further comprises one or more solvents or excipients selected from the group consisting of chlorobutanol, benzalkonium (benzalkonium), methyl 4-hydroxybenzoate, propyl 4-hydroxybenzoate, acetic acid, citric acid, glycerol, sodium chloride, sodium monohydrogen phosphate, sorbitol, and water. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition further comprises chlorobutanol, acetic acid, and water.

In some embodiments, a magnesium-containing oxytocin peptide formulation or composition comprising an oxytocin peptide and magnesium ions further comprises a chitosan-containing excipient (e.g.,http:// www. Archimedesphama. Com/product archidevchisys. Html). In some embodiments, the magnesium-containing oxytocin peptide formulation or composition further comprises about 1% of a chitosan-containing excipient. In some embodiments, chitosan glutamate may be preferred for nasal delivery due to its superior absorption enhancing capacity. In some embodiments, chitosan copolymer nanoparticles may be used, such as nanoparticles containing chitosan glutamate and a negatively charged polymer (e.g., pentasodium tripolyphosphate). Thiolated chitosans (e.g. with 2-iminothiolane)(iminothiolane) covalently modified chitosan), which has been used in microparticles containing insulin and reduced glutathione, may also be used as an excipient in the magnesium-containing oxytocin peptide formulations or compositions described herein.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition comprising an oxytocin peptide and magnesium ions further comprises one or more gelling agents such that the oxytocin peptide formulation forms a gel in the nasal cavity, thereby enhancing nasal absorption of the oxytocin peptide. The gelling system useful in the formulations and methods described herein can include any known gelling system, such as a chemically reactive pectin (pect) -based gelling system (e.g., pecSys) ^TM Archimedes Pharma) and thermoreactive polymer gelation systems (e.g.,F127，BASF)。PecSys ^TM is a low viscosity aqueous pectin based solution delivered as a fine mist, wherein each drop of the gel is in contact with calcium ions in the nasal mucosa. Other low methoxyl pectins may also be used, for example, at a concentration of about 1%.F127 contains an ethylene oxide/propylene oxide block copolymer. The gelation temperature varies depending on the component proportions and the amount of copolymer utilized in the final formulation. At about 18-20% wt/volGelation of F127 in the human nasal cavity has been demonstrated, for example, as used in vitamin B12 gel supplement (Enerb, nature's Bounty, N.Y.) and gelatinized sumatriptan, containing 18% wt/volF127 and 0.3% wt/vol Carbopol (anionic bioadhesive polymer C934P). The monomer ratio and concentration can be adjusted to the intended oxytocin formulation to ensure gelation at 25-37 ℃ (around the typical temperature in the nasal cavity, 34 ℃). If the gelation temperature is lower thanAt 25 ℃, the preparation can be gelled at room temperature; if the gelation temperature is above 37 deg.C, the formulation will not completely gel upon contact with the nasal mucosa. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition may further include a mucoadhesive such as Carbopol. The addition of mucoadhesive (e.g., up to 0.5% carbopol addition) can further reduce the gelation temperature.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition comprising an oxytocin peptide and magnesium ions further comprises a surfactant, such as a non-ionic surfactant (e.g., polysorbate-80), and one or more buffers, stabilizers, or modulators. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition further comprises a propellant. The pH of the nasal spray solution is optionally between about pH 3.0 and 8.5, but the pH is adjusted as desired to optimize delivery of the charged macromolecular species (e.g., therapeutic protein or peptide) in a substantially unionized state. The drug solvent utilized may also be a slightly acidic aqueous buffer solution (pH 3-6). Suitable buffers for use in these compositions are as described above or as otherwise known in the art. Other components may be added to enhance or maintain chemical stability, including preservatives, surfactants, dispersants, or gases. Suitable preservatives include, but are not limited to, phenol, methyl paraben, m-cresol, thimerosal, benzalkonium chloride, and the like. Suitable surfactants include, but are not limited to, oleic acid, sorbitan trioleate, polysorbate, lecithin, phosphatidyl (phosphatidyl) choline, and various long chain diglycerides and phospholipids. Suitable dispersing agents include, but are not limited to, ethylenediaminetetraacetic acid (EDTA) and the like. Suitable gases include, but are not limited to, nitrogen, helium, chlorofluorocarbons (CFCs), hydrofluorocarbons (HFCs), carbon dioxide, air, and the like. Suitable stabilizers and tonicity agents (tonicity agents) include sugars and other polyols, amino acids, and organic and inorganic salts. In some embodiments, the magnesium-containing oxytocin peptide formulation or composition further comprises a citrate salt, a succinate salt, or a pyrophosphate salt.

In some embodiments, the magnesium-containing oxytocin peptide formulation or composition comprising an oxytocin peptide and a magnesium ion further comprises an agent capable of upregulating oxytocin receptor expression, such as IL-6.

To further enhance mucosal delivery of oxytocin peptides, enzyme inhibitors, in particular protease inhibitors, may further be included in the formulation. Protease inhibitors may include, but are not limited to, antinocin (antipain), oxigenin (orphamenine) A and B, anisole hydrochloride, AEBSF, CA-074, calpain inhibitors I and II, calpain inhibitor (calpeptin), pepsin inhibitor A, actinonin, aminopeptidase inhibitor, bestatin, boroleucine (boroleucin), captopril (captopril), chloroacetyl-HOLeu-Ala-Gly-NH ₂ DAPT, dipeptidyl (diprotin) a and B, ebelactone a and B, medroxypin, leupeptin, phosphoramidon, bovine trypsin inhibitor, puromycin, BBI, soy trypsin inhibitor, phenylmethanesulfonyl fluoride, E-64, chymotrypsin inhibitor, 1, 10-phenanthroline, EDTA, and EGTA. Other enzyme inhibitors such as bacitracin may also be included in the formulation.

To enhance delivery and/or absorption of the oxytocin peptide and magnesium ions into or across mucosal surfaces, absorption enhancers may be included in the formulation. These enhancers may enhance the release or solubility (e.g., from the formulation delivery vehicle), diffusion rate, penetration ability and time, uptake, residence time, stability, effective half-life, peak or sustained concentration levels, clearance, and other desirable mucosal delivery characteristics of the composition (e.g., as measured at the delivery site). Thus, enhancement of mucosal delivery may occur by any of a variety of mechanisms, for example, by increasing diffusion, transport, persistence, or stability of oxytocin peptides, increasing membrane fluidity, modulating the availability or action of calcium and other ions that modulate intracellular or paracellular penetration, solubilizing mucosal membrane components (e.g., lipids), altering non-protein and protein sulfhydryl levels in mucosal tissue, increasing water flux across mucosal surfaces, modulating epithelial junction physiology, reducing viscosity of mucus on mucosal epithelium, reducing mucociliary clearance rates, and other mechanisms.

Mucosal absorption enhancing compounds may include, but are not limited to, surfactants, bile salts, dihydrofusidates (dihydrofusidates), bioadhesives/mucoadhesives, phospholipid additives, mixed micelles, liposomes, or carriers, alcohols, enamines, cationic polymers, NO donor compounds, long chain amphiphiles, small hydrophobic penetration enhancers; sodium or salicylic acid derivatives, glycerol acetoacetate, cyclodextrin or β -cyclodextrin derivatives, medium-chain fatty acids, chelating agents, amino acids or salts thereof, N-acetyl amino acids or salts thereof, mucolytic (mucolytic) agents, enzymes specific to selected membrane components, fatty acid synthesis inhibitors and cholesterol synthesis inhibitors.

All of the peptides described and/or contemplated herein can be prepared by chemical synthesis using automated or manual solid phase synthesis techniques generally known in the art. Peptides may also be prepared using molecular recombinant techniques known in the art.

Delivery system

The magnesium-containing oxytocin peptide formulation or composition may be suitable for craniofacial mucosal administration (e.g., nasal, buccal, sublingual, or ocular administration). In some embodiments, the composition may further comprise a device for mucosal delivery. In some embodiments, the composition is suitable for buccal and/or sublingual mucosal delivery, which may further comprise devices for buccal and/or sublingual mucosal administration, such as unit dose containers, pump sprayers, droppers (droppers), squeeze bottles, airless and preservative-free nebulizers, dose inhalers, and pressurized dose inhalers. In some embodiments, the composition is suitable for ocular delivery, which may further comprise a device for conjunctival administration, such as a dropper or squeeze bottle. In some embodiments, the composition is suitable for intranasal administration, which may further comprise an intranasal administration device such as a dropper, pump spray, squeeze bottle, airless and preservative-free nebulizer, or a nasal pump device, e.g., a nasal pump device comprising a container bottle attached to an aerosolizer.

Intranasal drug delivery has been the subject of research and development for many years, although vehicle systems for efficient delivery of substances have been designed only in the last decade. (Sayani and Chien, clinical Reviews in therapeutic drug carriers Systems 1996, 13. Intranasal delivery has several advantageous features, including relatively high bioavailability, rapid absorption kinetics, and avoidance of first-pass effects in the liver. In some aspects, intranasal administration may allow for delivery of the oxytocin peptide to the nasal cavity, while in other aspects, intranasal administration may allow for targeted delivery to the nose and/or cranial nerves of the brain. Without wishing to be bound by any particular theory, intranasal administration of oxytocin peptide may target the olfactory nervous system or the trigeminal nervous system or both. The oxytocin peptide may be delivered intranasally in any applicable form, including but not limited to a liquid formulation, a solid formulation (e.g., a dry powder formulation), a gel formulation, or an emulsion formulation.

In embodiments where the combination of oxytocin and magnesium ions is administered intranasally, the composition may be prepared as a liquid aerosol formulation in combination with a dispersant and/or a pharmaceutically acceptable diluent. Alternatively, dry powder aerosol formulations are contemplated and may contain the subject compounds and dispersing agents in finely divided solid form, allowing for easy dispersion of the dry powder particles. In the case of liquid or dry powder aerosol formulations, the formulation is aerosolized into small, liquid or solid particles in order to ensure that the aerosolized dose reaches the mucosa of the nasal passages or lungs. The term "aerosol particles" is used herein to describe liquid or solid particles suitable for sufficiently small particle size for nasal (in the range of about 10 microns) or pulmonary (in the range of about 2-5 microns) distribution to the targeted mucosal or alveolar membrane. Other considerations include the structure of the delivery device, additional components in the formulation, and particle characteristics. These aspects of nasal or pulmonary administration of drugs are well known in the art, and the handling of the formulation, the aerosolization means, and the structure of the delivery device are within the level of ordinary skill in the art.

In some embodiments, the magnesium-containing oxytocin peptide formulations or compositions useful in the methods described herein (in which the oxytocin peptide and magnesium ions are in amounts that produce a synergistic or enhanced effect when used in treating an autism spectrum disorder) are administered using a device for intranasal delivery. The device may be any device suitable for intranasal administration of a magnesium-containing oxytocin peptide formulation. In some embodiments, the device is adapted to deliver oxytocin peptides and magnesium ions to specific regions within the nasal cavity. In some embodiments, the device is adapted to deliver oxytocin peptide and magnesium ions to the lower two thirds of the nasal cavity. In some embodiments, the device is adapted to deliver oxytocin peptide and magnesium ions to the upper third of the nasal cavity. In some embodiments, the device is adapted to deliver oxytocin peptides throughout the nasal passage.

In some embodiments, the device for intranasal delivery is a nasal pump device. In some embodiments, the nasal pump device includes a container bottle attached to a pump actuator. In some embodiments, the pump actuator is metered to deliver a specified volume (e.g., about 5 to about 1000 μ Ι _, preferably about 50 to about 150 μ Ι _, more preferably about 50 μ Ι _orabout 100 μ Ι _) at a specified droplet distribution size. In some embodiments, the nasal pump device comprises a container bottle attached to an aerosolizer, such as an equel pump sold by Aptar Pharma. In some embodiments, the means for nasal administration is active regardless of the pressure applied to the pump once the threshold is reached. In some embodiments, the device for nasal administration is a mucosal aerosolization device that can be added to a syringe (e.g.,MAD NASAL ^TM ). For administration in large mammals, a nasal pump device may include a container bottle attached to a pump actuator that is metered to deliver a larger volume (e.g., about 100 μ Ι _ to about 600 μ Ι _ or higher).

In some embodiments, the device for intranasal delivery is designed to deliver multiple doses of the pharmaceutical formulation. For example, a nasal pump device may include a container vial attached to a pump actuator, where the container vial contains multiple doses of a liquid formulation and the pump actuator is metered to deliver a specified volume that is part of the liquid formulation contained in the container vial. In some embodiments, the pump actuator is metered to deliver about 50 μ Ι _ of liquid formulation per spray. The nasal pump device may include a filter for preventing backflow to reduce contaminants (e.g., bacteria) from entering the container bottle. In some embodiments, the nasal pump device includes a metal-free pathway for delivering the liquid formulation (e.g., a plastic pathway). In some embodiments, the pump device utilizes a plastic material that is stable to gamma radiation (used to sterilize the nasal device). In some embodiments, the device for intranasal delivery is equipped with a multi-dose pump that includes a microbial filter and an automatic occlusion mechanism in the pump actuator, e.g., a spray device as described in U.S. patent No. 5,988,449.

In some embodiments, the device for intranasal delivery is a breath-actuated nasal delivery device, such as the devices described in U.S. Pat. nos. 7,784,460 and 7,854,227. Such a device may improve delivery to a target site deep within the nasal cavity. In some embodiments, a standard metered dose spray device is incorporated into a housing that allows a patient to insufflate an mouthpiece (mouthpiece) to actuate the device. In some embodiments, the device consists of a tapered sealing nosepiece (nosepiece) and mouthpiece incorporated into a conventional mechanical spray pump (e.g., an equel pump sold by Aptar Pharma), a chargeable spring, and a breath actuation mechanism. The system may be used for single dose or multiple dose delivery. One example of such a liquid delivery device is OptiMist sold by OptiNose ^TM Provided is a device. In use, the nosepiece of the device is inserted into a nostril and the mouthpiece is insufflated. This closes the soft palate, delivers pressure to the nostrils, opens a channel that provides airflow behind the nasal septum and allows air to leave the other nostril (bi-directional flow). Since the device is breath actuated, small particles cannot enter the lungs. Modification of flow rate and particle size allows for targeting of specific nasal regions.

In some embodiments, the device for intranasal delivery is a unit dose metered spray device suitable for a single administration of the magnesium-containing oxytocin peptide formulation or composition. In some embodiments, the device for intranasal delivery is a multi-dose metered spray pump device suitable for repeated administration of oxytocin peptides.

Droplet size, plume volume, and flow rate may be modified to target specific nasal regions. The liquid nebulizer may provide a droplet size of between 5 and 50 microns in order to target the olfactory and/or respiratory epithelium. The larger droplets move primarily down the nasopharynx and are swallowed, while the smaller droplets are targeted to the lung tissue. Mass Median Equivalent Aerodynamic Diameter (Mass media Equivalent Aerodynamic Diameter, MMAD) was used to specify the droplet size. The pH of the nasal spray is optimized to deliver the charged peptide in a largely unionized state. The nose will typically tolerate solutions having a pH of about 3-8. The nasal mucosa can typically absorb a volume of about 100 μ L before saturation occurs and liquid begins to spill out of the nose. Thus, the plume volume may be up to (and including) 100 μ L. For use in large mammals, the plume volume may be up to (and include) 150 μ L or more (e.g., 600 μ L or more). For infant use, or veterinary use in smaller animals (e.g., rodents, cats), a smaller plume volume (5-50 μ Ι _ can be used.

In some embodiments, the device for intranasal delivery is ergonomically designed to promote patient compliance, such as a pump device with a side-actuated trigger mechanism. In some embodiments, the device for intranasal delivery comprises a metered spray pump that operates as a closed system that does not allow air to enter the pump device to prevent contamination from airborne pathogens. In some embodiments, the device for intranasal delivery comprises a metered spray pump working with a filter. Ventilating air is drawn through a filter fitted within the pump to keep airborne pathogens out of the pump unit. In some embodiments, the intranasal delivery device including the nasal pump device may further include a microelectronic device that may facilitate data transmission and therapy monitoring.

In some embodiments, a magnesium-containing oxytocin peptide formulation or composition comprises an oxytocin peptide and a magnesium ion, wherein the oxytocin peptide and the magnesium ion are contained in any one of the devices for intranasal delivery described herein, and wherein the concentration of the oxytocin peptide and the magnesium ion is within any of the concentration ranges described herein as if each and every combination of the devices and concentrations were described separately.

Method

The term "Autism Spectrum Disorder (ASD)" or "autism" refers to a complex group of brain developmental disorders. These disorders manifest to varying degrees as difficulties in social interaction, verbal and nonverbal communication, and repetitive behaviors. With respect to the fifth edition Diagnostic and Statistical Manual of Mental Disorders (DSM-5), published in 5.2013, all autism Disorders were combined as an umbrella diagnosis of ASD. Previously, they were considered to be distinct subtypes, including autism, childhood disintegrations, pervasive developmental disorders-not otherwise specified (PDD-NOS) and Asperger syndrome. See http:// www.austismseeds.org/what-austism. Those skilled in the art will recognize that the symptoms of autism spectrum disorders overlap considerably with many other psychiatric disorders. Examples of disorders exhibiting symptoms similar to those shown in autism spectrum disorders include, but are not limited to, social anxiety disorder, obsessive compulsive disorder, social (pragmatic) communication disorders, and neurodevelopmental disorders including, but not limited to, attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, frangle-X syndrome, rett syndrome, and Williams syndrome.

DSM-5 provides diagnostic criteria for ASD, including: (A) Persistent barriers in social communications and social interactions across multiple contexts, as currently or historically shown by the following illustrative examples: (1) Defects in social-emotional reciprocity (social-emotional recency), for example, range from abnormal social methods and failure of normal back-and-forth conversations; to reduce sharing interests, emotions or feelings; to failure to initiate or respond to a social interaction; (2) Deficiencies in non-verbal communication behavior for social interaction, for example, ranging from poorly integrated verbal and nonverbal communication; abnormalities in eye contact and body language or deficiencies in understanding and use of gestures; to complete lack of facial expression and non-verbal communication; and (3) deficiencies in developing, maintaining, and understanding relationships, e.g., difficulties ranging from adjusting behavior to adapting to various social situations; to difficulty in sharing imaginative games or friendships; to lack of interest in the peer; and (B) restricted, repetitive behaviors, interests, or activity patterns, as presently or historically shown by at least two of the following illustrative examples: (1) Stereotypy or repetitive motion, use of a subject, or speech (e.g., simple motor stereotypies, arraying toys or turning objects, imitating speech, monster phrases); (2) Adherence to identity, inflexibly adherence to regular, or ceremonial patterns or verbal non-verbal behavior (e.g., extreme pain with small changes, difficulty in transition, rigid thinking patterns, greeting ceremonies, need to take the same route or eat food daily); (3) A high limit of intensity or focus anomalies, a fixed interest (e.g., strong adherence or concentration to an anomalous object, over-restriction, or sustained interest); and (4) high or low reactivity to sensory input or unusual interest in environmental sensory aspects (e.g., significant apathy to pain/temperature, adverse reactions to specific sounds or textures, excessive sniffing or touching of objects, visual fascination of light or movement). See http:// www.austspeaks.org/what-aust/diagnosis/dsm-5-diagnostic-criticia.

Autism Spectrum Disorder (ASD) is characterized by difficulties in social interaction, communication challenges, and a tendency to engage in repetitive behaviors. However, the symptoms and their severity vary widely among these three core regions. ASD may be associated with intellectual disability, motor coordination difficulties and attention and physical health problems such as sleep and gastrointestinal disturbances. ASD may be associated with psychiatric symptoms including anxiety and depression. See, for example, kim et al, autism2000,4 (2): 117-132.

Oxytocin is known to treat a variety of conditions including anxiety and social and communication deficits in autistic spectrum disorders. However, the effect of oxytocin in the treatment of social and communication deficits in autism spectrum disorders has been observed to vary greatly between patients. Changes in the receptor availability and receptor affinity of oxytocin may be responsible for the changes in effect. Commercial preparations of oxytocin are used (e.g.,) Clinical efforts to treat ASD are compromised by lack of efficacy and poor tolerability. Due to the low efficiency of currently available oxytocin formulationsAnd high volume, so that the amount of drug absorbed is insufficient to exert efficacy when administered by nasal spray. The present invention provides methods of administering oxytocin peptides in more effective formulations and lower volumes such that an effective amount of the formulation can be delivered using a nasal device for treating autism spectrum disorders, disorders exhibiting one or more symptoms associated with autism spectrum disorders, or social and communication deficits.

In one aspect, provided is a method for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering to a subject in need thereof an effective amount of oxytocin peptide and magnesium ion, wherein the effective amount is delivered by intranasal administration of a volume that is readily absorbed in the nasal cavity. In some embodiments, the volume in which the oxytocin peptide and magnesium ions are readily absorbed in the nasal cavity is between about 5 μ L and about 1000 μ L. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is about 1.

In one aspect, provided is a method comprising administering to a subject in need thereof an effective amount of an oxytocin peptide and a magnesium ion, wherein the effective amount is delivered by intranasal administration in a volume of between about 5 μ Ι _ and about 1000 μ Ι _. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is about 1. In some embodiments, the methods are used to treat one or more symptoms associated with an autism spectrum disorder. In some embodiments, the methods are used to treat a disorder exhibiting one or more symptoms associated with an autism spectrum disorder. In some embodiments, the method is used to reduce social and communication deficits. In some embodiments, the methods are used to treat or reduce anxiety.

Magnesium is involved in many aspects of life and health, such as energy production, oxygen uptake, central nervous system function, electrolyte balance, glucose metabolism, and muscle activity. Magnesium has also been found to be clinically effective in reducing social and communication deficits in children with autism spectrum disorders. See Mousain-Bosc et al, magnes. Res.2006,19 (1): 53-62. Co-administration of oxytocin and magnesium ions according to the invention results in a synergistic or enhanced social behaviour improvement and anxiety reduction relative to oxytocin administration alone. The underlying mechanism of these effects is not clear, but may involve noncompetitive blockade of N-methyl D-aspartate (NMDA) neurotransmitter receptors, or an increase in the affinity of oxytocin receptor action as an allosteric modulator, or both.

In some aspects, provided are methods for treating autism spectrum disorders, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect. In some aspects, provided are methods for alleviating or reducing one or more symptoms associated with an autism spectrum disorder, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect. In some aspects, provided are methods for treating a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or enhanced effect. In some embodiments, the oxytocin peptide and magnesium ion are administered in a dose that produces an overall effect of alleviating or reducing symptoms that is greater than the sum of the effects of the oxytocin peptide and magnesium salt administered alone in equal doses. In some embodiments, the oxytocin peptide and the magnesium ion are administered at doses that produce a faster onset and/or longer duration of effect than would occur following administration of each agent alone in equal amounts. Examples of symptoms associated with autism spectrum disorders include, but are not limited to, persistent deficits in social communication and social interactions, social anxiety, and limited repetitive behaviors, interests, and activities. Other behaviors and characteristics also observed in patients with autism spectrum disorders include aversion to physical contact, generalized anxiety, monotonous or unregulated volume of sounds, failure to develop companion relationships, lack of shared enjoyment and interest, and lack of social or emotional reciprocity. Examples of disorders that present symptoms similar to those exhibited in autism spectrum disorders include, but are not limited to, social anxiety disorder, obsessive compulsive disorder, social (Prader-Willi) communication disorders, and neurodevelopmental disorders including, but not limited to, attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, fragile-X syndrome, rett syndrome, and Williams syndrome.

Prader-Willi syndrome is a complex genetic condition that affects many parts of the body and is caused by loss of gene function in specific regions of chromosome 15. Individuals with Prader-Willi syndrome often have mild to moderate intellectual impairment and learning difficulties, as well as many exhibiting behavioral problems including splenic agitation, stubborn, manipulation behavior, and compulsive behavior including scratching the skin. Other symptoms often observed in individuals with Prader-Willi syndrome are persistent deficits in social communication and social interaction, anxiety and irritability, and sleep problems.

In some aspects, provided are methods for treating Prader-Willi syndrome comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions. In some embodiments, the co-administration of the oxytocin peptide and the magnesium ion produces a synergistic or potentiating effect. In some embodiments, the oxytocin peptide and the magnesium ion are administered in a dose that produces an overall effect that is greater than the sum of the effects of administering an equivalent dose of the oxytocin peptide and the magnesium salt alone. In some embodiments, the oxytocin peptide and the magnesium ion are administered in doses that produce a faster onset and/or longer duration of effect than would occur following administration of each agent alone in equal amounts. In one embodiment, the invention provides a method for treating Prader-Willi syndrome comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions, wherein the effective dose of the oxytocin peptide and magnesium ions are administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is from about 1.

In some aspects, provided are methods for alleviating or reducing one or more symptoms associated with Prader-Willi syndrome, comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions. Examples of symptoms associated with Prader-Willi syndrome include, but are not limited to, persistent deficits in social communication and social interaction, anxiety and irritability, and sleep problems. In some embodiments, the co-administration of the oxytocin peptide and the magnesium ion produces a synergistic or potentiating effect. In some embodiments, the oxytocin peptide and magnesium ion are administered in a dose that produces an overall effect of alleviating or reducing symptoms that is greater than the sum of the effects of the oxytocin peptide and magnesium salt administered alone in equal doses. In some embodiments, the oxytocin peptide and the magnesium ion are administered in doses that produce a faster onset and/or longer duration of effect than would occur following administration of each agent alone in equal amounts. Examples of symptoms associated with Prader-Willi syndrome include, but are not limited to, persistent deficits in social communication and social interaction, anxiety and irritability, and sleep problems. In one embodiment, the invention provides a method for treating Prader-Willi syndrome comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ions, wherein the effective dose of oxytocin peptide and magnesium ions are administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is about 1.

In some aspects, provided are methods for treating anxiety associated with Prader-Willi syndrome comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions. In some embodiments, the co-administration of the oxytocin peptide and the magnesium ion produces a synergistic or potentiating effect. In some embodiments, the oxytocin peptide and the magnesium ion are administered in a dose that produces an overall effect of relieving or reducing anxiety that is greater than the sum of the effects of administering an equivalent dose of the oxytocin peptide and the magnesium salt alone. In some embodiments, the oxytocin peptide and the magnesium ion are administered in doses that produce a faster onset and/or longer duration of effect than would occur following administration of each agent alone in equal amounts. In one embodiment, the invention provides a method for treating anxiety associated with Prader-Willi syndrome comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions, wherein the effective dose of the oxytocin peptide and magnesium ions are administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is about 1.

In one aspect, the invention provides a method for treating social and communication deficits comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and magnesium ions, wherein co-administration of the oxytocin peptide and magnesium ions produces a synergistic or potentiating effect. In one aspect, the invention provides a method for treating anxiety comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and the magnesium ion produces a synergistic or potentiating effect. In some embodiments, the oxytocin peptide and the magnesium ion are administered in a dose that produces an overall effect of reducing social and communication deficits and/or anxiety that is greater than the sum of the effects of the oxytocin peptide and the magnesium salt administered alone in equal doses. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is from about 1. In some embodiments, social and communication deficits are deficits in communication skills and/or social interactions, lack of eye contact, and/or inability to form and/or maintain social relationships.

In some aspects, provided are methods for treating anxiety associated with autism spectrum disorders comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ions. In some embodiments, the co-administration of the oxytocin peptide and the magnesium ion produces a synergistic or potentiating effect. In some embodiments, the oxytocin peptide and the magnesium ion are administered in a dose that produces an overall effect of relieving or reducing anxiety that is greater than the sum of the effects of administering an equivalent dose of the oxytocin peptide and the magnesium salt alone. In some embodiments, the oxytocin peptide and the magnesium ion are administered in doses that produce a faster onset and/or longer duration of effect than would occur following administration of each agent alone in equal amounts. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is from about 1.

The oxytocin peptide and the magnesium ion may be administered simultaneously or sequentially. In some embodiments, the oxytocin peptide and the magnesium ion are administered simultaneously in the same unit dose. In some embodiments, the oxytocin peptide is administered simultaneously with the magnesium ion but in separate unit doses or formulations. In some embodiments, the oxytocin peptide and the magnesium ion are administered sequentially. In some embodiments, the magnesium ion is administered to the subject in a first administration, and the oxytocin peptide is subsequently administered to the subject in a second administration. In some of these embodiments, the oxytocin peptide is administered between about 10 minutes and about 2 hours after the administration of the magnesium ion. In some of these embodiments, the oxytocin peptide is administered between about 10 minutes and about 2 hours, between about 10 minutes and about 1 hour, between about 10 minutes and about 30 minutes, between about 20 minutes and about 2 hours, between about 20 minutes and about 1 hour, between about 30 minutes and about 2 hours, or between about 30 minutes and about 1 hour after the administration of the magnesium ion. In some of these embodiments, the oxytocin peptide is administered about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 60 minutes, about 90 minutes, or about 120 minutes after the administration of the magnesium ion. In some of these embodiments, the oxytocin peptide is administered about 10 minutes, about 15 minutes, about 20 minutes, or about 30 minutes after the administration of the magnesium ion. In one embodiment, the oxytocin peptide is administered to the subject first, and the magnesium ion is subsequently administered to the subject. In some embodiments, the subject is a human.

Interleukin-6 (IL-6) has been demonstrated to cause an increase in oxytocin receptor expression in various tissues (e.g., young et al, j. Neuroendocrinology,1997 859-65. Thus, serum IL-6 levels can be used as a biomarker of the potential efficacy of oxytocin, for example, when administered nasally with magnesium.

In some aspects, IL-6 is used as a biomarker of efficacy of administration of oxytocin peptide in a subject according to the methods described herein for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety; and for selecting subjects for use in the method. In some embodiments, IL-6 is used to select a subject (e.g., a human) for administration of the oxytocin peptide (e.g., nasal administration of the oxytocin peptide in combination with magnesium ions).

In some embodiments, the subject is selected for treatment based on the subject having a high level of IL-6. The level of IL-6 may be high compared to a control or reference. In some embodiments, the level of IL-6 is higher than a control or reference if the level of IL-6 is determined to be significantly greater than the control or reference by appropriate statistical analysis. In some embodiments, if the level of IL-6 is greater than the control or reference by at least one standard deviation, then the level of IL-6 is greater than the control or reference. In some embodiments, the control is a level value of IL-6 determined in an age and gender matched healthy subject. In some embodiments, the reference is a reported value for IL-6 level, such as a reported value for IL-6 in an age and gender matched healthy subject. In some embodiments, the level of IL-6 is determined as the level of IL-6 in a sample (e.g., a tissue or fluid sample) from the subject, including but not limited to whole blood, serum, plasma, tears, and the like. IL-6 levels in a sample can be determined by any method known in the art, for example by an immunoassay, such as an ELISA-based assay. See, e.g., yang, C-j., et al neuroscience 284, 290-296,2015; emanuule, E., et al, neurosciences letters 471 (3): 162-165,2010; ashwood, P., et al, brain, behavior, and immunity 25 (1): 40-45,2011; and Malik, M., et al, immunology 216 (1): 80-85,2011.

In some embodiments, a method for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprises measuring a level of IL-6 (e.g., a serum level of IL-6) in a subject and administering an effective dose of an oxytocin peptide and magnesium ions to a subject with a high IL-6 level.

In one aspect, a method for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect, further comprises administering to the subject an effective amount of interleukin-6 (IL-6). In some embodiments, an effective amount of IL-6 results in increased expression of oxytocin receptor (OTR) in a subject.

In some embodiments, the oxytocin peptide and IL-6 may be administered simultaneously or sequentially according to any of the methods described herein in which IL-6 is administered to a subject. In some embodiments, the oxytocin peptide is administered simultaneously with IL-6 in the same unit dose. In some embodiments, the oxytocin peptide is administered simultaneously with IL-6 but in separate unit doses or formulations. In some embodiments, the oxytocin peptide and IL-6 are administered sequentially. In some embodiments, IL-6 is administered to the subject in a first administration, and the oxytocin peptide is subsequently administered to the subject in a second administration. In some of these embodiments, the oxytocin peptide is administered between about 1 minute and about 4 hours after the administration of IL-6. In some of these embodiments, the oxytocin peptide is administered between about 1 minute and about 4 hours, between about 10 minutes and about 3 hours, between about 10 minutes and about 2 hours, between about 10 minutes and about 1 hour, between about 10 minutes and about 30 minutes, between about 20 minutes and about 4 hours, between about 20 minutes and about 3 hours, between about 20 minutes and about 2 hours, between about 20 minutes and about 1 hour, between about 30 minutes and about 4 hours, between about 30 minutes and about 3 hours, between about 30 minutes and about 2 hours, or between about 30 minutes and about 1 hour after the administration of IL-6. In some of these embodiments, the oxytocin peptide is administered about 1 minute, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 60 minutes, about 90 minutes, about 120 minutes, about 150 minutes, about 180 minutes, about 210 minutes, or about 240 minutes after the administration of IL-6. In some of these embodiments, the oxytocin peptide is administered about 10 minutes, about 15 minutes, about 20 minutes, or about 30 minutes after the administration of IL-6. In one embodiment, the oxytocin peptide is administered to the subject first and the IL-6 is subsequently administered to the subject. In some embodiments, the subject is a human. In some of these embodiments, the magnesium ion is administered simultaneously with the oxytocin peptide and/or IL-6 prior to either or both of the oxytocin peptide and IL-6 or after either or both of the oxytocin peptide and IL-6.

The oxytocin peptide and the magnesium ion can be administered to a subject in need thereof by the same route or different routes. In some embodiments, the oxytocin peptide is administered by craniofacial mucosal administration (e.g., nasal, buccal, sublingual, or ocular administration). In one embodiment, both the oxytocin peptide and the magnesium ion are administered intranasally in the same formulation. In one embodiment, the oxytocin peptide is administered through the craniofacial mucosa and the magnesium ion is administered systemically, e.g., intravenously, intramuscularly, orally, subcutaneously, or intrathecally.

In some embodiments, the oxytocin peptide is administered by intranasal administration. In some embodiments, the oxytocin peptide and the magnesium ion are administered by intranasal administration. Oxytocin peptide and/or magnesium ions can be administered to mucosal tissue within the nasal cavity using a suitable device for intranasal delivery, such as the nasal delivery devices described herein. Suitable areas within the nasal cavity include, but are not limited to, the lower two-thirds of the nasal cavity, or the upper third, or the entire nasal passage. In some embodiments, the oxytocin peptide and/or magnesium ion is administered into the upper third of the nasal cavity. In some embodiments, the oxytocin peptide and/or magnesium ion is administered into the lower two thirds of the nasal cavity. In some embodiments, the oxytocin peptide and/or magnesium ion is specifically administered to reach both the lower two-thirds and the upper one-third of the nasal cavity. In some embodiments, methods are provided for treating an autism spectrum disorder, one or more symptoms associated with an autism spectrum disorder, or a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, comprising intranasally administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect. In some embodiments, the methods are used to treat social and communication deficits or anxiety.

In some embodiments, according to any of the methods described herein, wherein IL-6 is administered to the subject, IL-6 is administered by intranasal administration. IL-6 can be administered to mucosal tissue within the nasal cavity using a suitable device for intranasal delivery, such as the nasal delivery devices described herein. In some embodiments, IL-6 is administered systemically, e.g., intravenously, intramuscularly, orally, subcutaneously, or intrathecally.

In some embodiments, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1). In some embodiments, the effective dose of oxytocin peptide is about 0.5 μ g to about 2000 μ g. In some embodiments, the effective dose of oxytocin peptide is about 0.5 μ g to about 1000 μ g, about 1 μ g to about 1000 μ g, or about 1 μ g to about 2000 μ g. In some embodiments of the present invention, the substrate is, an effective dose of oxytocin peptide is about 4 μ g to about 1000 μ g, about 8 μ g to about 800 μ g, about 8 μ g to about 500 μ g, about 8 μ g to about 400 μ g, about 8 μ g to about 300 μ g, about 8 μ g to about 200 μ g, about 8 μ g to about 100 μ g, about 8 μ g to about 80 μ g, about 8 μ g to about 50 μ g, about 10 μ g to about 1000 μ g, about 10 μ g to about 500 μ g, about 10 μ g to about 200 μ g, about 10 μ g to about 100 μ g, about 16 μ g to about 1000 μ g, about 16 μ g to about 800 μ g, about 16 μ g to about 500 μ g, about 16 μ g to about 400 μ g, about 16 μ g to about 200 μ g, about 16 μ g to about 160 μ g, about 16 μ g to about 120 μ g, about 16 μ g to about 80 μ g, about 20 μ g to about 1000 μ g, about 20 μ g to about 800 μ g, about 20 μ g to about 500 μ g, about 20 μ g to about 200 μ g, about 20 μ g to about 100 μ g, about 30 μ g to about 1000 μ g, about 30 μ g to about 500 μ g, about 30 μ g to about 300 μ g, about 30 μ g to about 120 μ g, about 30 μ g to about 90 μ g, about 50 μ g to about 1000 μ g, about 50 μ g to about 500 μ g, about 50 μ g to about 250 μ g, about 50 μ g to about 100 μ g, or about 50 μ g to about 80 μ g. In some embodiments, the effective dose of oxytocin peptide is about 8 μ g, about 16 μ g, about 32 μ g, about 48 μ g, about 64 μ g, about 80 μ g, about 96 μ g, about 128 μ g, about 256 μ g, about 10 μ g, about 20 μ g, about 30 μ g, about 40 μ g, about 50 μ g, about 60 μ g, about 70 μ g, about 80 μ g, about 90 μ g, about 100 μ g, about 120 μ g, about 150 μ g, about 200 μ g, about 400 μ g, about 600 μ g, about 800 μ g, or about 100 μ g. In preferred embodiments, the effective dose of oxytocin peptide is about 8 μ g to about 120 μ g, about 15 μ g to about 120 μ g, about 30 μ g to about 120 μ g, or about 66 μ g.

In some embodiments, the effective dose of oxytocin peptide is about 0.25IU to about 1000IU. In some embodiments, the effective dose of oxytocin peptide is about 0.25IU to about 500IU, about 0.5IU to about 500IU, or about 0.5IU to about 1000IU. In some embodiments of the present invention, the substrate is, an effective dose of oxytocin peptide is about 2IU to about 500IU, about 4IU to about 400IU, about 4IU to about 250IU, about 4IU to about 200IU, about 4IU to about 150IU, about 4IU to about 100IU, about 4IU to about 50IU, about 4IU to about 40IU, about 4IU to about 25IU, about 5 to about 500IU, about 5IU to about 250IU, about 5IU to about 100IU, about 5 to about 50IU, about 8IU to about 500IU, about 8IU to about 400IU, about 8IU to about 250, about 8IU to about 200IU, about from about 8IU to about 100IU, from about 8IU to about 80IU, from about 8IU to about 60IU, from about 8IU to about 40IU, from about 10IU to about 500IU, from about 10IU to about 400IU, from about 10IU to about 250IU, from about 10IU to about 100IU, from about 10IU to about 50IU, from about 15IU to about 500IU, from about 15IU to about 250IU, from about 15IU to about 150IU, from about 15IU to about 60IU, from about 15IU to about 45IU, from about 25IU to about 500IU, from about 25IU to about 250IU, from about 25IU to about 125, from about 25IU to about 50, or from about 25IU to about 40IU. In some embodiments, the effective dose of oxytocin peptide is about 4IU, about 8IU, about 16IU, about 24IU, about 32IU, about 40IU, about 48IU, about 64IU, about 128IU, about 5IU, about 10IU, about 15IU, about 20IU, about 25IU, about 30IU, about 35IU, about 40IU, about 45IU, about 50IU, about 60IU, about 75IU, about 100IU, about 200IU, about 300IU, about 400, or about 50IU. In preferred embodiments, the effective dose of oxytocin peptide is about 4IU to about 60IU, about 7.5IU to about 60IU, about 15IU to about 60IU, or about 30IU.

In one embodiment, the dose or amount of oxytocin in the combination is effective to provide a clinically measurable improvement in the symptoms of an autism spectrum disorder or related disorder. The combination of oxytocin and magnesium ions provides a synergistic or potentiating effect to ameliorate autism spectrum disorders or related disorders. In some embodiments, oxytocin is administered in a sub-therapeutically effective dose relative to the dose of oxytocin administered as a single agent. The dose of oxytocin as a single dose depends in part on the route of administration. Thus, the dosage of oxytocin in the combination therapies described herein will also depend in part on the route of administration.

The optimal dosage of magnesium ion may depend on the particular disorder or condition, the type of synergistic or potentiating effect desired, and other factors such as the route of administration. The optimum dosage may be measured in the total amount of magnesium ions administered, or the concentration of magnesium ions in the formulation administered. In some embodiments, an effective dose of magnesium ions administered is from about 50 μ g to about 68mg. In some embodiments, an effective dose of magnesium ions administered is from about 50 μ g to about 34mg, or from about 1mg to about 3mg. In some embodiments, an effective dose of magnesium ions administered is about 1.3mg, or about 2.6mg. In some embodiments, the effective dose of magnesium ions administered is about 1.2mg, or about 2.4mg. In some embodiments of the present invention, the substrate is, an effective dose of magnesium ions administered is about 50 μ g to about 17mg, about 50 μ g to about 8mg, about 50 μ g to about 4mg, about 50 μ g to about 2mg, about 50 μ g to about 1mg, about 50 μ g to about 500 μ g, about 100 μ g to about 68mg, about 100 μ g to about 34mg, about 100 μ g to about 17mg, about 100 μ g to about 8mg, about 100 μ g to about 4mg, about 100 μ g to about 2mg, about 100 μ g to about 1mg, about 100 μ g to about 500 μ g, about 200 μ g to about 68mg, about 200 μ g to about 34mg, about 200 μ g to about 17mg, about 200 μ g to about 8mg, about 200 μ g to about 4mg, about 200 μ g to about 2mg, about 200 μ g to about 1mg, about 200 μ g to about 500 μ g, about 500 μ g to about 68mg about 500 μ g to about 34mg, about 500 μ g to about 17mg, about 500 μ g to about 8mg, about 500 μ g to about 5mg, about 500 μ g to about 4mg, about 500 μ g to about 3mg, about 500 μ g to about 2mg, about 500 μ g to about 1mg, about 1mg to about 68mg, about 1mg to about 34mg, about 1mg to about 17mg, about 1mg to about 8mg, about 1mg to about 6mg, about 1mg to about 5mg, about 1mg to about 4mg, about 1mg to about 3mg, about 1mg to about 2mg, about 1.5mg to about 8mg, about 1.5mg to about 6mg, about 1.5mg to about 5mg, about 1.5mg to about 4mg, about 1.5mg to about 3mg, about 1.5mg to about 2mg, about 1.3mg to about 2.6mg, or about 1.2mg to about 2.2 mg. In some embodiments, magnesium ions are provided using a magnesium salt (e.g., magnesium citrate and/or magnesium chloride).

In some embodiments, the magnesium salt administered comprises magnesium chloride and an effective amount of magnesium salt of about 0.48mg to about 600mg magnesium chloride hexahydrate (MgCl) ₂ ·6H ₂ O, MW 203.3). In some embodiments, the effective dose of magnesium chloride hexahydrate is about 0.48mg to about 300mg, about 0.5mg to about 150mg, about 0.5mg to about 75mg, about 5mg to about 150mg, about 5mg to about 75mg, about 5mg to about 50mg, about 10mg to about 600mg, about 10mg to about 300mg, about 10mg to about 150mg, about 10mg to about 75mg, about 10mg to about 50mg, about 10mg to about 30mg, or about 12mg to about 24mg. In some preferred embodiments, the effective dose of magnesium chloride hexahydrate is about 6mg, about 12mg, about 18mg, about 24mg, or about 30mg.

In some embodiments, the magnesium salt administered is magnesium citrate and the effective dose of magnesium salt is about 0.48mg to about 600mg of magnesium citrate. In some embodiments, an effective dose of magnesium citrate (e.g., dibasic magnesium citrate anhydrous, mw.214.4) is about 0.48mg to about 300mg, about 0.5mg to about 150mg, about 0.5mg to about 75mg, about 5mg to about 150mg, about 5mg to about 75mg, about 5mg to about 50mg, about 10mg to about 600mg, about 10mg to about 300mg, about 10mg to about 150mg, about 10mg to about 75mg, about 10mg to about 50mg, about 10mg to about 30mg, or about 12mg to about 24mg. In some preferred embodiments, the effective dose of magnesium citrate (e.g., dibasic anhydrous magnesium citrate, mw.214.4) is about 6mg, about 12mg, about 18mg, about 24mg, or about 30mg. In some embodiments, an effective dose of magnesium citrate is about 0.48mg to about 12mg, about 0.5mg to about 10mg, about 0.5mg to about 8mg, about 0.5mg to about 5mg, about 0.5mg to about 2.5mg, about 0.5mg to about 1mg, about 1mg to about 10mg, about 1mg to about 8mg, about 1mg to about 5mg, about 1mg to about 2mg, about 2mg to about 10mg, about 2mg to about 8mg, about 2mg to about 6mg, about 2mg to about 4mg, about 3mg to about 10mg, about 4mg to about 8mg, about 4mg to about 6mg, about 5mg to about 10mg, about 5mg to about 8mg, about 5mg to about 7mg, about 5mg to about 6mg, about 6mg to about 10mg, about 6mg to about 8mg, or about 6mg to about 7mg. If other magnesium salts are substituted for magnesium citrate, the effective amount of the magnesium salt provides an amount of magnesium ions equal to the amount provided by magnesium citrate.

It is intended and understood that each and every dose of magnesium ions described herein can be combined with each and every dose of oxytocin peptide described herein as if each and every combination were individually recited. For example, in some embodiments, the effective dose of oxytocin peptide is about 0.5 μ g to about 2000 μ g and the effective dose of magnesium ion is about 50 μ g to about 68mg of magnesium. In some embodiments, the effective dose of oxytocin peptide is about 15 μ g to about 120 μ g (e.g., about 60 μ g or about 66 μ g) and the effective dose of magnesium ions is equivalent to the amount of magnesium ions provided by about 10mg to about 30mg (e.g., about 12mg or about 24 mg) of magnesium citrate.

In some embodiments, provided is a method for treating an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering (e.g., by intranasal administration) an effective dose of oxytocin peptide and magnesium ions to a subject in need thereof, wherein the weight ratio between the dose of oxytocin peptide administered and the dose of magnesium ions administered is between about 1. In some embodiments, the method is provided for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, social and communication deficits, or anxiety, comprising administering (e.g., by intranasal administration) an effective dose of oxytocin peptide and magnesium ions to a subject in need thereof, wherein the molar ratio between the dose of oxytocin peptide administered and the dose of magnesium ions administered is between about 1 40040 to about 1. In some of these embodiments, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1). In some of such embodiments, the magnesium ions are provided by magnesium citrate and/or magnesium chloride. In some of these embodiments, social and communication deficiencies are deficiencies in communication skills and/or social interactions, lack of eye contact, and/or inability to form and/or maintain social relationships.

In one embodiment, methods are provided for treating autism spectrum disorders, disorders exhibiting one or more symptoms associated with autism spectrum disorders, social and communication deficits, or anxiety comprising intranasally administering to a subject in need thereof an oxytocin peptide in a dose of about 0.5 μ g to about 2000 μ g (e.g., about 8 μ g to about 300 μ g, about 15 μ g to about 120 μ g, or about 66 μ g) and a magnesium or magnesium ion in a dose of about 50 μ g to about 68mg, about 50 μ g to about 34mg, about 1mg to about 3mg, about 1.3mg, or about 2.6mg. In one embodiment, the method comprises intranasally administering to a subject in need thereof an effective amount of a magnesium-containing oxytocin peptide formulation or composition described herein. In one embodiment, the method comprises administering intranasally to a subject in need thereof an effective amount of an oxytocin peptide and a magnesium ion in a liquid formulation having a volume of between about 5 μ Ι _andabout 1000 μ Ι _. In one embodiment, the method comprises intranasally administering to a subject in need thereof an effective amount of a magnesium-containing oxytocin peptide formulation or composition comprising between about 0.01mg/mL and about 16mg/mL (e.g., about 0.1mg/mL and about 16 mg/mL) oxytocin and between about 1mg/mL and about 30mg/mL magnesium or magnesium ions. In one embodiment, the method comprises intranasally administering to a subject in need thereof an effective amount of a magnesium-containing oxytocin peptide formulation comprising between about 0.01mg/mL and about 16mg/mL (e.g., about 0.1mg/mL and about 16mg/mL or about 0.15mg/mL and about 1.5 mg/mL) oxytocin and between about 1% and about 25% (by weight) (e.g., about 1% to about 15% or about 10% to about 14%) magnesium citrate. In one embodiment, the method comprises intranasally administering to a subject in need thereof an effective amount of a magnesium-containing oxytocin peptide formulation comprising between about 5IU/mL and about 8000IU/mL (e.g., about 50IU/mL and about 8000IU/mL or about 75IU/mL and about 750 IU/mL) oxytocin and between about 1% and about 25% (by weight) (e.g., about 1% to about 15%, about 10% to about 14%, or about 12%) magnesium citrate. In one embodiment, the method comprises intranasally administering to a subject in need thereof an effective amount of a magnesium-containing oxytocin peptide formulation comprising between about 0.01mg/mL and about 16mg/mL (e.g., about 0.1mg/mL and about 16mg/mL or about 0.15mg/mL and about 1.5 mg/mL) oxytocin and between about 1% and about 25% (by weight) (e.g., about 1% to about 15%, about 8% to about 12%, or about 10%) magnesium chloride hexahydrate. In one embodiment, the method comprises intranasally administering to a subject in need thereof an effective amount of a magnesium-containing oxytocin peptide formulation comprising between about 5IU/mL and about 8000IU/mL (e.g., about 50IU/mL and about 8000IU/mL or about 75IU/mL and about 750 IU/mL) oxytocin and between about 1% and about 25% (by weight) (e.g., about 1% to about 15%, about 8% to about 12%, or about 10%) magnesium chloride hexahydrate.

In some embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 0.5 μ g (or 0.25 IU) to about 2000 μ g (or 1000 IU) of oxytocin peptide administered in an aqueous solution containing about 0.1% to about 2.8% (w/v) magnesium. In some embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 8 μ g (or 4 IU) to about 1000 μ g (or 500 IU) of oxytocin peptide administered in an aqueous solution containing about 0.11% to about 1.65% (w/v) magnesium. In some embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 15 μ g (or 7.5 IU) to about 120 μ g (or about 60 IU) (e.g., about 60 μ g or 30 IU) of oxytocin peptide administered in an aqueous solution containing about 1.1% to about 1.6% (e.g., about 1.2% or about 1.35%) magnesium. In one embodiment, the effective dose of oxytocin peptide and magnesium ions comprises about 60 μ g (or 30 IU) of oxytocin peptide administered in an aqueous solution containing about 1.2% or about 1.35% magnesium.

In some embodiments, the effective dose of oxytocin peptide and magnesium ion is administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _. In some embodiments, the volume administered is between about 5 μ L and about 500 μ L, between about 5 μ L and about 250 μ L, between about 5 μ L and about 100 μ L, between about 5 μ L and about 50 μ L, between about 10 μ L and about 1000 μ L, between about 10 μ L and about 500 μ L, between about 10 μ L and about 250 μ L, between about 10 μ L and about 100 μ L, between about 25 μ L and about 1000 μ L, between about 25 μ L and about 500 μ L, between about 25 μ L and about 250 μ L, between about 25 μ L and about 100 μ L, between about 50 μ L and about 1000 μ L, between about 50 μ L and about 750 μ L, between about 50 μ L and about 500 μ L, between about 50 μ L and about 450 μ L, between about 50 μ L and about 400 μ L, between about 50 μ L and about 350 μ L, between about 50 μ L and about 300 μ L, between about 50 μ L and about 250 μ L, between about 50 μ L and about 200 μ L, between about 150 μ L and about 100 μ L, between about 100 μ L, or between about 100 μ L and about 100 μ L. In some embodiments, the volume administered is about 50 μ L, about 100 μ L, about 150 μ L, about 200 μ L, about 250 μ L, about 300 μ L, about 350 μ L, about 400 μ L, about 450 μ L, or about 500 μ L. In some embodiments, an effective dose of oxytocin peptide and magnesium ion is administered intranasally in a liquid formulation contained in a nasal device as described herein.

The combinations of oxytocin peptides and magnesium ions described herein may be used to treat any social and communication deficits treatable by oxytocin, such as a deficit in communication skills and/or social interactions, lack of eye contact, and/or inability to form and/or maintain social relationships. Thus, provided are methods for treating social and communication deficits, comprising administering to a subject in need thereof an effective dose of an oxytocin peptide and a magnesium ion, wherein the social and communication deficits are a deficit in communication skills and/or social interactions, lack of eye contact, and/or an inability to form and/or maintain social relationships. In one embodiment, the method comprises administering intranasally to a subject in need thereof an effective dose of an oxytocin peptide and a magnesium ion. In some embodiments, the molar ratio between the amount of oxytocin peptide and the amount of magnesium or magnesium ions is about 1.

In one embodiment, methods are provided for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising intranasally administering to a subject in need thereof (e.g., a human or veterinary patient) an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect. In some embodiments, the oxytocin peptide is a human oxytocin consisting of Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly (SEQ. ID NO: 1). In some embodiments, an effective dose of oxytocin peptide is about 0.5 μ g (or 0.25 IU) to about 2000 μ g (or 1000 IU), preferably about 8 μ g (or 4 IU) to about 1000 μ g (or 500 IU), more preferably about 15 μ g (or 7.5 IU) to about 120 μ g (or 60 IU). In some embodiments, an effective dose of magnesium ions is from about 50 μ g to about 68mg. In some embodiments, magnesium ions are provided using a magnesium salt (e.g., magnesium chloride and/or magnesium citrate) administered in an amount to provide about 50 μ g to about 68mg of magnesium. In some embodiments, an effective dose of magnesium ions is provided by using from about 0.48mg to about 600mg of magnesium citrate. In some embodiments, an effective dose of magnesium ions is provided by using from about 0.42mg to about 540mg of magnesium chloride hexahydrate. In some embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 15 μ g (or 7.5 IU) to about 120 μ g (or 60 IU) (e.g., about 60 μ g or 30 IU) of oxytocin peptide — administered in an aqueous solution containing about 1.1% to about 1.54% (e.g., about 1.2% or about 1.35%) (w/v) magnesium. In some embodiments, the effective dose of oxytocin peptide and magnesium ion comprises about 10 μ g to about 120 μ g (e.g., about 66 μ g) of oxytocin peptide administered in an aqueous solution containing about 10% to about 14% (e.g., about 12%) (w/v) magnesium citrate.

Suit set

Provided herein are kits for performing any of the methods described herein. The kit is provided for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety. In some embodiments, the kit comprises oxytocin peptide and magnesium ion in a suitable package, wherein the oxytocin peptide and magnesium ion are in amounts to produce a synergistic or enhanced effect upon treatment of an autism spectrum disorder, a disorder exhibiting one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, and a device for craniofacial mucosal administration (e.g., intranasal administration). The kit may further comprise a protease inhibitor and/or at least one absorption enhancer. The kit may further comprise IL-6. Other kits may further include instructions for providing information to the user and/or healthcare provider for performing any of the methods described herein. The kit may further comprise reagents/means for measuring IL-6 levels in a subject; and optionally instructions for anticipating the efficacy of nasal oxytocin and magnesium ions.

Also provided are kits comprising a device for craniofacial mucosal administration (e.g., an intranasal administration device such as a nasal pump device) and a magnesium-containing oxytocin peptide formulation as described herein in a suitable package. The kit may further comprise instructions for administering the magnesium-containing oxytocin peptide formulation in a subject in need thereof.

The instructions relating to the use of the kit for carrying out the invention generally describe how the contents of the kit can be used to carry out the method of the invention. The instructions provided in the kit of the present invention are typically written instructions on a label or package insert (e.g., a sheet of paper included in the kit), but machine-readable instructions (e.g., instructions carried on a magnetic or optical storage disc) are also acceptable.

Examples

The invention may be further understood by reference to the following examples, which are provided by way of illustration and are not intended to be limiting.

Example 1: exemplary preparation of magnesium-containing oxytocin peptide formulations

Example 1A

A pharmaceutical product formulation that is hypertonic and targeted at ph4.5 consists of oxytocin USP (150 IU/mL); magnesium chloride USP (as the hexahydrate or anhydrous salt); citric acid USP (as anhydrous or monohydrate form); sodium hydroxide NF; sterile water for injection USP. The quantitative compositions are provided in table 1. The molar ratio of oxytocin to magnesium ions in the formulation is about 1. All ingredients meet the pharmacopoeia (USP/NF) requirements of the corresponding monograph.

TABLE 1

¹ The exact amount of oxytocin used is based on the oxytocin activity of the supplier assay certificate.

² The composition value of magnesium chloride represents the composition value of hexahydrate; anhydrous salts may also be used and the composition adjusted accordingly.

³ The composition value of citric acid represents the composition value of the anhydrous form; monohydrate can also be used with corresponding adjustments in composition.

Pharmaceutical products are manufactured by dissolving the ingredients in sterile water for injection, sterile filtering and rapidly filling into vials on preservative-free pumps, and are typically tested according to the FDA nasal spray guidelines (FDA nasal spray guidelines) on month 7 of 2002.

In one embodiment, a 10-L batch (batch) of magnesium-containing oxytocin formulation is prepared according to the composition provided in table 1 as follows: the formulation container was filled with water to about 60% of the volume of the desired batch. While stirring at ambient temperature, the required amounts were added in the following order: sodium chloride, citric acid and magnesium chloride hexahydrate. The material dissolves easily. No heating was required, only gentle stirring. With the addition of 1N NaOH, the pH of the solution was adjusted to 4.5. (if over-titrated, 10% HCl may be used to back-titrate to pH 4.5). The required amount of oxytocin was added and stirred until dissolved. Water was added to bring the batch to final weight/volume. Stirring until the solution is homogeneous.

Example 1B

A pharmaceutical product formulation that is isotonic and targeted at pH4.5 consists of oxytocin USP (150 IU/mL); magnesium citrate; sodium chloride USP; sodium acetate trihydrate USP; glacial acetic acid USP; and sterile water for injection USP. The quantitative compositions are provided in table 2. The molar ratio of oxytocin to magnesium ions in the formulation is about 1. A target pH of 4.5 was selected-based on optimal formulation stability at or near this pH (Hawe, et al. Pharmaceut. Res.2009, 26. All ingredients meet the pharmacopoeia (USP/NF) requirements of the corresponding monograph.

To prepare a stock oxytocin solution, lyophilized oxytocin (2 mg) was added to 1mL of water (USP), 0.9% normal saline or phosphate buffered saline in a 5mL glass container. The solution was stirred until all oxytocin was dissolved and the pH was adjusted to between 3.5 and 8.5, yielding 1mL of a 2mg/mL (about 1000 IU/mL) liquid oxytocin formulation.

For use as a clinical material, oxytocin and excipients were manufactured under current Good Manufacturing practices (Good Manufacturing Practice) and terminally sterilized (sterile filtration through 0.2 micron membrane filters) before filling into glass container vials and sealing with pump actuators. Various formulation concentrations can be produced by this example by increasing or decreasing the amount of oxytocin. Approximately 10 doses of oxytocin were obtained from this 1mL batch volume.

TABLE 2

Composition (I)	Concentration (mg/mL)
		Oxytocin USP	0.283
Citric acid magnesium salt	120
		Sodium chloride USP	4.675
Sodium acetate trihydrate USP	6.805
		Citric acid USP	pH 4.5
Sterile water for injection USP	qs

Example 2 rat social behavior model

Rats were treated intranasally with 20 μ l (10 μ l/nostril) of a solution containing saline, 10 μ g oxytocin, a combination of 12% magnesium citrate and 10 μ g oxytocin (oxytocin to magnesium ion molar ratio of about 1 1127), or 12% magnesium citrate. Eight (8) rats were used in each treatment group. Forty minutes after nasal drug administration, two animals from the same treatment group were partnered and placed in the test compartment and their social behavior (sniffing, following, climb and climb down, groom each other [ modification partners ], and alarm) was recorded for 10 minutes. The time spent on the social interaction is shown in fig. 1. The results show evidence of an enhanced effect of the combination of 12% magnesium citrate and 10 μ g oxytocin on improving social behaviour.

Example 3 rat anxiety model

Example 3A

Rats were treated intranasally with 20 μ l (10 μ l/nostril) of a solution containing saline, 10 μ g oxytocin, a combination of 12% magnesium citrate and 10 μ g oxytocin (oxytocin to magnesium ion molar ratio of about 1 1127), or 12% magnesium citrate. Eight (8) rats were used in each treatment group. Fifty minutes after nasal drug administration, animals were placed in an eight arm maze (radial arm maze) and their anxiety was assessed by the number of open arm entries made by the animals during the 5 minute time period. The number of open arm entries observed is shown in fig. 2. The results show evidence of a synergistic effect of a combination of 12% magnesium citrate and 10 μ g oxytocin on reducing anxiety.

Example 3B

Rats were treated intranasally with 20 μ l (10 μ l/nostril) of a solution containing saline, 3% magnesium citrate, 6% magnesium citrate, 16 μ g oxytocin, 10 μ g oxytocin, a combination of 3% magnesium citrate and 16 μ g oxytocin (oxytocin to magnesium ion molar ratio of about 1. Eight (8) rats were used in each treatment group. Thirty minutes after nasal drug administration, animals were exposed to 5 minutes of elevated platform stress, followed immediately by placement in the elevated plus maze for 5 minutes. Their anxiety was assessed by the number of open arm entries made during the 5 minute time period. The number of open arm entries observed is shown in fig. 3A and 3B. Animals treated with the combination of 3% magnesium citrate and 16 μ g oxytocin (oxytocin to magnesium ion molar ratio of about 1 to 176) had lower anxiety as indicated by increased open arm entry times compared to animals treated with 3% magnesium citrate alone or 16 μ g oxytocin alone. In contrast, animals treated with a combination of 6% magnesium citrate and 10 μ g oxytocin (oxytocin to magnesium ion molar ratio of about 1).

To further assess anxiety, the latency of open arm entry, the time spent in open arm, and the number of closed arm entries were determined.

These experiments were repeated with additional amounts of magnesium citrate and oxytocin, including, for example, 6% magnesium citrate alone, 20 μ g oxytocin alone, and a combination of 6% magnesium citrate and 20 μ g oxytocin (oxytocin to magnesium ion molar ratio of about 1) 281.

Example 4: single subject case study

Every morning and evening, subjects (e.g., children) diagnosed with autism spectrum disorder were intranasally administered a liquid formulation containing between 12 and 24IU of oxytocin for a period of 3 days. The subject was evaluated for social function and anxiety. After 4 days of rinsing, subjects were intranasally administered a liquid formulation containing between 3% and 12% magnesium citrate daily in the morning and evening for a period of 3 days and evaluated for social function and anxiety in subjects. After 4 days of washing, the subjects were intranasally administered a liquid formulation containing a combination of between 12 and 24IU oxytocin and between 3% and 12% magnesium citrate daily in the morning and evening for a period of 3 days and evaluated for social function and anxiety of the subjects.

Example 5: human clinical test

The effect of a 6-week course of intranasal treatment with a combination of oxytocin and magnesium twice daily was tested in male and female subjects aged 18 to 55 years diagnosed with autism spectrum disorder using a double-blind, randomized, placebo-controlled, parallel design. The primary efficacy endpoint was the change in social reciprocity score measured by Autism Diagnostic Observation table-II (the Autism Diagnostic evaluation Schedule-II) before and after the double-blind treatment period. The secondary endpoint consists of one or more of:

(1) Autism diagnosis before and at completion of the double-blind treatment period observed changes in the scores of communication and restriction and repetitive behaviors measured in table-II;

(2) Changes in Anxiety as measured by the State and Trait Anxiety scale (the State and train Anxiety Inventory) assessed before and at the completion of a double-blind treatment session;

(3) Changes in Depression measured by the epidemiological study Center Depression Scale (the Center for epidemic Studies Depression Scale) assessed before and at the completion of the double-blind treatment period;

(4) Eye fixation changes to social cues assessed before and at completion of the double-blind treatment session;

(5) Changes in facial and phonetic expressions analyzed from videos recorded every two weeks during the trial; and

(6) Changes in Clinical Global Impression and Global Assessment of function scores (Clinical Global Impression and Global Assessment of functions) assessed every two weeks during the trial.

The study inclusion criteria consisted of:

1) (ii) diagnosis of autism spectrum disorder based on DSM-V;

2) Qualitative abnormalities of reciprocal social interaction (Domain A) in the Autism Diagnostic Interview-revision (Automation Diagnostic Interview-reviewed) exceed the limit; and

3) Language IQ85 or more, full IQ 80 or more measured using Wechsler Adult intelligence Scale III (Wechsler Adult Intelligent Scale-III).

The study inclusion criteria consisted of:

1) A primary psychiatric diagnosis other than inclusion criteria 1);

2) Current instability due to comorbid psychiatric diagnostics;

3) (ii) a history of drug or drug dose variation of the psychotropic drug over a month of randomization;

4) Currently, treatment with more than two classes of psychotropic drugs;

5) Current treatment with atomoxetine (atomoxetine) or methylphenidate;

6) History of oxytocin sustained treatment;

7) History of sensitivity to oxytocin;

8) A history of over 5 minutes of seizure or traumatic brain injury loss of consciousness; and

9) History of alcoholism or substance abuse or addiction.

Example 6: effects of oxytocin and magnesium in social anxiety disorders

Subjects who met the generalized social phobia criteria of the fourth edition of the Diagnostic and Statistical Manual of Mental Disorders (Diagnostic and Statistical Manual of Mental Disorders) were randomized to treatment with nasal placebo (saline) -treatment A, oxytocin (30 IU) alone-treatment B, magnesium alone (10%) -treatment C, or oxytocin (30 IU) plus magnesium (10%) -treatment D.

After a 1-week, single-blind, placebo, run-in period, patients received double-blind, 11-week courses of treatment a, B, C, or D. Patients received treatment twice daily, approximately 12 hours apart.

Optionally, serum IL-6 levels are collected at the end of the 1-week break-in period and at the end of the 11-week course of treatment.

Measuring the number of respondents ("substantial Improvement" or "very good Improvement") based on the Clinical Global Impression Global Improvement Item(s); average change from baseline in the Liebowitz Social Anxiety Scale (the Liebowitz Social Anxiety Scale) total score. Optionally, the serum level of IL-6 is correlated with the degree of efficacy of IL-6 as a predictive biomarker for efficacy to determine effectiveness.

The effect of the treatment group was analyzed.

Exemplary embodiments

The invention is further described by the following embodiments. The features of each of the embodiments may be combined with any of the other embodiments, where appropriate and practical.

Embodiment 1 in one embodiment, there is provided a method for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein co-administration of the oxytocin peptide and magnesium ion produces a synergistic or potentiating effect.

Embodiment 2. In a further embodiment of embodiment 1, the oxytocin peptide is administered simultaneously with the magnesium ion.

Embodiment 3. In a further embodiment of embodiment 1, the oxytocin peptide is administered before or after administration of magnesium ions.

Embodiment 4. In a further embodiment of any one of embodiments 1 to 3, the oxytocin peptide is administered by craniofacial mucosal administration.

Embodiment 5. In a further embodiment of embodiment 4, the oxytocin peptide is administered by intranasal administration.

Embodiment 6. In a further embodiment of embodiment 5, the oxytocin peptide and the magnesium ion are administered by intranasal administration.

Embodiment 7. In a further embodiment of any one of embodiments 1 to 6, the effective dose of oxytocin peptide is about 0.5 μ g to about 2000 μ g.

Embodiment 8. In a further embodiment of any one of embodiments 1 to 7, the effective dose of magnesium ions is about 50 μ g to about 68mg.

Embodiment 9. In a further embodiment of any one of embodiments 1 to 8, magnesium ions are provided using magnesium chloride and/or magnesium citrate.

Embodiment 10. In a further embodiment of embodiment 1, the effective doses of the oxytocin peptide and the magnesium ion comprise about 15 μ g to about 120 μ g of the oxytocin peptide administered in an aqueous solution containing about 1.1% to about 1.6% (w/v) magnesium.

Embodiment 11. In a further embodiment of embodiment 1, the effective dose of oxytocin peptide and magnesium ion has a oxytocin to magnesium molar ratio of between about 1.

Embodiment 12 in a further embodiment of any one of embodiments 1 to 11, the method is used to treat autism spectrum disorders.

Embodiment 13 in a further embodiment of any one of embodiments 1 to 11, the methods are used to treat a disorder exhibiting one or more symptoms associated with an autism spectrum disorder.

Embodiment 14. In a further embodiment of embodiment 13, the disorder is social anxiety disorder, obsessive compulsive disorder, a social (pragmatic) communication disorder, a neurodevelopmental disorder, attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, frangle-X syndrome, rett syndrome, or Williams syndrome.

Embodiment 15. In a further embodiment of any one of embodiments 1 to 11, the method is used to treat social and communication deficits.

Embodiment 16 in a further embodiment of any one of embodiments 1 to 11, the methods are used to treat anxiety.

Embodiment 17. In a further embodiment of any one of embodiments 1 to 16, the oxytocin peptide is human oxytocin (SEQ. ID NO: 1).

Embodiment 18 in one embodiment, there is provided a method for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, comprising administering to a subject in need thereof an effective dose of oxytocin peptide and magnesium ion, wherein the effective dose of oxytocin peptide and magnesium ion are administered intranasally in a liquid formulation, and the volume of the liquid formulation administered is between about 5 μ Ι _ and about 1000 μ Ι _.

Embodiment 19. In a further embodiment of embodiment 18, the effective dose of oxytocin peptide is about 0.5 μ g to about 2000 μ g.

Embodiment 20. In a further embodiment of embodiment 18, the effective dose of magnesium ions is about 50 μ g to about 68mg.

Embodiment 21. In a further embodiment of embodiment 18, the effective doses of the oxytocin peptide and the magnesium ion include about 15 μ g to about 120 μ g of the oxytocin peptide administered in an aqueous solution containing about 1.1% to about 1.6% (w/v) magnesium.

Embodiment 22. In a further embodiment of embodiment 18, the effective dose of oxytocin peptide and magnesium ion has a molar ratio of oxytocin to magnesium between about 1.

Embodiment 23. In a further embodiment of embodiment 21 or 22, the liquid formulation is administered in a volume of between about 50 μ L and about 200 μ L.

Embodiment 24. In a further embodiment of embodiment 23, the liquid formulation is administered at 1 to 4 units, about 50 μ L/unit, using a metered nasal device.

Embodiment 25 in a further embodiment of any one of embodiments 18 to 24, the method is used for treating an autism spectrum disorder.

Embodiment 26 in a further embodiment of any one of embodiments 18 to 24, the method is used to treat a disorder exhibiting one or more symptoms associated with an autism spectrum disorder.

Embodiment 27. In a further embodiment of embodiment 26, the disorder is social anxiety disorder, obsessive compulsive disorder, a social (pragmatic) communication disorder, a neurodevelopmental disorder, attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, frangle-X syndrome, rett syndrome, or Williams syndrome.

Embodiment 28. In a further embodiment of any one of embodiments 18 to 24, the method is used to treat social and communication deficits.

Embodiment 29 in a further embodiment of any one of embodiments 18 to 24, the methods are used to treat anxiety.

Embodiment 30. In a further embodiment of any one of embodiments 18 to 29, the oxytocin peptide is human oxytocin (SEQ. ID NO: 1).

Embodiment 31. In a further embodiment of embodiment 18, the liquid formulation is contained within an intranasal administration device.

Embodiment 32. In a further embodiment of embodiment 31, the intranasal administration device is a nasal pump device.

Embodiment 33 in a further embodiment of embodiment 32, the nasal pump device comprises a container bottle attached to the pump actuator.

Embodiment 34 in a further embodiment of embodiment 33, the pump actuator is metered to deliver a specified volume of about 50 μ Ι _.

Embodiment 35 in a further embodiment of embodiment 32, the nasal pump apparatus comprises a container bottle attached to the aerosolizer.

Embodiment 36 in a further embodiment of any one of embodiments 32 to 35, the nasal pump device comprises one or more of:

(i) A filter for preventing a back flow of the liquid,

(ii) A metal-free fluid path, and

(iii) A plastic material stable to gamma radiation.

Embodiment 37. In one embodiment, a composition is provided comprising an oxytocin peptide and magnesium ions, wherein the oxytocin peptide and magnesium ions are in amounts that produce a synergistic or potentiating effect when used in the treatment of anxiety.

Embodiment 38. In a further embodiment of embodiment 37, the oxytocin peptide is human oxytocin (SEQ. ID NO: 1).

Embodiment 39. In a further embodiment of embodiment 37, the composition is a liquid formulation comprising between about 0.01mg/mL and about 16mg/mL oxytocin peptide.

Embodiment 40. In a further embodiment of embodiment 37, the composition is a liquid formulation comprising an amount of magnesium salt that provides between about 3mg/mL and about 30mg/mL of magnesium.

Embodiment 41. In a further embodiment of embodiment 37, the molar ratio of oxytocin peptide and magnesium ion is between about 1.

Embodiment 42. In a further embodiment of embodiment 41, the molar ratio is between about 1.

Embodiment 43. In a further embodiment of embodiment 41, the molar ratio is between about 1.

Embodiment 44 in a further embodiment of any one of embodiments 37 to 43, the composition further comprises a device for craniofacial mucosal administration.

Embodiment 45 in a further embodiment of embodiment 44, the oxytocin peptide and the magnesium ion are comprised in a device for craniofacial mucosal administration.

Embodiment 46. In a further embodiment of embodiment 45, the device is for intranasal administration.

Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be obvious to those skilled in the art that certain changes and modifications may be practiced without departing from the invention. Therefore, the description and examples should not be construed as limiting the scope of the invention.

All patents, patent applications, documents, and articles cited herein are hereby incorporated by reference in their entirety.

Claims (19)

1. Use of a composition comprising an effective dose of an oxytocin peptide and an effective dose of magnesium ions in the manufacture of a medicament for treating an autism spectrum disorder, a disorder showing one or more symptoms associated with an autism spectrum disorder, a social and communication deficit, or anxiety, wherein the medicament is formulated as an aqueous solution for administration by craniofacial mucosal administration, wherein the effective dose of the oxytocin peptide and the effective dose of magnesium ions have an oxytocin peptide/magnesium ion molar ratio of 1:1100 to 1:1600, and wherein said effective dose of said oxytocin peptide and said effective dose of said magnesium ion produce a synergistic effect.

2. The use of claim 1, wherein the oxytocin peptide/magnesium ion molar ratio is between about 1:1100.

3. the use of claim 1, wherein the craniofacial administration is intranasal administration.

4. The use of claim 1, wherein the effective dose of oxytocin peptide is between 0.5 μ g and 2000 μ g.

5. The use of claim 1, wherein the effective dose of magnesium ions is between 50 μ g and 68mg.

6. Use according to claim 1, wherein the magnesium ions are provided by means of magnesium chloride and/or magnesium citrate.

7. The use of claim 1, wherein the oxytocin peptide is human oxytocin of SEQ ID No. 1.

8. The use of claim 1, wherein the medicament is for the treatment of autism spectrum disorders.

9. The use of claim 1, wherein the medicament is used to treat a disorder exhibiting one or more symptoms associated with an autism spectrum disorder.

10. The use of claim 9, wherein the disorder is social anxiety disorder, obsessive compulsive disorder, social communication disorder, attention deficit hyperactivity disorder, prader-Willi syndrome, timothy syndrome, fragile-X syndrome, rett syndrome, or Williams syndrome.

11. The use of claim 9, wherein the disorder is a neurodevelopmental disorder.

12. The use of claim 1, wherein the medicament is used to treat social and communication deficits.

13. The use according to claim 1, wherein the medicament is for the treatment of anxiety.

14. The use of claim 3, wherein the medicament is contained in an intranasal administration device.

15. The use of claim 14, wherein the intranasal administration device is a nasal pump device.

16. The use of claim 15, wherein the nasal pump device comprises a container bottle attached to a pump actuator.

17. The use of claim 16, wherein the pump actuator is metered to deliver a specified volume of about 50 μ L.

18. The use of claim 17, wherein the nasal pump device comprises a container bottle attached to an aerosolizer.

19. The use of any one of claims 15 to 18, wherein the nasal pump device comprises one or more of:

(i) A filter for preventing a back flow of the liquid,

(ii) A metal-free fluid path, and

(iii) A plastic material stable to gamma radiation.