Methods and compositions for the treatment of optic neuropathy

Abstract

The present invention provides, among other things, improved methods and compositions for treating optic neuropathy comprising administering to a subject suffering from optic neuropathy an angiotensin (1-7) peptide, wherein the administration results in a reduction in the intensity, severity, duration, or frequency of at least one symptom of optic neuropathy (e.g. non-arteritic anterior ischemic neuropathy).

Description

[0001] METHODS AND COMPOSITIONS FOR THE TREATMENT OF OPTIC NEUROPATHY

[0002] BACKGROUND

[0003] Optic neuropathy occurs when the optic nerve is damaged due to blocked blood flow, inflammation or abnormalities caused by various conditions, or trauma. For example, Nonarteritic anterior ischemic optic neuropathy (NA-AION or NAION) is a form of acute anterior ischemic optic neuropathy that refers to loss or impairment of blood flow to the optic nerve supplied by the posterior ciliary artery that typically causes sudden vision loss in one eye, without any pain.

[0004] Although colloquially referred to as eye stroke, NAION is dissimilar to stroke in that there are no signs of emboli, thrombosis, or other vascular occlusive disease. Instead, the disease resembles a compartment syndrome, and consists of initial tissue vessel leakage followed by a cycle of edema, subsequent closure of small vessels within the scleral canal and anterior optic nerve, and infarction. Clinical Toxicology, 2009.

[0005] NAION is a form of anterior ischemic optic neuropathy which affects between 2 to 10 individuals out of 100,000 but comprises nearly 85% of all cases of anterior ischemic optic neuropathy. There is an estimated 6000 new cases per year in the US, and most often affects a middle-aged and elderly population over the age of 50 causing a crippling visual impairment that may remain stable or progress over weeks to months. While initially affecting one eye, in 20% of patients it can affect the second eye as well.

[0006] There is currently no cure for NAION. Eyeglasses cannot correct vision loss due to NAION. Current therapy includes corticosteroid treatment, typically a moderate to high dose of prednisone. Prolonged systemic corticosteroid use causes side-effects and remains of limited use in effecting visual improvement, so there is a need for improved therapies for NAION as well as other debilitating forms of optic neuropathy. SUMMARY OF THE INVENTION

[0007] The present invention provides, among other things, methods and compositions for treating optic neuropathy, for example, anterior ischemic optic neuropathy, e.g., nonarteritic anterior ischemic optic neuropathy (NAION) comprising administering to a subject suffering from the optic neuropathy, an angiotensin (1-7) receptor agonist, in particular an angiotensin (1-7) peptide. The present invention importantly provides therapy for optic neuropathy that currently has no other cure and causes serious impairment or loss of vision, affecting one or both eyes, often progressing bilaterally once an eye is affected.

[0008] The present invention also provides a method of prophylactically treating optic neuropathy comprising administering the angiotensin-(l-7) receptor agonist, preferably to a subject at increased risk for optic neuropathy.

[0009] In preferred embodiments, the angiotensin (1-7) peptide has an amino acid sequence of Asp1-Arg2-Val3-Tyr4-Ile5- His6-Pro7(SEQ ID NO: 1), Asp1- Arg2-Val3- Ser4-Ile5-His6-Cys7(SEQ ID NO: 2), or Ala1- Arg2-Val3-Ser4-Ile5-His6-Cys7(SEQ ID NO: 3).

[0010] In some embodiments, the angiotensin (1-7) receptor agonist is a non- peptidic angiotensin (1-7) receptor agonist. In a preferred embodiment, the non- peptidic angiotensin (1-7) receptor agonist is a compound with the following structure: or a pharmaceutically acceptable salt thereof. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] The following figures are for illustration purposes only and not for limitation.

[0012] FIG. 1 shows images from analysis of L-E rat optic nerve head edema, using spectral domain optical coherence tomography (SD-OCT). FIG. 1A shows a flat on (en face) image of the retina and optic nerve of a control eye. FIG. IB shows images of measurement of the cross-sectional retina and optic nerve head (ON) in an uninduced rat, where arrowheads indicate the ON width of a single section of 322 pm. FIG. 1C shows an en face image of post-rNAION optic nerve head edema showing swelling of the intraocular optic nerve. Individual cross sections that can be measured are shown as lines. FIG. ID shows a crosssection of optic nerve head edema post-rNAION where arrowheads indicate expansion to 552 pm. The three largest contiguous sections are used to estimate mean optic nerve head diameter.

[0013] FIG. 2 shows a graph of stratification of optomotry results showing a comparison of visual acuity, as measured with optomotry (a commerical device that measures the visual acuity of an animal by induction of optokinetic nystagmus) with degree of edema. A visual acuity of 500-600 pm was categorized as mild edema, which corresponds to the least degree of ON damage, visual acuity of 600-700 pm was categorized as moderate edema and greater than 700 pm was categorized as severe edema. Treatment with Ang- (1-7) was associated with a small improvement in visual acuity relative to control in cases of mild edema. Relative improvement in visual acuity upon Ang-(l-7) was greater relative to control in eyes with a moderate degree of edema. The greatest improvement upon Ang-(l-7) treatment relative to control was observed in eyes with the most severe amount of edema.

[0014] FIG. 3 shows graphs of visual function, using visual electrophysiology as measured by flash visual evoked potential (VEP). Animals were administered Ang-(l-7) (Img / kg / d) for 28 days beginning Id post-induction. SD-OCTs were used to evaluate degree of ONH edema and presumptive severity of ON ischemia (Mild: 500-600 pm; Moderate: 600-700 pm; Severe: >700 pm). Progressively greater retention of residual VEP function was observed in Ang-(l-7)-treated animals with a more severe ischemic lesion than in vehicle-treated animals.

[0015] FIG. 4 shows a graph of VEP in animals suffering moderate and severe edema (600-850 pm) treated with Ang-(l-7) relative to control animals. Although improvement was seen in every subgroup i.e. mild and moderate edema, the greatest improvement was seen in animals with the most severe edema.

[0016] FIG. 5 : ONH edema results 1 d post-rNAION for both rat groups prior to treatment (both sexes). Mean ONH edema for each animal was based on results for three largest contiguous diameters measured by SD-OCT. Mean ONH diameter of 16 uninduced (contralateral eyes used for comparison). Animals were eliminated, if ONH edema was <450 pm. Vehicle: N=26; Treatment: N=24. The relative spread of the edema is similar in both groups.

[0017] FIG. 6: ANG(l-7) improves visual acuity by OptoMotry after rNAION. A. Regression analysis using analysis of covariance (ANCOVA). ANCOVA was performed to assess any differences in regression lines in either slope or intercept. Regression lines for both vehicle- (n=22) and ANG(l-7)-treated (n=21) are shown. ANCOVA reveals a significant difference in the intercepts between TXA-127- and vehicle-treated rats. [F(l, 40) = 7.68, p = 0.0084], B. Estimation plot for OptoMotry for animals with moderate- severe edema. Difference between treated and vehicle yielded an overall improvement of 16% for ANG(l-7)-treated animals, compared with vehicle-treated animals with moderate-severe ONH edema, defined as edema >600pm [t(25)=2.753, p=0.0108], C. subgroup analysis of ANG(l-7) treatment effects post-rNAION. ANG(l-7) provides an increasingly robust improvement in animals with progressively more severe ONH edema. There was 3.6% improvement in the mild group (>450pm but <600pm), 13% improvement in the moderate group (>600pm to 699pm), and 18% improvement in the severe group (700pm-850pm). The vertical bars in panels B and C indicate 95% confidence intervals.

[0018] FIG. 7 : ANG(l-7) administered 1 day after rNAION improves fVEP -based visual function as measured using waveform amplitudes: Analyses. A. fVEP vs Edema regression analysis using analysis of covariance (ANCOVA). ANCOVA regression analysis reveals significant difference in the intercepts between ANG(l-7) (n=22)- and vehicle-treated (n=18) rats [F(l, 37) = 4.64, p = 0.0378], B. Estimation plot for fVEP differences for vehicle vs treated animals with moderate-severe ONH edema. There is a 20% overall improvement in the fVEP of treated animals with moderate-severe ONH edema, defined as edema >600um. C. Stratified fVEP results for mild (>450pm- <600pm), moderate (600pm-699pm) and severe (700pm-850pm) ONH edema. There was an 8.0% improvement in the mild group, 21% improvement in the moderate group, and 19% improvement in the severe group. ANG(l-7) neuroregenerative / neuroreparative effects are strongest in animals with the moderate-severe amount of ONH edema. The vertical bars in panels B and C indicate 95% confidence intervals.

[0019] FIG. 8: RGC loss in vehicle- and ANG(l-7)-treated animals. There is a similar distribution of both samples. Mean value for vehicle-treated animals was 74.29±5.39 % (n=17), whereas ANG(l-7)-treated animals yielded a mean RGC loss of 63.09±7.17 % (n=14). This difference is not significant [t(29)=l .257, p=0.219],

[0020] FIG. 9: ANG(l-7) effects on late cellular inflammation after rNAION. All tissues from animals 30d post-treatment. Only animals with similar levels of RGC loss were used for comparing vehicle- and ANG(l-7) treated responses. A. Immunostaining of rat control ON (Green: Iba-1; Red: ED1 / CD68). Ramified inactive microglia are distributed throughout the ON diameter, with little EDI activity. B. Quantification of inflammation: Comparison graphs. Left plot (left bar: naive, middle bar: Vehicle-rNAION, right bar: TXA-rNAION, y-axis: % IBA-1): Vehicle treated rNAION induced ON has ~5X the Iba- 1 expression of naive (10.7±0.7% vs 2.2±0.2% naive). Iba-1 expression in ANG(l-7) treated animals is 8.5±1.0% sem (n=5 / group). The difference between vehicle- and ANG(l-7) treatment is non-significant (Mann-Whitney two tailed U test, p=0.211). Right plot (left bar: Vehicle-rNAION, right bar: TXA-rNAION, y-axis: ED1 / CD68 (Densitometric units): ED1 / CD68 expression is shown in densitometric units; EDI expression in naive ONH is virtually 0. The pattern of ED1 / CD68 expression between vehicle 162 (- and ANG(l-7) treatment was similar to that seen with Iba-1 (N=7 / group; nonsignificant, Mann-Whitney 2 tailed U test: p=0.522). C. Immunostaining of an rNAION-induced vehicle-treated animal. RGC loss at 30d=88%. The majority of microglia / macrophages are amoeboid / active, with strong upregulation of Iba-1 activity. EDI activity is also elevated, confirming activation the majority of inflammatory cells. D. Immunostaining of an rNAION-induced ANG(l-7)-treated animal. RGC loss=84%. The overall expression of both Iba-1 and ED1 / CD68 are considerably lower than that seen in vehicle-treated rNAION induced ON. DEFINITIONS

[0021] In order for the present invention to be more readily understood, certain terms are first defined below. Additional definitions for the following terms and other terms are set forth throughout the specification.

[0022] Animal: As used herein, the term “animal” refers to any member of the animal kingdom. In some embodiments, “animal” refers to humans, at any stage of development. In some embodiments, “animal” refers to non-human animals, at any stage of development. In certain embodiments, the non-human animal is a mammal (e.g., a rodent, a mouse, a rat, a rabbit, a monkey, a dog, a cat, a sheep, cattle, a primate, and / or a pig). In some embodiments, animals include, but are not limited to, mammals, birds, reptiles, amphibians, fish, insects, and / or worms. In some embodiments, an animal may be a transgenic animal, genetically-engineered animal, and / or a clone.

[0023] Approximately or about: As used herein, the term “approximately” or “about,” as applied to one or more values of interest, refers to a value that is similar to a stated reference value. In certain embodiments, the term “approximately” or “about” refers to a range of values that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value).

[0024] Biologically active'. As used herein, the phrase “biologically active” refers to a characteristic of any agent that has activity in a biological system, and particularly in an organism. For instance, an agent that, when administered to an organism, has a biological effect on that organism, is considered to be biologically active. In particular embodiments, where a peptide is biologically active, a portion of that peptide that shares at least one biological activity of the peptide is typically referred to as a “biologically active” portion. In certain embodiments, a peptide has no intrinsic biological activity but that inhibits the effects of one or more naturally- occurring angiotensin compounds is considered to be biologically active.

[0025] Carrier or diluent. As used herein, the terms “carrier” and “diluent” refers to a pharmaceutically acceptable (e.g., safe and non-toxic for administration to a human) carrier or diluting substance useful for the preparation of a pharmaceutical formulation. Exemplary diluents include sterile water, bacteriostatic water for injection (BWFI), a pH buffered solution (e.g. phosphate-buffered saline), sterile saline solution, Ringer's solution or dextrose solution.

[0026] Complication'. As used herein, the term “complication” refers to an unfavorable evolution of a disease including the development of one or more signs, symptoms or, in some embodiments, even new pathological changes that manifest for a sustained period of time (e.g., weeks, months or years). In some embodiments, complication(s) may include a progression of a sign, symptom or other pathological change, for example, a minor memory loss growing worse over time, or a difficulty with one or more motor functions progressing to paralysis.

[0027] Dosage form: As used herein, the terms “dosage form” and “unit dosage form” refer to a physically discrete unit of a therapeutic agent for the patient to be treated. Each unit contains a predetermined quantity of active material calculated to produce the desired therapeutic effect. It will be understood, however, that the total dosage of the composition will be decided by the attending physician within the scope of sound medical judgment.

[0028] Dosing regimen: A “dosing regimen” (or “therapeutic regimen”), as that term is used herein, is a set of unit doses (typically more than one) that are administered individually to a subject, typically separated by periods of time. In some embodiments, a given therapeutic agent (e.g, an angiotensin (1-7) peptide) has a recommended dosing regimen, which may involve one or more doses. In some embodiments, a dosing regimen comprises a plurality of doses each of which are separated from one another by a time period of the same length; in some embodiments, a dosing regimen comprises a plurality of doses and at least two different time periods separating individual doses. In some embodiments, the therapeutic agent is administered continuously over a predetermined period. In some embodiments, the therapeutic agent is administered once a day (QD) or twice a day (BID).

[0029] Functional equivalent or derivative'. As used herein, the term “functional equivalent” or “functional derivative” denotes, in the context of a functional derivative of an amino acid sequence, a molecule that retains a biological activity (either function or structural) that is substantially similar to that of the original sequence. A functional derivative or equivalent may be a natural derivative or is prepared synthetically. Exemplary functional derivatives include amino acid sequences having substitutions, deletions, or additions of one or more amino acids, provided that the biological activity of the protein is conserved. The substituting amino acid desirably has chemico-physical properties which are similar to that of the substituted amino acid. Desirable similar chemico-physical properties include similarities in charge, bulkiness, hydrophobicity, hydrophilicity, and the like, such as a measurement in the same individual prior to initiation of the treatment described herein, or a measurement in a control subject (or multiple control subject) in the absence of the treatment described herein. A “control subject” is a subject afflicted with the same form of disease as the subject being treated, who is about the same age as the subject being treated.

[0030] In vitro'. As used herein, the term “ / ' / / vitro” refers to events that occur in an artificial environment, e.g., in a test tube or reaction vessel, in cell culture, etc. , rather than within a multi- cellular organism.

[0031] In vivo'. As used herein, the term “z' / z vivo” refers to events that occur within a multi- cellular organism, such as a human and a non-human animal. In the context of cell-based systems, the term may be used to refer to events that occur within a living cell (as opposed to, for example, in vitro systems).

[0032] Isolated. As used herein, the term “isolated” refers to a substance and / or entity that has been (1) separated from at least some of the components with which it was associated when initially produced (whether in nature and / or in an experimental setting), and / or (2) produced, prepared, and / or manufactured by the hand of man. Isolated substances and / or entities may be separated from at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 98%, about 99%, substantially 100%, or 100% of the other components with which they were initially associated. In some embodiments, isolated agents are more than about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, substantially 100%, or 100% pure. As used herein, a substance is “pure” if it is substantially free of other components. As used herein, the term “isolated cell” refers to a cell not contained in a multi-cellular organism.

[0033] Prevent. As used herein, the term “prevent” or “prevention”, when used in connection with the occurrence of a disease, disorder, and / or condition, refers to reducing the risk of developing the disease, disorder and / or condition. See the definition of “risk.” Polypeptide'. The term “polypeptide” as used herein refers a sequential chain of amino acids linked together via peptide bonds. The term is used to refer to an amino acid chain of any length, but one of ordinary skill in the art will understand that the term is not limited to lengthy chains and can refer to a minimal chain comprising two amino acids linked together via a peptide bond. As is known to those skilled in the art, polypeptides may be processed and / or modified.

[0034] Protein-. The term “protein” as used herein refers to one or more polypeptides that function as a discrete unit. If a single polypeptide is the discrete functioning unit and does not require permanent or temporary physical association with other polypeptides in order to form the discrete functioning unit, the terms “polypeptide” and “protein” may be used interchangeably. If the discrete functional unit is comprised of more than one polypeptide that physically associate with one another, the term “protein” refers to the multiple polypeptides that are physically coupled and function together as the discrete unit.

[0035] Risk: As will be understood from context, a “risk” of a disease, disorder, and / or condition comprises a likelihood that a particular individual will develop a disease, disorder, and / or condition e.g., optic neuropathy). In some embodiments, risk is expressed as a percentage. In some embodiments, risk is from 0,1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90 up to 100%. In some embodiments risk is expressed as a risk relative to a risk associated with a reference sample or group of reference samples. In some embodiments, a reference sample or group of reference samples have a known risk of a disease, disorder, condition and / or event (e.g., optic neuropathy). In some embodiments a reference sample or group of reference samples are from individuals comparable to a particular individual. In some embodiments, relative risk is 0,1, 2, 3, 4, 5, 6, 7, 8, 9, 10, ormore.

[0036] Alternatively or additionally, risk can mean the presence of one of more risk factors for the optic neuropathy, such as age of 40 years or older (or 50 years or older, or 60 years or older), having an ancestry associated with increased risk (such as Caucasian), hypertension, diabetes mellitus (in particular type 2), atherosclerosis (in particular atherosclerosis of branches of the ophthalmic artery), intimal thickening of the short posterior ciliary artery, sleep apnea, nocturnal hypotension, anemia, smoking, migraine, hyperlipidemia, being under a GLP-1 agonist therapy or having small optic nerve openings. Preferably, the risk factor is being under a GLP-1 agonist therapy or the risk factor is having small optic nerve openings (‘disk at risk’), more preferably having small optic nerve openings.

[0037] In this context, “a subject at increased risk for optic neuropathy” herein refers to a subject having one or more of the above risk factors, preferably being under a GLP-1 agonist therapy or the risk factor is having small optic nerve openings (‘disk at risk’), more preferably having small optic nerve openings.

[0038] Sign: As used herein, the term “sign” refers to a departure from normal body function that indicates the presence of a disease or abnormality that is noticed by a person other than the patient (as opposed to a symptom, see below).

[0039] Stability: As used herein, the term “stable” refers to the ability of the therapeutic agent to maintain its therapeutic efficacy (e.g., all or the majority of its intended biological activity and / or physiochemical integrity) over extended periods of time. The stability of a therapeutic agent, and the capability of the pharmaceutical composition to maintain stability of such therapeutic agent, may be assessed over extended periods of time (e.g., for at least 1, 3, 6, 12, 18, 24, 30, 36 months or more). In certain embodiments, pharmaceutical compositions described herein have been formulated such that they are capable of stabilizing, or alternatively slowing or preventing the degradation, of one or more therapeutic agents formulated therewith. In the context of a formulation a stable formulation is one in which the therapeutic agent therein essentially retains its physical and / or chemical integrity and biological activity upon storage and during processes (such as freeze / thaw, mechanical mixing and lyophilization).

[0040] Subject. As used herein, the term “subject” refers to a human or any nonhuman animal (e.g., mouse, rat, rabbit, dog, cat, cattle, swine, sheep, horse or primate). A human includes pre- and post-natal forms. In many embodiments, a subject is a human being. A subject can be a patient, which refers to a human presenting to a medical provider for diagnosis or treatment of a disease. The term “subject” is used herein interchangeably with “individual” or “patient.” A subject can be afflicted with or is susceptible to a disease or disorder but may or may not display symptoms of the disease or disorder.

[0041] Substantially. As used herein, the term “substantially” refers to the qualitative condition of exhibiting total or near-total extent or degree of a characteristic or property of interest. One of ordinary skill in the biological arts will understand that biological and chemical phenomena rarely, if ever, go to completion and / or proceed to completeness or achieve or avoid an absolute result. The term “substantially” is therefore used herein to capture the potential lack of completeness inherent in many biological and chemical phenomena.

[0042] Suffering from'. An individual who is “suffering from” a disease, disorder, and / or condition has been diagnosed with or displays one or more symptoms of the disease, disorder, and / or condition.

[0043] Susceptible to An individual who is “susceptible to” a disease, disorder, and / or condition has not been diagnosed with the disease, disorder, and / or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and / or condition may not exhibit symptoms of the disease, disorder, and / or condition. In some embodiments, an individual who is susceptible to a disease, disorder, condition, or event (for example, optic neuropathy) may be characterized by one or more of the following: (1) a genetic mutation associated with development of the disease, disorder, and / or condition; (2) a genetic polymorphism associated with development of the disease, disorder, and / or condition; (3) increased and / or decreased expression and / or activity of a protein associated with the disease, disorder, and / or condition; (4) habits and / or lifestyles associated with development of the disease, disorder, condition, and / or event (5) having undergone, planning to undergo, or requiring a transplant. In some embodiments, an individual who is susceptible to a disease, disorder, and / or condition will develop the disease, disorder, and / or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and / or condition will not develop the disease, disorder, and / or condition.

[0044] Symptom: As used herein, the term “symptom” refers to a departure from normal body function that indicates the presence of a disease or abnormality that is noticed by the subject or patient.

[0045] Therapeutically effective amount. As used herein, the term “therapeutically effective amount” of a therapeutic agent means an amount that is sufficient, when administered to a subject suffering from or susceptible to a disease, disorder, and / or condition, to treat, diagnose, prevent, and / or delay the onset of the symptom(s) of the disease, disorder, and / or condition. It will be appreciated by those of ordinary skill in the art that a therapeutically effective amount is typically administered via a dosing regimen comprising at least one unit dose.

[0046] Treating-. As used herein, the term “treat,” “treatment,” or “treating” refers to any method used to partially or completely alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of and / or reduce incidence of one or more symptoms or features of a particular disease, disorder, and / or condition. Treatment may be administered to a subject who does not exhibit signs of a disease and / or exhibits only early signs of the disease for the purpose of decreasing the risk of developing pathology associated with the disease.

[0047] DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS

[0048] As used in this application, the terms “about” and “approximately” are used as equivalents. Any citations to publications, patents, or patent applications herein are incorporated by reference in their entirety. Any numerals used in this application with or without about / approximately are meant to cover any normal fluctuations appreciated by one of ordinary skill in the relevant art.

[0049] Other features, objects, and advantages of the present invention are apparent in the detailed description that follows. It should be understood, however, that the detailed description, while indicating embodiments of the present invention, is given by way of illustration only, not limitation. Various changes and modifications within the scope of the invention will become apparent to those skilled in the art from the detailed description.

[0050] The present invention provides, among other things, improved methods and compositions for treating optic neuropathy comprising administering to a subject suffering from the optic neuropathy, an angiotensin (1-7) receptor agonist. In other words, the invention provides an angiotensin (1-7) receptor agonist for use in treating the optic neuropathy. In yet other words, the invention provides the use of an angiotensin (l-7)receptor agonist in the manufacture of a medicament for treating the optic neuropathy.

[0051] The angiotensin (1-7) receptor agonist, wherever mentioned herein and unless indicated otherwise, is preferably an angiotensin (1-7) peptide, in particular angiotensin (1-7), a functional variant and / or a precursor thereof.

[0052] In some embodiments, the optic neuropathy is an ischemic optic neuropathy. In some embodiments, the optic neuropathy is one or more of anterior ischemic optic neuropathy or posterior ischemic optic neuropathy, and combinations thereof. In some embodiments, the anterior ischemic optic neuropathy is arteritic anterior ischemic optic neuropathy (A-AION), or non-arteritic anterior ischemic neuropathy (NA-AION). In some embodiments, the arteritic anterior ischemic optic neuropathy is associated with autoimmune inflammation and / or giant cell arteritis. In some embodiments, the posterior ischemic optic neuropathy is arteritic posterior ischemic optic neuropathy (A-PION), non-arteritic posterior ischemic optic neuropathy (NA-PION), trauma-related posterior ischemic optic neuropathy (T-PION) or perioperative posterior ischemic optic neuropathy (P-PION). In some embodiments, the optic neuropathy is anterior ischemic optic neuropathy, e.g., nonarteritic anterior ischemic optic neuropathy (NAION).

[0053] In some aspects, provided herein is a method for treating nonarteritic anterior ischemic optic neuropathy (NAION) comprising administering to a subject suffering from nonarteritic anterior ischemic optic neuropathy (NAION), an angiotensin (1-7) receptor agonist.

[0054] Various aspects of the invention are described in detail in the following sections. The use of sections is not meant to limit the invention. Each section can apply to any aspect of the invention. In this application, the use of “or” means “and / or” unless stated otherwise.

[0055] Optic Neuropathy

[0056] The optic nerve is composed of the axons of retinal ganglion cell neurons that emerge from the retina. Nerve fibers from the optic nerve head traverse through a sieve-like structure called the lamina cribrosa into the extraocular space, and posterior to the sclera is myelinated. The optic nerve then traverses into intracranial space where right and left optic nerves cross each other, forming a chiasma, which finally leads to the visual cortex in the occipital lobe.

[0057] The optic nerve is mainly divided into anterior and posterior. A portion of the optic nerve between the retina and sclera is supplied by a network of 6-12 short posterior ciliary arteries that form a plexus known as the circle of Zinn-Haller in the sclera that surrounds the optic nerve, and give rise to feeder vessels in the anterior optic nerve.

[0058] Optic neuropathy refers to damage to the optic nerve, for example, due to blocked blood flow, inflammation, structural abnormalities or trauma. In some embodiments, the optic neuropathy is ischemic optic neuropathy, due to lack of proper blood flow to the optic nerve. Optic neuropathy is of many types, but there are two different types of ischemic optic neuropathy that are defined based on the location of damage (anterior and posterior) and two forms of anterior ischemic optic neuropathy. In some embodiments, the ischemic optic neuropathy is one or more of anterior ischemic optic neuropathy or posterior ischemic optic neuropathy, and combinations thereof.

[0059] In a preferred embodiment, the optic neuropathy is associated with white matter stroke.

[0060] The optic neuropathy may be mild, moderate or severe optic neuropathy, preferably it is moderate or severe optic neuropathy and more preferably it is severe neuropathy. Severity can be expressed as visual field loss (VF loss) measured in decibels (dB) e.g. by Humphrey Visual Field (HVF) testing. Preferably, mild means a visual field loss of 5 dB or less, moderate means a visual field loss of above 5 dB to 15 dB or less, and severe means a visual field loss of above 15 dB. Alternatively or in addition, the forms may be distinguished by one or more of:

[0061] Symptoms. Mild: Minimal visual impairment, vision slightly blurred, minor difficulties with color perception, subtle changes in contrast sensitivity, and / or visual field testing shows small, localized defects. Moderate: noticeable vision loss, moderate blurring or difficulty with color perception, clear reduction in contrast sensitivity, and / or larger areas of visual field loss (e.g., central scotoma or altitudinal field defects) affecting daily activities. Severe: significant to profound vision loss, severe blurring or even blindness in the affected eye(s), significant color vision loss, poor contrast sensitivity, and / or extensive visual field defects, often with very small preserved areas or complete loss of visual fields (e.g., a central scotoma may encompass most of the visual field).

[0062] - Fundoscopic exam. Mild: Mild pallor of the optic disc (if any) and / or subtle swelling e.g. due to optic neuritis; not necessarily noticeable in early stages of all optic neuropathies. Moderate: evident pallor or atrophy of the optic disc, moderate optic disc swelling If the cause is inflammation (e.g., optic neuritis), and / or nerve fiber layer loss (visible on optical coherence tomography (OCT)). Severe: Marked pallor or atrophy of the optic disc with evidence of extensive damage, and / or chalky white optic disc.

[0063] Visual acuity. Mild: Near normal or slightly reduced. Moderate: Moderate reduction, often to between 20 / 40 and 20 / 200. Severe: Significantly reduced, often below 20 / 200, patient may be completely blind (no light perception, or NLP (no light perception)). Others. Mild: VEP (visual evoked potential) test may show mild delays in signal transmission indicating early damage to the optic nerve. Moderate: VEP test may show moderate delays in transmission and / or OCT (optical coherence tomography) or visual field testing demonstrates optic nerve damage. Severe: VEP shows severe delays or absent responses, and / or OCT may reveal extensive thinning of the retinal nerve fiber layer (RNFL) and ganglion cell layer.

[0064] Anterior Ischemic Optic Neuropathy

[0065] Anterior Ischemic Optic Neuropathy is a sudden loss of vision due to an interruption of blood flow to the anterior or front portion of the optic nerve, also known as the optic nerve head. Blocking blood flow to the optic nerve prevents delivery of oxygen and nutrients to the nerves and causes loss of vision, proportional to the extent of damage to the optic nerve. A swollen optic disc may be observed.

[0066] Arteritic Anterior Ischemic Optic Neuropathy: In some embodiments, the anterior ischemic optic neuropathy is arteritic anterior ischemic optic neuropathy (A-AION). In some embodiments, the arteritic anterior ischemic optic neuropathy is associated with autoimmune inflammation and / or giant cell arteritis. In some embodiments, the anterior ischemic optic neuropathy is associated with autoimmune inflammation and / or giant cell arteritis. In some embodiments, autoimmune inflammation leads to giant cell arteritis. In some embodiments, the giant cell arteritis causes inflammation of arteries that in some embodiments, damages the entire optic nerve head leading to permanent vision loss, and is potentially fatal. A-AION is three times as common in women than men, and frequently affects those over the age of 55. Giant cell arteritis is associated with one or more symptoms selected from pain in the temples, jaw, scalp, neck, arms or legs, fatigue, unexplained weight loss, loss of appetite and fever. In some embodiments, the giant cell arteritis is occult giant cell arteritis, which is typically symptomless and is less common. In some embodiments, A-AION presents with loss of vision without pain which may progress to bilateral and irreversible vision loss.

[0067] Non-arteritic Anterior Ischemic Optic Neuropathy: In some embodiments, the anterior ischemic optic neuropathy is non-arteritic anterior ischemic neuropathy (NA- AION). NA-AION is the most prevalent form of anterior ischemic optic neuropathy, frequently affecting those over 50, men and women alike. Sudden painless loss of vision or blurring of vision usually upon waking up from a nap or in the morning is the main symptom of NA-AION, perhaps triggered by the body’s normal drop in pressure during sleep that interrupts blood flow to the optic nerve. NA-AION is not caused by inflammation of the arteries, but instead is caused by other factors that decrease blood supply to the optic nerve, for example, low7blood pressure, increased eyeball pressure, narrowed arteries, for exampie, due to arteriosclerosis, increased blood viscosity or decreased blood flow7to the optic nerve for any other reason. Factors that increase propensity for NA-AION include, high blood pressure, diabetes mellitus, high cholesterol, smoking, sleep apnea, heart disease, blocked arteries, anemia, sudden drop in blood pressure, sickle cell disease, vasculitis, liver disease, kidney disease, erectile dysfunction, GLP-1 agonist therapy and combinations thereof.

[0068] Posterior Ischemic Optic Neuropathy

[0069] Posterior Ischemic Optic Neuropathy is an acute optic neuropathy due to ischemia in the posterior (retrobulbar) portion of the optic nerve supplied by the pial capillary plexus. It causes rapid vision loss in one or both eyes, without pain, and is associated with a relative afferent pupillary defect (RAPD) and a normal fundus (retrobulbar optic neuropathy). In contrast to AION, it is associated with a normal optic nerve head.

[0070] Arteritic Posterior Ischemic Optic Neuropathy: In some embodiments, the posterior ischemic optic neuropathy is arteritic posterior ischemic optic neuropathy (A- PION). A-PION is not as common as NA-PION, and is often seen in older individuals, due to giant cell arteritis of the posterior ciliary arteries but can also be due to other arteries. It causes severe vision loss with poor prognosis.

[0071] Non-arteritic Posterior Ischemic Optic Neuropathy: In some embodiments, the posterior ischemic optic neuropathy is non-arteritic posterior ischemic optic neuropathy (NA-PION). NA-PION is relatively rare and usually occurs as a result of small vessel vascular disease and is associated with diabetes mellitus, hypertension, atherosclerosis, glaucoma, carotid artery dissection, carotid cavernous fistula, migraine, hemodialysis, head injury, or GLP-1 agonist therapy.

[0072] Trauma-related Posterior Ischemic Optic Neuropathy: In some embodiments, the posterior ischemic optic neuropathy is trauma-related posterior ischemic optic neuropathy (T-PION). T-PION can be caused by direct or indirect ocular or head trauma that leads to penetrating damage to the optic nerve, causing serious vision loss with poor chances of recovery. Perioperative Posterior Ischemic Optic Neuropathy: In some embodiments, the posterior ischemic optic neuropathy is perioperative posterior ischemic optic neuropathy (P-PION). Ischemia occurs in pial plexus, extending to the optic nerve involving centripetal blood supply which may be affected during surgical interventions in close proximity to the optic chiasm. Patients may be younger and have bilateral vision loss in 70% of cases. Some risk factors include prolonged intraoperative arterial hypotension, postoperative anemia, increased intraocular pressure, facial swelling, and combinations thereof. Spinal surgery, blepharoplasty, sinus surgery and orbital decompression for thyroid eye disease are some examples of surgeries associated with perioperative posterior ischemic optic neuropathy.

[0073] Symptoms

[0074] In some embodiments, the one or more symptoms of optic neuropathy is selected from vision loss or impairment characterized by blurring, blind spots, shadows obscuring upper or lower visual fields, impaired visual contrast, light sensititvity, decreased color vision, increased intraocular pressure, and combinations thereof. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by blurring. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by blind spots. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by shadows obscuring upper or lower visual fields. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by impaired visual contrast. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by light sensitivity. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by decreased color vision. In some embodiments, a symptom of the optic neuropathy is vision loss or impairment characterized by increased intraocular pressure.

[0075] In some embodiments, the vision loss or impairment is bilateral. In some embodiments, the vision loss or impairment is unilateral.

[0076] In some embodiments, the optic neuropathy is associated with one or more of high blood pressure, anemia, diabetes mellitus, high cholesterol, cardiac disease, arteriosclerosis, vasculitis, liver disease, kidney disease, erectile dysfunction, GLP-1 agonist therapy and combinations thereof. In some embodiments, the optic neuropathy is associated with high blood pressure. In some embodiments, the optic neuropathy is associated with anemia. In some embodiments, the optic neuropathy is associated with diabetes mellitus. In some embodiments, the optic neuropathy is associated with high cholesterol. In some embodiments, the optic neuropathy is associated with cardiac disease. In some embodiments, the optic neuropathy is associated with arteriosclerosis. In some embodiments, the optic neuropathy is associated with vasculitis. In some embodiments, the optic neuropathy is associated with liver disease. In some embodiments, the optic neuropathy is associated with kidney disease. In some embodiments, the optic neuropathy is associated with erectile dysfunction. In some embodiments, optic neuropathy is associated with one or more of high blood pressure, anemia, diabetes mellitus, high cholesterol, cardiac disease, arteriosclerosis, vasculitis, erectile dysfunction, GLP-1 agonist therapy, in any number of combinations and in any order.

[0077] Administration

[0078] In some embodiments, the angiotensin-(l-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy is reduced in intensity, severity, duration, or frequency or has delayed onset relative to a control. In some embodiments, the angiotensin-(l-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy is reduced in intensity relative to a control. In some embodiments, the angiotensin-(l-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy is reduced in severity relative to a control. In some embodiments, the angiotensin-(l-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy is reduced in duration relative to a control. In some embodiments, the angiotensin-(l-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy is reduced in frequency relative to a control. In some embodiments, the angiotensin-(l-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy has delayed onset relative to a control. In some embodiments, the angiotensin (1-7) receptor agonist is administered once per day, once per week, three times per month, twice per month, or once per month.

[0079] In some embodiments, the administration is parenteral, rectal, intranasal, oral, or a combination thereof. In some embodiments, the administration is parenteral. In some embodiments, the administration is rectal. In some embodiments, the administration is intranasal. In some embodiments, the administration is rectal. In some embodiments, the administration is oral. In some embodiments, the administration is a combination of one or more of parenteral, rectal, intranasal, or oral.

[0080] In some embodiments, the parenteral administration is intravenous, subcutaneous, inhalation, intradermal, transdermal, and / or transmucosal administration. In some embodiments, the parenteral administration is intravenous. In some embodiments, the parenteral administration is subcutaneous. In some embodiments, the parenteral administration is inhalation. In some embodiments, the parenteral administration is intradermal. In some embodiments, the parenteral administration is transdermal. In some embodiments, the parenteral administration is transmucosal administration.

[0081] Angiotensin (1-7) peptides

[0082] In some aspects, provided herein is a method for treating optic neuropathy comprising administering to a subject suffering from optic neuropathy, an angiotensin (1-7) peptide.

[0083] As used herein, the term “angiotensin (1-7) peptide” refers to both “naturally- occurring Angiotensin (1-7)” (also referred to as “Angiotensin (1-7)” or “Ang-(l-7)” herein) and any functional equivalent, analogue or derivative of naturally-occurring Angiotensin (1-7), preferably a functional variant and / or a precursor of naturally- occurring Angiotensin (1-7). As used herein, “peptide” and “polypeptide” are interchangeable terms and refer to two or more amino acids bound together by a peptide bond. As used herein, the terms “peptide” and “polypeptide” include both linear and cyclic peptide. The terms “angiotensin-(l-7)”, “Angiotensin-(l-7)”, and “Ang-(l-7)” are used interchangeably. Naturally -occurring Angiotensin ( 1-7)

[0084] Naturally-occurring Angiotensin (1-7) (also referred to as Ang-(l-7)) is a seven amino acid peptide: Asp1-Arg2-Val3-Tyr4-Ile5-His6-Pro7(SEQ ID NO: 1). It is part of the renin-angiotensin system and is converted from a precursor, also known as Angiotensinogen, which is an a-2-globulin that is produced constitutively and released into the circulation mainly by the liver. Angiotensinogen is a member of the serpin family and also known as renin substrate. Human angiotensinogen is 452 amino acids long, but other species have angiotensinogen of varying sizes. Typically, the first 12 amino acids are the most important for angiotensin activity: Asp'-Arg2- Val3-Tyr4-Ile5-His6-Pro7-Phe8-His9-Leu10-Val11-Ile12(SEQ ID NO: 4), in particular the first 10 amino acids thereof.

[0085] Different types of angiotensin may be formed by the action of various enzymes. For example, Angiotensin (1-7) is generated by action of Angiotensin-converting enzyme 2 (ACE 2).

[0086] Angiotensin (1-7) is an endogenous ligand for Mas receptor (or G Protein- Coupled Mas Receptor, Uniprot identifier for human Mas receptor: P04201), for Mas- related G-protein coupled receptor member D (MrgD receptor, Uniprot identifier for human MrgD receptor: Q8TDS7), and, at high concentrations (e.g. 10'5M or higher) for the Angiotensin II receptor type 2 (AT2 receptor, Uniprot identifier for human AT2 receptor: P50052). Mas receptors are G-protein coupled receptor containing seven transmembrane spanning regions. As used herein, the term “angiotensin-(l-7) receptor” encompasses the above receptors, in particular the G Protein-Coupled Mas Receptors.

[0087] As used herein, the term “naturally-occurring Angiotensin (1-7)” includes any Angiotensin (1-7) peptide purified from natural sources and any recombinantly produced or chemically synthesized peptides that have an amino acid sequence identical to that of the naturally- occurring Angiotensin (1-7).

[0088] Functional equivalents, analogs or derivatives of Ang-(l-7)

[0089] In some embodiments, an angiotensin (1-7) peptide suitable for the present invention is a functional equivalent of naturally-occurring Ang-(l-7). As used herein, a functional equivalent of naturally-occurring Ang-(l-7) refers to any peptide that shares amino acid sequence identity to the naturally-occurring Ang-(l-7) and retain substantially the same or similar activity as the naturally-occurring Ang-(l-7). For example, in some embodiments, a functional equivalent of naturally-occurring Ang- (1-7) described herein has pro-angiogenic activity as determined using methods described herein or known in the art, or an activity such as nitric oxide release, vasodilation, improved endothelial function, antidiuresis, or one of the other properties discussed herein, that positively impacts angiogenesis. In some embodiments, a functional equivalent of naturally-occurring Ang-(l-7) described herein can bind to or activate an angiotensin-(l-7) receptor (e.g., the G protein-coupled Mas receptor) as determined using various assays described herein or known in the art. In some embodiments, a functional equivalent of Ang-(l-7) is also referred to as an angiotensin (1-7) analogue or derivative, or functional derivative. In some embodiments, a functional equivalent of Ang-(l-7) is a non-cyclic peptide.

[0090] Typically, a functional equivalent of angiotensin (1-7) shares amino acid sequence similarity to the naturally-occurring Ang-(l-7). In some embodiments, a functional equivalent of Ang-(l-7) according to the invention contains a sequence that includes at least 3 (e.g., at least 4, at least 5, at least 6, at least 7) amino acids from the seven amino acids that appear in the naturally- occurring Ang-(l-7), wherein the at least 3 (e.g., at least 4, at least 5, at least 6, or at least 7) amino acids maintain their relative positions and / or spacing as they appear in the naturally-occurring Ang- (1-7).

[0091] In some embodiments, a functional equivalent of Ang-(l-7) may encompass any peptide that contains a sequence at least 50% (e.g., at least 60%, 70%, 80%, or 90%) identical to the amino acid sequence of naturally-occurring Ang-(l-7). Percentage of amino acid sequence identity can be determined by alignment of amino acid sequences. Alignment of amino acid sequences can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. Preferably, the WU-BLAST-2 software is used to determine amino acid sequence identity (Altschul etal. , Methods in Enzymology 266, 460-480 (1996); http: / / blast.wustl / edu / blast / README.html). WU-BLAST-2 uses several search parameters, most of which are set to the default values. The adjustable parameters are set with the following values: overlap span=l, overlap fraction=0.125, word threshold (T)= 11. HSP score (S) andHSP S2 parameters are dynamic values and are established by the program itself, depending upon the composition of the particular sequence, however, the minimum values may be adjusted and are set as indicated above.

[0092] In some embodiments, a functional equivalent, analogue or derivative of Ang- (1-7) is a fragment of the naturally-occurring Ang-(l-7). In some embodiments, a functional equivalent, analogue or derivative of Ang-(l-7) contains amino acid substitutions, deletions and / or insertions in the naturally-occurring Ang-(l-7). Ang- (1-7) functional equivalents, analogues or derivatives can be made by altering the amino acid sequences by substitutions, additions, and / or deletions. For example, one or more amino acid residues within the sequence of the naturally-occurring Ang-(l-7) (SEQ ID NO: 1) can be substituted by another amino acid of a similar polarity, which acts as a functional equivalent, resulting in a silent alteration. Substitution for an amino acid within the sequence may be selected from other members of the class to which the amino acid belongs (conservative substitution). For example, the positively charged (basic) amino acids include arginine, lysine, and histidine. The nonpolar (hydrophobic) amino acids include leucine, isoleucine, alanine, phenylalanine, valine, proline, tryptophan, and methionine. The uncharged polar amino acids include serine, threonine, cysteine, tyrosine, asparagine, and glutamine. The negatively charged (acid) amino acids include glutamic acid and aspartic acid. The amino acid glycine may be included in either the nonpolar amino acid family or the uncharged (neutral) polar amino acid family. Substitutions made within a family of amino acids are generally understood to be conservative substitutions. For example, the amino acid sequence of a peptide inhibitor can be modified or substituted.

[0093] Examples of Ang-(l-7) functional equivalents, analogues and derivatives are described in the section entitled “Exemplary Angiotensin(l-7) Peptides” below.

[0094] An angiotensin-(l-7) peptide can be of any length. In some embodiments, an angiotensin-(l-7) peptide according to the present invention can contain, for example, from 4-25 amino acids (e.g., 4-20, 4-15, 4-14, 4-13, 4-12, 4-11, 4-10, 4-9, 4-8, 4-7 amino acids). In some embodiments, the linear peptide contains 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acids.

[0095] In some embodiments, an angiotensin-(l-7) peptide contains one or more modifications to increase protease resistance, serum stability and / or bioavailability. In some embodiments, suitable modifications are selected from pegylation, acetylation, glycosylation, biotinylation, substitution with D-amino acid and / or unnatural amino acid, and / or cyclization of the peptide. As used herein, the term “amino acid,” in its broadest sense, refers to any compound and / or substance that can be incorporated into a polypeptide chain. In certain embodiments, an amino acid has the general structure H2N-C(H)(R)-C00H. In certain embodiments, an amino acid is a naturally-occurring amino acid. In certain embodiments, an amino acid is a synthetic or un-natural amino acid (e.g., a,a- disubstituted amino acids, N-alkyl amino acids); in some embodiments, an amino acid is a d-amino acid; in certain embodiments, an amino acid is an 1-amino acid. “Standard amino acid” refers to any of the twenty standard amino acids commonly found in naturally occurring peptides including both 1- and d- amino acids which are both incorporated in peptides in nature. “Nonstandard” or “unconventional amino acid” refers to any amino acid, other than the standard amino acids, regardless of whether it is prepared synthetically or obtained from a natural source. As used herein, “synthetic or un-natural amino acid” encompasses chemically modified amino acids, including but not limited to salts, amino acid derivatives (such as amides), and / or substitutions. Amino acids, including carboxy- and / or amino- terminal amino acids in peptides, can be modified by methylation, amidation, acetylation, and / or substitution with other chemical groups that can change the peptide’ s circulating halflife without adversely affecting its activity. Examples of unconventional or un-natural amino acids include, but are not limited to, citrulline, ornithine, norleucine, norvaline, 4-(£)-butenyl-4(A)-methyl-N- methylthreonine (MeBmt), N-m ethyl -leucine (MeLeu), aminoisobutyric acid, statine, andN-methyl- alanine (MeAla). Amino acids may participate in a disulfide bond. The term “amino acid” is used interchangeably with “amino acid residue,” and may refer to a free amino acid and / or to an amino acid residue of a peptide. It will be apparent from the context in which the term is used whether it refers to a free amino acid or a residue of a peptide.

[0096] In some embodiments, angiotensin-(l-7) peptides contain one or more L- amino acids, D-amino acids, and / or un-natural amino acids. In addition to peptides containing only naturally occurring amino acids, peptidomimetics or peptide analogs are also encompassed by the present invention. Peptide analogs are commonly used in the pharmaceutical industry as non-peptide drugs with properties analogous to those of the template peptide. The non-peptide compounds are termed “peptide mimetics” or peptidomimetics (Fauchere et al., Infect. Immun. 54:283-287 (1986); Evans et al., J. Med. Chem. 30: 1229-1239 (1987)). Peptide mimetics that are structurally related to therapeutically useful peptides and may be used to produce an equivalent or enhanced therapeutic or prophylactic effect. Generally, peptidomimetics are structurally similar to the paradigm polypeptide (i.e., a polypeptide that has a biological or pharmacological activity) such as naturally- occurring receptor- binding polypeptides, but have one or more peptide linkages optionally replaced by linkages such as -CH2NH-, -CH2S-, -CH2-CH2-, -CH-CH- (cis and trans), -CH2SO-, -CH(0H)CH2-, -COCH2- etc., by methods well known in the art (Spatola, Peptide Backbone Modifications, Vega Data, 1(3):267 (1983); Spatola et al. Life Sci. 38: 1243-1249 (1986); Hudson et al. hit. J. Pept. Res. 14: 177- 185 (1979); and Weinstein. B., 1983, Chemistry and Biochemistry, of Amino Acids, Peptides and Proteins, Weinstein eds, Marcel Dekker, New-York,). Such peptide mimetics may have significant advantages over naturally-occurring polypeptides including more economical production, greater chemical stability, enhanced pharmacological properties (e.g., half-life, absorption, potency, efficiency, etc.), reduced antigenicity and others.

[0097] Ang-(l-7) peptides also include other types of peptide derivatives containing additional chemical moieties not normally part of the peptide, provided that the derivative retains the desired functional activity of the peptide. Examples of such derivatives include (1) N-acyl derivatives of the amino terminal or of another free amino group, wherein the acyl group may be an alkanoyl group (e.g., acetyl, hexanoyl, octanoyl) an aroyl group (e.g., benzoyl) or a blocking group such as F-moc (fluorenylmethyl-O-CO-); (2) esters of the carboxy terminal or of another free carboxy or hydroxyl group; (3) amide of the carboxy-terminal or of another free carboxyl group produced by reaction with ammonia or with a suitable amine; (4) phosphorylated derivatives; (5) derivatives conjugated to an antibody or other biological ligand and other types of derivatives; and (6) derivatives conjugated to a polyethylene glycol (PEG) chain.

[0098] Ang-(l-7) peptides may be obtained by any method of peptide synthesis known to those skilled in the art, including synthetic (e.g., exclusive solid phase synthesis, partial solid phase synthesis, fragment condensation, classical solution synthesis, native-chemical ligation) and recombinant techniques. For example, the peptides or peptides derivatives can be obtained by solid phase peptide synthesis, which in brief, consist of coupling the carboxyl group of the C-terminal amino acid to a resin (e.g., benzhydrylamine resin, chloromethylated resin, hydroxymethyl resin) and successively adding N-alpha protected amino acids. The protecting groups may be any such groups known in the art. Before each new amino acid is added to the growing chain, the protecting group of the previous amino acid added to the chain is removed. Such solid phase synthesis has been disclosed, for example, by Merrifield, J. Am. Chem. Soc. 85: 2149 (1964); Vale et al., Science 213: 1394-1397 (1981), in U.S. Patent Numbers 4, 305, 872 and 4,316, 891, Bodonsky et al. Chem. Ind. (London), 38: 1597 (1966); and Pietta and Marshall, Chem. Comm. 650 (1970) by techniques reviewed in Lubell et al. “Peptides” Science of Synthesis 21.11, Chemistry of Amides. Thieme, Stuttgart, 713-809 (2005). The coupling of amino acids to appropriate resins is also well known in the art and has been disclosed in U.S. Patent Number 4,244,946. (Reviewed in Houver-Weyl, Methods of Organic Chemistry. Vol E22a. Synthesis of Peptides and Peptidomimetics, Murray Goodman, Editor-in-Chief, Thieme. Stuttgart. New York 2002).

[0099] Unless defined otherwise, the scientific and technological terms and nomenclature used herein have the same meaning as commonly understood by a person of ordinary skill to which this invention pertains. Generally, the procedures of cell cultures, infection, molecular biology methods and the like are common methods used in the art. Such standard techniques can be found in reference manuals such as, for example, Ausubel et al., Current Protocols in Molecular Biology, Wiley Interscience, New York, 2001; and Sambrooket al., Molecular Cloning: A Laboratory Manual, 3rdedition, Cold Spring Harbor Laboratory Press, N.Y., 2001.

[0100] During any process of the preparation of an Ang-(l-7) peptide, it may be desirable to protect sensitive reactive groups on any of the molecule concerned. This may be achieved by means of conventional protecting groups such as those described in Protective Groups In Organic Synthesis by T.W. Greene & P.G.M. Wuts, 1991, John Wiley and Sons, New-York; and Peptides: chemistry and Biology by Sewald and Jakubke, 2002, Wiley-VCH, Wheinheim p.142. For example, alpha amino protecting groups include acyl type protecting groups (e.g., trifluoroacetyl, formyl, acetyl), aliphatic urethane protecting groups (e.g., t-butyloxycarbonyl (BOC), cyclohexyl oxy carbonyl), aromatic urethane type protecting groups (e.g., fluorenyl-9- methoxy-carbonyl (Fmoc), benzyl oxy carbonyl (Cbz), Cbz derivatives) and alkyl type protecting groups (e.g., triphenyl methyl, benzyl). The amino acids side chain protecting groups include benzyl (for Thr and Ser), Cbz (Tyr, Thr, Ser, Arg, Lys), methyl ethyl, cyclohexyl (Asp, His), Boc (Arg, His, Cys) etc. The protecting groups may be removed at a convenient subsequent stage using methods known in the art. Further, Ang-(l-7) peptides may be synthesized according to the FMOC protocol in an organic phase with protective groups. Desirably, the peptides are purified with a yield of 70% with high-pressure liquid chromatography (HPLC) on a C18 chromatography column and eluted with an acetonitrile gradient of 10-60%. The molecular weight of a peptide can be verified by mass spectrometry (reviewed in Fields, G.B. “Solid-Phase Peptide Synthesis” Methods in Enzymology. Vol. 289, Academic Press, 1997).

[0101] Alternatively, Ang-(l-7) peptides may be prepared in recombinant systems using, for example, polynucleotide sequences encoding the polypeptides. It is understood that a polypeptide may contain more than one of the above-described modifications within the same polypeptide.

[0102] While peptides may be effective in eliciting a biological activity in vitro their effectiveness in vivo might be reduced by the presence of proteases. Serum proteases have specific substrate requirements. The substrate must have both L-amino acids and peptide bonds for cleavage. Furthermore, exopeptidases, which represent the most prominent component of the protease activity in serum, usually act on the first peptide bond of the peptide and require a free N- terminus (Powell et al., Pharm. Res. 10: 1268-1273 (1993)). In light of this, it is often advantageous to use modified versions of peptides. The modified peptides retain the structural characteristics of the original L-amino acid peptides that confer the desired biological activity of Ang-(1- 7) but are advantageously not readily susceptible to cleavage by protease and / or exopeptidases.

[0103] Systematic substitution of one or more amino acids of a consensus sequence with D-amino acid of the same type (e.g., D-lysine in place of L-lysine) may be used to generate more stable peptides. Thus, a peptide derivative or peptidomimetic of the present invention may be all L, all D or mixed D, L peptide, in either forward or reverse order. The presence of an N-terminal or C-terminal D-amino acid increases the in vivo stability of a peptide since peptidases cannot utilize a D-amino acid as a substrate (Powell et al., Pharm. Res. 10: 1268-1273 (1993)). Reverse-D peptides are peptides containing D-amino acids, arranged in a reverse sequence relative to a peptide containing L-amino acids. Thus, the C-terminal residue of an L-amino acid peptide becomes N-terminal for the D-amino acid peptide, and so forth. Reverse D-peptides retain the same secondary conformation and therefore similar activity, as the L-amino acid peptides, but are more resistant to enzymatic degradation in vitro and in vivo, and thus can have greater therapeutic efficacy than the original peptide (Brady and Dodson, Nature 368:692-693 (1994); Jameson et al., Nature 368:744-746 (1994)). Similarly, a reverse-L peptide may be generated using standard methods where the C-terminus of the parent peptide becomes takes the place of the N-terminus of the reverse-L peptide. It is contemplated that reverse L-peptides of L-amino acid peptides that do not have significant secondary structure (e.g., short peptides) retain the same spacing and conformation of the side chains of the L-amino acid peptide and therefore often have the similar activity as the original L- amino acid peptide. Moreover, a reverse peptide may contain a combination of L- and D-amino acids. The spacing between amino acids and the conformation of the side chains may be retained resulting in similar activity as the original L-amino acid peptide.

[0104] Another effective approach to confer resistance to peptidases acting on the N- terminal or C-terminal residues of a peptide is to add chemical groups at the peptide termini, such that the modified peptide is no longer a substrate for the peptidase. One such chemical modification is glycosylation of the peptides at either or both termini. Certain chemical modifications, in particular N-terminal glycosylation, have been shown to increase the stability of peptides in human serum (Powell et al., Pharm. Res. 10: 1268-1273 (1993)). Other chemical modifications which enhance serum stability include, but are not limited to, the addition of an N- terminal alkyl group, consisting of a lower alkyl of from one to twenty carbons, such as an acetyl group, and / or the addition of a C-terminal amide or substituted amide group. In particular, the present invention includes modified peptides consisting of peptides bearing an N-terminal acetyl group and / or a C-terminal amide group.

[0105] Substitution of non-naturally-occurring amino acids for natural amino acids in a subsequence of the peptides can also confer resistance to proteolysis. Such a substitution can, for instance, confer resistance to proteolysis by exopeptidases acting on the N-terminus without affecting biological activity. Examples of non-naturally- occurring amino acids include alpha, alpha - disubstituted amino acids, N-alkyl amino acids, homo amino acids, beta homo-amino acids, and alpha-methyl amino acids. Amino acids analogs useful in the present invention may include, but are not limited to, D-alanine, norvaline, norleucine, 4-aminobutyric acid, ornithine, hydroxyproline, sarcosine, citrulline, cysteic acid, cyclohexylalanine, 2-aminoisobutyric acid, 6- aminohexanoic acid, t- butylglycine, phenylglycine, o-phosphoserine, N-acetyl serine, N-formylmethionine, 3-methylhistidine and other unconventional amino acids. Furthermore, the synthesis of peptides with non-naturally-occurring amino acids is routine in the art.

[0106] In addition, constrained peptides comprising a consensus sequence or a substantially identical consensus sequence variation may be generated by methods well known in the art (Rizo and Gierasch, Ann. Rev. Biochem. 61:387-418 (1992)). For example, constrained peptides may be generated by adding cysteine residues capable of forming disulfide bridges and, thereby, resulting in a cyclic peptide. Cyclic peptides can be constructed to have no free N- or C- termini. Accordingly, they are not susceptible to proteolysis by exopeptidases, although they may be susceptible to endopeptidases, which do not cleave at peptide termini. The amino acid sequences of the peptides with N-terminal or C-terminal D-amino acids and of the cyclic peptides are usually identical to the sequences of the peptides to which they correspond, except for the presence of N- terminal or C-terminal D-amino acid residue, or their circular structure, respectively.

[0107] Cyclic Peptides

[0108] In some embodiments, a functional equivalent, analogue or derivative of naturally- occurring Ang-(l-7) is a cyclic peptide. As used herein, a cyclic peptide has an intramolecular covalent bond between two non-adjacent residues. The intramolecular bond may be a backbone to backbone, side-chain to backbone or sidechain to side-chain bond (i.e., terminal functional groups of a linear peptide and / or side-chain functional groups of a terminal or interior residue may be linked to achieve cyclization). Typical intramolecular bonds include disulfide, amide and thioether bonds. A variety of means for cyclizing polypeptides are well known in the art, as are many other modifications that can be made to such peptides. For a general discussion, see International Patent Publication Nos. WO 01 / 53331 and WO 98 / 02452, the contents of which are incorporated herein by reference. Such cyclic bonds and other modifications can also be applied to the cyclic peptides and derivative compounds of this invention.

[0109] Cyclic peptides as described herein may comprise residues of L-amino acids, D- amino acids, or any combination thereof. Amino acids may be from natural or nonnatural sources, provided that at least one amino group and at least one carboxyl group are present in the molecule; a- and P-amino acids are generally preferred. Cyclic peptides may also contain one or more rare amino acids (such as 4-hydroxyproline or hydroxylysine), organic acids or amides and / or derivatives of common amino acids, such as amino acids having the C-terminal carboxylate esterified (e.g., benzyl, methyl or ethyl ester) or amidated and / or having modifications of the N-terminal amino group (e.g., acetylation or alkoxy carbonylation), with or without any of a wide variety of side-chain modifications and / or substitutions (e.g., methylation, benzylation, t- butylation, tosylation, alkoxy carbonylation, and the like). Suitable derivatives include amino acids having an N-acetyl group (such that the amino group that represents the N-terminus of the linear peptide prior to cyclization is acetylated) and / or a C-terminal amide group (i.e., the carboxy terminus of the linear peptide prior to cyclization is amidated). Residues other than common amino acids that may be present with a cyclic peptide include, but are not limited to, penicillamine, P,P-tetramethylene cysteine, P,P- pentamethylene cysteine, P -mercaptopropionic acid, P,P- pentamethylene-P-mercaptopropionic acid, 2-mercaptobenzene, 2-mercaptoaniline, 2-mercaptoproline, ornithine, di aminobutyric acid, a- aminoadipic acid, m- aminomethylbenzoic acid and a,P-diaminopropionicacid.

[0110] Following synthesis of a linear peptide, with or without N-acetylation and / or C- amidation, cyclization may be achieved by any of a variety of techniques well known in the art. Within one embodiment, a bond may be generated between reactive amino acid side chains. For example, a disulfide bridge may be formed from a linear peptide comprising two thiol-containing residues by oxidizing the peptide using any of a variety of methods. Within one such method, air oxidation of thiols can generate disulfide linkages over a period of several days using either basic or neutral aqueous media. The peptide is used in high dilution to minimize aggregation and intermolecular side reactions. Alternatively, strong oxidizing agents such as I2 and KsFe(CN)6 can be used to form disulfide linkages. Those of ordinary skill in the art will recognize that care must be taken not to oxidize the sensitive side chains of Met, Tyr, Trp or His. Within further embodiments, cyclization may be achieved by amide bond formation. For example, a peptide bond may be formed between terminal functional groups (i.e., the amino and carboxy termini of a linear peptide prior to cyclization). Within another such embodiment, the linear peptide comprises a D- amino acid.

[0111] Alternatively, cyclization may be accomplished by linking one terminus and a residue side chain or using two side chains, with or without an N-terminal acetyl group and / or a C-terminal amide. Residues capable of forming a lactam bond include lysine, ornithine (Om), a- amino adipic acid, m-aminomethylbenzoic acid, a,P-diaminopropionic acid, glutamate or aspartate. Methods for forming amide bonds are generally well known in the art. Within one such method, carbodiimide-mediated lactam formation can be accomplished by reaction of the carboxylic acid with DCC, DIC, ED AC or DCCI, resulting in the formation of an O-acylurea that can be reacted immediately with the free amino group to complete the cyclization.

[0112] Alternatively, cyclization can be performed using the azide method, in which a reactive azide intermediate is generated from an alkyl ester via a hydrazide.

[0113] Alternatively, cyclization can be accomplished using activated esters. The presence of electron withdrawing substituents on the alkoxy carbon of esters increases their susceptibility to aminolysis. The high reactivity of esters of p-nitrophenol, N- hydroxy compounds and polyhalogenated phenols has made these "active esters" useful in the synthesis of amide bonds. Within a further embodiment, a thioether linkage may be formed between the side chain of a thiol -containing residue and an appropriately derivatized a-amino acid. By way of example, a lysine side chain can be coupled to bromoacetic acid through the carbodiimide coupling method (DCC, ED AC) and then reacted with the side chain of any of the thiol containing residues mentioned above to form a thioether linkage. In order to form dithioethers, any two thiol containing side-chains can be reacted with dibromoethane and diisopropylamine inDMF.

[0114] Exemplary Angiotensin-( 1-7) Peptides

[0115] In certain aspects, the invention provides non-cyclic (e.g., linear) angiotensin- (1-7) peptides. As discussed above, the structure of naturally-occurring Ang-(l-7) is asfollows: Asp1-Arg2-Val3-Tyr4-Ile5-His6-Pro7(SEQ ID NO: 1)

[0116] The peptides and peptide analogs of the invention can be generally represented by the following sequence: Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7, or a pharmaceutically acceptable salt thereof. Xaa1is any amino acid or a dicarboxylic acid. In certain embodiments, Xaa1is Asp, Glu, Asn, Acpc (1 -aminocyclopentane carboxylic acid), Ala, MeiGly (N,N-dimethylglycine), Pro, Bet (betaine, 1-carboxy- N,N,N-trimethylmethanaminium hydroxide), Glu, Gly, Asp, Sar (sarcosine) or Sue (succinic acid). In certain such embodiments, Xaa1is a negatively-charged amino acid, such as Asp or Glu, typically Asp. Xaa2is Arg, Lys, Ala, Cit (citrulline), Om (ornithine), acetylated Ser, Sar, D-Arg and D- Lys. In certain embodiments, Xaa2is a positively-charged amino acid such as Arg or Lys, typically Arg. Xaa3is Vai, Ala, Leu, Nle (norleucine), He, Gly, Lys, Pro, HydroxyPro (hydroxyproline), Aib (2- aminoisobutyric acid), Acpc or Tyr. In certain embodiments, Xaa3is an aliphatic amino acid such as Vai, Leu, He or Nle, typically Vai or Nle. Xaa4is Tyr, Tyr(POs), Thr, Ser, homoSer (homoserine), azaTyr (aza-od-homo-L- tyrosine) or Ala. In certain embodiments, Xaa4is a hydroxyl-substituted amino acid such as Tyr, Ser or Thr, typically Tyr. Xaa5is He, Ala, Leu, norLeu, Vai or Gly. In certain embodiments, Xaa5is an aliphatic amino acid such as Vai, Leu, He or Nle, typically He. Xaa6is His, Arg or 6-NH2-Phe (6-aminophenylalaine). In certain embodiments, Xaa6is a fully or partially positively-charged amino acid such as Arg or His. Xaa7is Cys, Pro or Ala.

[0117] In certain embodiments, one or more of Xaa^Xaa7is identical to the corresponding amino acid in naturally-occurring Ang-(l-7). In certain such embodiments, all but one or two of Xaa1- Xaa7are identical to the corresponding amino acid in naturally-occurring Ang-(l-7). In other embodiments, all of Xaax-Xaa6are identical to the corresponding amino acid in naturally-occurring Ang-(l-7).

[0118] In certain embodiments, Xaa3is Nle. When Xaa3is Nle, one or more of Xaa1- Xaa2and Xaa4'7are optionally identical to the corresponding amino acid in naturally- occurring Ang-(l-7). In certain such embodiments, all but one or two of Xaax-Xaa2and Xaa4'7are identical to the corresponding amino acid in naturally-occurring Ang- (1-7). In other embodiments, all of Xaa1-Xaa2and Xaa4'7are identical to the corresponding amino acid in naturally-occurring Ang-(l-7), resulting in the amino acid sequence: Asp1-Arg2-Nle3-Tyr4-He5-His6-Pro7(SEQ ID NO: 5).

[0119] In certain embodiments, the peptide has the amino acid sequence Asp^Arg2- Val3- Ser4-He5-His6-Cys7(SEQ ID NO: 2) or Ala1-Arg2-Val3-Ser4-He5-His6-Cys7(SEQ ID NO: 3).

[0120] In some embodiments, a linear angiotensin (1-7) peptide as described herein is a peptide having a sequence of Asp1-Arg2-Val3-Tyr4-He5-His6-Pro7-Phe8-His9(SEQ IDNO: 19), whichis identical to the sequence of Ang(l-9). In some embodiments, an angiotensin (1-7) peptide is a derivative of Ang (1-9). For exemplary Ang (1-9) peptides, including Ang(l-9) derivatives, see U.S. Patent Publication 2012 / 0172301, the disclosure of which is hereby incorporated by reference.

[0121] In some embodiments, a linear angiotensin (1-7) peptide is a peptide with an amino acid sequence of Ala1-Arg2-Val3-Tyr4-He5-His6-Pro7(SEQ ID NO: 20). Additional sequences derived from SEQ ID NO: 20 may be found in European Patent Application 2,264,048, the disclosure of which is hereby incorporated by reference.

[0122] Exemplary Cyclic Angiotensin (1-7) Peptides

[0123] In certain aspects, the invention provides a cyclic angiotensin-(l-7) (Ang-(1- 7)) peptide analog comprising a linkage, such as between the side chains of amino acids corresponding to positions Tyr4and Pro7in Ang. These peptide analogs typically comprise 7 amino acid residues, but can also include a cleavable sequence. As discussed in greater detail below, the invention includes fragments and analogs where one or more amino acids are substituted by another amino acid (including fragments). One example of such an analog is Asp1-Arg2-Val3-Ser4-Ile5-His6-Cys7(SEQ ID NO: 2), wherein a linkage is formed between Ser4and Cys7. Another example of such an analog is Ala1-Arg2-Val3-Ser4-Ile5-His6-Cys7(SEQ ID NO: 3), wherein a linkage is formed between Ser4and Cys7. In some embodiments, a cyclic angiotensin (1-7) peptide analog is a cyclic analog that does not have a sequence according to SEQ ID NO: 1. In some embodiments, a cyclic angiotensin (1-7) peptide analog is a cyclic analog that does not have a sequence according to SEQ ID NO: 2. In some embodiments, a cyclic angiotensin (1-7) peptide analog is a cyclic analog that does not have a sequence according to SEQ ID NO: 3.

[0124] Although the following section describes aspects of the invention in terms of a thioether bond linking residues at the 4- and 7-positions, it should be understood that other linkages (as described above) could replace the thioether bridge and that other residues could be cyclized. A thioether bridge is also referred to as a monosulfide bridge or, in the case of Ala-S-Ala, as a lanthionine bridge. Thioether bridgecontaining peptides can be formed by two amino acids having one of the following formulas:

[0125] Formula (I)

[0126]

[0127] Formula (III)

[0128] In these formulae, R1, R2, R3, R4, R5and R6are independently -H, an alkyl (e.g., Ci-Ce alkyl, C1-C4 alkyl) or an aralkyl group, where the alkyl and aralkyl groups are optionally substituted with one or more halogen, -OH or -NRR’ groups (where R and R’ are independently -H or C1-C4 alkyl). In certain embodiments, R1, R2, R3, R4, R5and R6are each independently -H or -CH3, such where all are -H.

[0129] In certain embodiments, the invention provides an Ang analog or derivative comprising a thioether bridge according to formula (I). Typically, R1, R2, R3and R4are independently selected from -H and -CH3. Peptides comprising a thioether bridge according to formula (I) can be produced, for example, by lantibiotic enzymes or by sulfur extrusion of a disulfide. In one example, the disulfide from which the sulfur is extruded can be formed by D-cysteine in position 4 and L- cysteine in position 7 or by D-cysteine in position 4 and L-penicillamine in position 7 (see, e.g., Galande, Trent and Spatola (2003) Biopolymers 71, 534-551).

[0130] In other embodiments, the linkage of the two amino acids can be the bridges depicted in Formula (II) or Formula (III). Peptides comprising a thioether bridge according to Formula (II) can be made, for example, by sulfur extrusion of a disulfide formed by D-homocysteine in position 4 and L-cysteine in position 7. Similarly, peptides comprising a thioether bridge as in Formula (III) can be made, for example, by sulfur extrusion of a disulfide formed by D-cysteine in position 4 and L- homocysteine in position 7.

[0131] As discussed above, the Ang analogs and derivatives of the invention vary in length and amino acid composition. The Ang analogs and derivatives of the invention preferably have biological activity or are an inactive precursor molecule (inactive with regard to Ang-(l-7) function, the precursor may nevertheless have other function and be active in that regard) that can be proteolytically activated (such as how angiotensin(I), with 10 amino acids, is converted to active fragments by cleavage of 2 amino acids). The size of an Ang analog or derivative can vary but is typically between from about 5 to 10 amino acids, as long as the "core" pentameric segment comprising the 3-7 Nle-thioether-ring structure is encompassed. The amino acid sequence of an analog or derivative of the invention can vary, typically provided that it is biologically active or can become proteolytically activated.

[0132] Generally, a precursor of Ang-(l-7) or a functional variant thereof is a molecule, preferably a peptide, which can be proteolytically activated; e.g. like Ang- (1-7) is generated by the cleavage of Angiotensin II by ACE2, by the cleavage of Angiotensin I by neprilysin (NEP), or by the cleavage of Ang-(l-9) by ACE / NEP). Thus, the precursor is a molecule that can be proteolytically activated to have the function of being a Mas and / or MrgD ligand, more specifically to have the functions described herein for the functional variant. The proteolytical activation, which preferably is a proteolytical cleavage, results in Ang-(l-7) or the functional variant thereof.

[0133] The precursor can be of any length. It may be 6-25 amino acids (e.g., 6-20, 6-15, 6-14, 6-13, 6-12, 6-11, 6-10, 6-9, 6-8, 6-7 amino acids) or preferably 7-25 amino acids (e.g., 7-20, 7-15, 7-14, 7-13, 7-12, 7-11, 7-10, 7-9, 7-8, 7 amino acids) long, but more preferably it is from 8-25 amino acids (e.g., 8-20, 8-15, 8-14, 8-13, 8-12, 8-11, 8-10, 8- 9, 8 amino acids), even more preferably from 9-25 amino acids (e.g., 9-20, 9-15, 9-14, 9- 13, 9-12, 9-11, 9-10, 9 amino acids) or even more preferably 10-25 amino acids (e.g., 10- 20, 10-15, 10-14, 10-13, 10-12, 10-11, 10 amino acids) long, narrower ranges being preferred. In some embodiments, the precursor is 6, 7, 8, 9, 10, 11, 12, 13, 14, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acids long. In embodiments in which it is 6 or 7 amino acids long, it preferably has a deletion of the first two or the first amino acid position(s), respectively, corresponding to positions 1 and 2 of the heptamers described herein (e.g. SEQ ID NO: 1). In other words, the precursor preferably comprises amino acids downstream of the position corresponding to position 7 of the heptamers described herein, that can be cleaved off.

[0134] In a specific embodiment, the precursor is a peptide having a sequence of Asp1- Arg2-Val3-Tyr4-Ile5-His6-Pro7-Phe8-His9(SEQ ID NO: 19), which is the sequence of Ang-(l-9). The functional variant is a molecule, preferably a peptide, which is functional substantially as Ang-(l-7). It encompasses analogs and derivatives described herein, in particular those which are not precursors as defined above. All definitions and embodiments described herein for analogs and derivatives apply to functional variants, unless the context dictates otherwise. “Functional” means, in line with the characterization of Ang-(l-7), that the variant is capable of activating Mas receptor, MrgD receptor and / or AT2 receptor. Preferably, like Ang-(l-7), it may also have an activity such as nitric oxide release, vasodilation, improved endothelial function, antidiuresis, or one of the other properties discussed herein.

[0135] The biological activity of an analog or derivative can be determined using methods known in the art, including radioligand binding studies, in vitro cell activation assays and in vivo experiments. See, for example, Godeny and Sayeski, (2006) Am. J. Physiol. Cell. Physiol. 291:0297-1307; Sarr e / rz / ., Cardiovasc. Res. (2006) 71 :794-802; and Koziarz et al., (1993) Gen. Pharmacol. 24:705- 713.

[0136] It should be understood that the functional variant (or the precursor, once processed to its active form) is one which has one or more functions of Ang-(l-7).

[0137] The functional variant can be of any length, preferably it is from 4-25 amino acids (e.g., 4-20, 4-15, 4-14, 4-13, 4-12, 4-11, 4-10, 4-9, 4-8, 4-7 amino acids), preferably from

[0138] 5-25 amino acids (e.g., 5-20, 5-15, 5-14, 5-13, 5-12, 5-11, 5-10, 5-9, 5-8, 5-7 amino acids), more preferably from 6-25 amino acids (e.g., 6-20, 6-15, 6-14, 6-13, 6-12, 6-11,

[0139] 6-10, 6-9, 6-8, 6-7 amino acids) or even more preferably 7-25 amino acids (e.g., 7-20, 7- 15, 7-14, 7-13, 7-12, 7-11, 7-10, 7-9, 7-8, 7 amino acids) long, narrower ranges being preferred. In some embodiments, the functional variant is 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acids long. In a preferred embodiment, the length of the functional variant is from 5 to 10 amino acids, preferably as long as the core pentameric segment comprising positions 4-7 of the heptamers described herein, e.g. SEQ ID NO: 1 or the 4-7 Nle-thioether-ring structure described below, is encompassed. Typically, the functional variant shares amino acid sequence identity with Ang- (1-7). Preferably, the functional variant contains a sequence that includes at least 3 (e.g., at least 4, at least 5, at least 6, or at least 7) amino acids from the seven amino acids of Ang-(l-7) in the same order. The functional variant of Ang-(l-7) can also be defined to have a sequence at least 50% (e.g., at least 60%, 70%, 80%, 85%, or 90%) identical to the amino acid sequence of Ang-(l-7). The functional variant may contain amino acid substitutions, deletions and / or insertions in the sequence of Ang-(l-7). For example, one or more amino acid residues within Ang-(l-7) can be substituted by another amino acid. Preferred positions of Ang-(l-7) for substitution are 1, 4 and / or 7 (e.g. 1, 4, 7, 1+4, 1+7, 4+7 or 1+4+7). Substitution for an amino acid within the sequence may be selected from other members of the class to which the amino acid belongs (conservative substitution), as described elsewhere herein.

[0140] The functional variant may also comprise one or more substitutions with D-amino acids and / or non-natural (i.e. un-natural) amino acids, which are described in more detail elsewhere herein. The functional variant may also comprise substitution of non-naturally- occurring amino acids for natural amino acids, which are described in more detail elsewhere herein. The functional variant can be generally represented by the following sequence: Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7, which is described in more detail elsewhere herein. In a preferred embodiment, the functional variant is a non-cyclic peptide, i.e. a linear peptide. An example of such a linear peptide is SEQ ID NO: 20. Particular preferred examples are SEQ ID NO: 2 and SEQ ID NO: 3 (in linear but also in cyclic form). However, the functional variant may instead be a cyclic peptide, as described in more detail elsewhere herein. The functional variant, while typically a peptide, may also be a peptide mimetic, as described in more detail elsewhere herein. The functional variant does not comprise Ang-(l-8), i.e. amino acids 1-8 of SEQ ID NO: 4. Preferably, it also does not comprise amino acids 1-10, 1-11 and / or 1-12 of SEQ ID NO: 11.

[0141] Ang analogs and derivatives where only the length of the peptide is varied include the following: a 4,7-cyclized analog designated [Cyc4'7]Ang-(l-7), which is derived from natural Ang-(l-7) (Asp1-Arg2-Val3-Cyc4-Ile5-His6-Cyc7, SEQ ID NO: 6). a 4,7-cyclized analog designated [Nle3, Cyc4'7]Ang-(l-10), which is derived from natural Angiotensin I (Ang-(l-lO)) (Asp1-Arg2-Nle3-Cyc4-Ile5-His6-Cyc7-Phe8-His9- Leu10, SEQ ID NO: 7); a 4,7-cyclized analog designated [Nle3, Cyc4'7]Ang-(l-8), which is derived from natural Angiotensin II (Ang-(l-8)) (Asp1-Arg2-Nle3-Cyc4-Ile5-His6-Cyc7-Phe8, SEQ ID NO: 8); a 4,7-cyclised analog designated [Nle3, Cyc4'7]Ang-(2-8), which is derived from natural Angiotensin III (Ang-(2-8)) (Arg2-Nle3-Cyc4-Ile5-His6-Cyc7-Phe8, SEQ ID NO: 9); a 4,7-cyclised analog designated [Nle3, Cyc4'7]Ang-(3-8), which is derived from natural Angiotensin IV (Ang-(3-8)) (Nle3-Cyc4-Ile5-His6-Cyc7-Phe8, SEQ ID NO: 10); a 4,7-cyclised analog designated [Nle3, Cyc4'7]Ang-(l-7) derived from natural Ang- (1-7) (Asp1-

[0142] Arg2-Nle3-Cyc4-Ile5-His6-Cyc7, SEQ ID NO: 11); and a 4,7-cyclised analog designated [Nle3, Cyc4'7]Ang-(l-9) derived from natural Ang-(1-

[0143] 9) (Asp1- Arg2-Nle3-Cyc4-Ile5-His6-Cyc7-Phe8-His9, SEQ ID NO: 12).

[0144] These analogs can have one of the thioether bridges shown in Formulae (I)- (III) as the Cyc4'7moiety, for example, where Cyc4and Cyc7are represented by Formula (I), such as where R'-p4are each -H or -CEE, typically -H.

[0145] As compared to the amino acid sequence of the natural angiotensin peptide, the amino acids at positions 4 and 7 of the Cyc4'7analog are modified to allow introduction of the thioether-ring structures shown above. In addition to the length of the Ang analogs, the amino acids at positions other than 3, 4 and 7 can be the same or different from the naturally-occurring peptide, typically provided that the analog retains a biological function. For analogs of inactive precursors, like [Cyc4'7]Ang-(l-

[0146] 10), biological function refers to one or both of an analog’s susceptibility to angiotensin-converting enzymes that can cleave it to a biologically active fragment (e.g. Ang-(l-8) or Ang-(l-7)) or the biological activity of the fragment itself. In certain embodiments, an Ang analog or derivative of the invention has no intrinsic function but inhibits the effects of one or more naturally- occurring angiotensin compounds.

[0147] In certain embodiments, an Ang analog of the invention is represented by Formula (IV): Xaa1-Xaa2-Xaa3-Cyc4-Xaa5-Xaa6-Cyc7(IV)

[0148] Generally, Cyc herein refers to an amino acid position mediating cyclizisation

[0149] Xaa1is any amino acid, but typically a negatively-charged amino acid such as Glu or Asp, more typically Asp. Xaa2is a positively-charged amino acid such as Arg or Lys, typically Arg. Xaa3is an aliphatic amino acid, such as Leu, He or Vai, typically Vai. Cyc4forms a thioether bridge in conjunction with Cyc7. Cyc4can be a D-stereoisomer and / or a L-stereoisomer, typically a D- stereoisomer. Examples of Cyc4(taken with Cyc7) are shown in Formulas (I), (II) and (III). Typically, the R groups in Formulae (I), (II) and (III) are -H or -CH3, especially -H. Xaa5is an aliphatic amino acid, such as Leu, He or Vai, typically He. Xaa6is His. Cyc7forms a thioether bridge in conjunction with Cyc4, such as in Formula (I), (II) or (III). Cyc7can be a D-stereoisomer and / or a L-stereoisomer, typically a L-stereoisomer. Examples of Cyc7(taken with Cyc4) are shown in Formulas (I), (II), (III) and (IV). Typically, the R groups in Formulae (I), (II),) and (III) and (IV) are -H or -CH3, especially -H.

[0150] In certain embodiments, one or more of Xaax-Xaa6(excluding Cyc4and Cyc7) is identical to the corresponding amino acid in naturally-occurring Ang-(l-7). In certain such embodiments, all but one or two of Xaax-Xaa6are identical to the corresponding amino acid in naturally-occurring Ang-(l-7). In other embodiments, all of Xaax-Xaa6are identical to the corresponding amino acid in naturally-occurring Ang-(l-7).

[0151] In certain embodiments, Cyc4and Cyc7are independently selected from Abu (2- aminobutyric acid) and Ala (alanine), where Ala is present in at least one position. Thus, cyclic analogs can have a thioether linkage formed by -Ala4-S-Ala7- (Formula (I), where R'-R4are each -H); - Ala4-S-Abu7- (Formula (I): R'-R3are -H and R4is - CH3) or -Abu4-S-Ala7- (Formula (I): R1, R3and R4are -H and R2is -CH3). Specific examples of cyclic analogs comprise a -Abu4-S-Ala7- or -Ala4-S-Ala7- linkage.

[0152] In certain embodiments, the invention provides an Ang-(l-7) analog with a thioether-b ridge between position 4 and position 7 having the amino acid sequence Asp1-Arg2-Val3- Abu4-He5-His6-Ala7(SEQ ID NO: 13) or the amino acid sequence Asp1-Arg2-Val3-Ala4-He5-His6-Ala7(SEQ ID NO: 14), which are represented by the following structural diagrams (specifically formulae):

[0153] In certain embodiments, an Ang analog or derivative of the invention is represented by Formula (V): Xaa1-Xaa2-Nle3-Cyc4-Xaa5-Xaa6-Cyc7-Xaa8-Xaa9- Xaa10(V)

[0154] As discussed above, one or more of Xaa1, Xaa2, Xaa8, Xaa9and Xaa10are absent in certain embodiments. For example, (1) Xaa10is absent, (2) Xaa9and Xaa10are absent, (3) Xaa8, Xaa9and Xaa10are absent, (4) Xaa1is absent, (5) Xaa1and Xaa10 are absent, (6) Xaa1, Xaa9and Xaa10are absent, (7) Xaa1, Xaa8, Xaa9and Xaa10are absent, (8) Xaa1and Xaa2are absent, (9) Xaa1, Xaa2and Xaa10are absent, (10) Xaa1, Xaa2, Xaa9and Xaa10are absent, or (11) Xaa1, Xaa2, Xaa8, Xaa9and Xaa10are absent. For each of these embodiments, the remaining amino acids have the values described below.

[0155] Xaa1, when present, is any amino acid, but typically a negatively charged amino acid such as Glu or Asp, more typically Asp. Xaa2, when present, is a positively charged amino acid such as Arg or Lys, typically Arg. Nle3is norleucine. Cyc4forms a thioether bridge in conjunction with Cyc7. Cyc4can be a D-stereoisomer and / or a L- stereoisomer, typically a D-stereoisomer. Examples of Cyc4(taken with Cyc7) are shown in Formulas (I), (II) and (III). Typically, the R groups in Formulae (I), (II) and (III) are -H or -CH?, especially -H. Xaa5is an aliphatic amino acid, such as Leu, Nle, He or Vai, typically He.

[0156] Xaa6is His. Cyc7forms a thioether bridge in conjunction with Cyc4, such as in Formula (I), (II) or (III). Cyc7can be a D-stereoisomer and / or a L-stereoisomer, typically a L-stereoisomer. Examples of Cyc7(taken with Cyc4) are shown in Formulas (I), (II) and (III). Typically, the R groups in Formulae (I), (II) and (III) are -H or -CH3, especially -H. Xaa8, when present, is an amino acid other than Pro, typically Phe or He. In certain embodiments, He results in an inhibitor of Ang(l-8). In certain embodiments, Phe maintains the biological activity of Ang(l-8) or Ang(l- 10).

[0157] Xaa9, when present, is His. Xaa10, when present, is an aliphatic residue, for example, He, Vai or Leu, typically Leu.

[0158] In certain embodiments, one or more of Xaa^Xaa10(excluding Nle3, Cyc4and Cyc7) is identical to the corresponding amino acid in naturally-occurring Ang (including Ang-(l-7), Ang(l-8), Ang(l-9), Ang(l-lO), Ang(2-7), Ang(2-8), Ang(2- 9), Ang(2-10), Ang(3-8), Ang(3-9) and Ang(3-10). In certain such embodiments, all but one or two of Xaa^Xaa10(for those present) are identical to the corresponding amino acid in naturally-occurring Ang. In other embodiments, all of Xaa^Xaa10(for those present) are identical to the corresponding amino acid in naturally-occurring Ang.

[0159] In certain embodiments, Cyc4and Cyc7are independently selected from Abu (2-aminobutyric acid) and Ala (alanine), where Ala is present at least one position. Thus, encompassed are cyclic analogs comprising a thioether linkage formed by - Ala4-S-Ala7- (Formula (I), where R'-R4are each -H); -Ala4-S-Abu7- (Formula (I): R1- R3are -H and R4is -CH3) or -Abu4-S-Ala7- (Formula (I): R1, R3and R4are -H and R2is -CH3). Specific cyclic analogs comprise a -Abu4-S-Ala7- or -Ala4-S-Ala7- linkage.

[0160] In particular, the invention provides an Ang-(l-7) analog or derivative with a thioether-b ridge between position 4 and position 7 having the amino acid sequence Asp'-Arg2-Nle3- Abu4-Ile5-His6-Ala7(SEQ ID NO: 15) or the amino acid sequence Asp1-Arg2-Nle3-Ala4-Ile5-His6-Ala7(SEQ ID NO: 16).

[0161] In another aspect, the invention provides an Ang-(l-8) analog or derivative with a thioether-b ridge between position 4 and position 7 having Ang-(l-8) antagonistic activity, in particular an Ang(l-8) analog or derivative having the amino acid sequence Asp1-Arg2-Nle3-Abu4-Ile5- His6-Ala7-Ile8(SEQ ID NO: 17), or the amino acid sequence Asp1-Arg2-Nle3-Ala4-Ile5-His6-Ala7-Ile8(SEQ ID NO: 18).

[0162] An alkyl group is a straight chained or branched non-aromatic hydrocarbon that is completely saturated. Typically, a straight chained or branched alkyl group has from 1 to about 20 carbon atoms, preferably from 1 to about 10. Examples of straight chained and branched alkyl groups include methyl, ethyl, n-propyl, iso-propyl, n- butyl, sec-butyl, tert-butyl, pentyl, hexyl, pentyl and octyl. A C1-C4 straight chained or branched alkyl group is also referred to as a "lower alkyl" group.

[0163] An aralkyl group is an alkyl group substituted by an aryl group. Aromatic (aryl) groups include carbocyclic aromatic groups such as phenyl, naphthyl, and anthracyl, and heteroaryl groups such as imidazolyl, thienyl, furyl, pyridyl, pyrimidyl, pyranyl, pyrazolyl, pyrrolyl, pyrazinyl, thiazolyl, oxazolyl, and tetrazolyl. Aromatic groups also include fused polycyclic aromatic ring systems in which a carbocyclic aromatic ring or heteroaryl ring is fused to one or more other heteroaryl rings. Examples include benzothienyl, benzofuryl, indolyl, quinolinyl, benzothiazole, benzoxazole, benzimidazole, quinolinyl, isoquinolinyl andisoindolyl. Ang-(l-7) Receptor Agonists

[0164] In some embodiments, the present invention provides methods of treating optic neuropathy including administering to a subject who is suffering from optic neuropathy an angiotensin (1-7) receptor agonist. As used herein, the term “angiotensin-(l-7) receptor agonist” encompasses any molecule that has a positive impact in a function of an angiotensin-(l-7) receptor, e.g. Mas receptor and / or MrgD receptor, in particular, the G-protein coupled Mas receptor. In some embodiments, an angiotensin- (1-7) receptor agonist directly or indirectly enhances, strengthens, activates and / or increases an angiotensin-(l-7) receptor (in particular , the Mas receptor) activity. In some embodiments, an angiotensin- (1-7) receptor agonist directly interacts with an angiotensin-(l-7) receptor (in particular , the Mas receptor). Such agonists can be peptidic or non-peptidic including, e.g., proteins, chemical compounds, small molecules, nucleic acids, antibodies, drugs, ligands, or other agents. In some embodiments, the angiotensin (1-7) receptor agonist is a non-peptidic agonist.

[0165] An exemplary class of angiotensin-(l-7) receptor agonists are l-(p- thienylbenzyl)imidazoles. Examples of these non-peptide angiotensin-(l-7) receptor agonists are represented by Structural Formula (VI):

[0166]

[0167] (VI), or pharmaceutically acceptable salts thereof, wherein:

[0168] R1is halogen, hydroxyl, (Ci-C4)-alkoxy, (Ci-Cs)-alkoxy wherein 1 to 6 carbon atoms are replaced by the heteroatoms O, S, or NH (preferably by O), (Ci-C4)-alkoxy substituted by a saturated cyclic ether such as tetrahydropyran or tetrahydrofuran, O-(Ci- C4)-alkenyl, O-(Ci-C4)-alkylaryl, or aryloxy that is unsubstituted or substituted by a substituent selected from halogen, (Ci-C3)-alkyl, (Ci- C3)-alkoxy and trifluoromethyl;

[0169] R2is CHO, COOH, or (3) CO-O-(Ci-C4)-alkyl; R3is (Ci-C4)-alkyl or aryl; R4is hydrogen, halogen (chloro, bromo, fluoro), or (Ci-C4)-alkyl; X is oxygen or sulfur;

[0170] Y is oxygen or -NH-; R5is hydrogen, (Ci-C6)-alkyl; or (Ci-C4)-alkylaryl, where R5is hydrogen when Y is -NH-; and R6is (Ci-Cs)-alkyl.

[0171] In certain embodiments, R1is not halogen when R2is COOH or CO-O-(Ci-C4)- alkyl.

[0172] In some embodiments, an angiotensin-(l-7) receptor agonist is AVE 0991, 5- formyl- 4-methoxy-2-phenyl-l[[4-[2-(ethylaminocarbonylsulfonamido)-5-isobutyl-3- thienyl]-phenyl]- m ethyl ]-imidazole, which is represented by the following structure:

[0173]

[0174] Another exemplary class of angiotensin-(l-7) receptor agonists are p- thienylbenzylamides. Examples of these non-peptide angiotensin-(l-7) receptor agonists are represented by Structural Formula (VII):

[0175] (VII), or a pharmaceutically acceptable salt thereof, wherein:

[0176] R1is (Ci-Cs)-alkyl that is unsubstituted or substituted by a radical chosen from NH2, halogen, O-(Ci-C3)-alkyl, CO-O-(Ci-C3)-alkyl andCO2H, (C3-C8)-cycloalkyl, (Ci- C3)-alkyl-(C3-C8)-cycloalkyl, (Ce-C )- aryl that is unsubstituted or substituted by a radical chosen from halogen and O-(Ci-C3)-alkyl, (Ci-C3)- alkyl-(C6-Cio)-aryl where the aryl radical is unsubstituted or substituted by a radical chosen from halogen and O-(Ci- C3)-alkyl, (Ci-C5)-heteroaryl, or (Ci-C3)-alkyl-(Ci-Cs)-heteroaryl; R2is hydrogen, (Ci- Cg)-alkyl that is unsubstituted or substituted by a radical chosen from halogen and O- (Ci-C3)-alkyl, (C3-Cs)-cycloalkyl, (Ci-C3)-alkyl-(C3-C8)-cycloalkyl, (Ce-Cio)-aryl that is unsubstituted or substituted by a radical chosen from among halogen, O-(Ci-C3)-alkyl and CO-O-(Ci-C3)-alkyl, or (Ci-C3)-alkyl-(Ce-Cio)-aryl that is unsubstituted or substituted by a radical chosen from halogen and O-(Ci-C3)-alkyl; R3is hydrogen, COOH, or COO-(Ci-C4)-alkyl; R4is hydrogen, halogen; or (Ci-C4)-alkyl; R5is hydrogen or (Ci-Ce)-alkyl; R6is hydrogen, (Ci-Ce)-alkyl, (Ci-C3)-alkyl-(C3-Cg)- cycloalkyl, or (C2-Ce)-alkenyl; and X is oxygen orNH.

[0177] Additional examples of angiotensin-(l-7) receptor agonists are described in U.S. Patent No. 6,235,766, the contents of which are incorporated by reference herein.

[0178] Various angiotensin-(l-7) receptor agonists described above can be present as pharmaceutically acceptable salts. As used herein, “a pharmaceutically acceptable salt” refers to salts that retain the desired activity of the peptide or equivalent compound, but preferably do not detrimentally affect the activity of the peptide or other component of a system, which uses the peptide. Examples of such salts are acid addition salts formed with inorganic acids, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, and the like. Salts may also be formed with organic acids such as, for example, acetic acid, oxalic acid, tartaric acid, succinic acid, maleic acid, fumaric acid, gluconic acid, citric acid, malic acid, ascorbic acid, benzoic acid, tannic acid, pamoic acid, alginic acid, polyglutamic acid, and the like. Salts formed from a cationic material may utilize the conjugate base of these inorganic and organic acids. Salts may also be formed with polyvalent metal cations such as zinc, calcium, bismuth, barium, magnesium, aluminum, copper, cobalt, nickel and the like or with an organic cation formed from N,N'- dibenzylethylenediamine or ethylenediamine, or combinations thereof (e.g., a zinc tannate salt).

[0179] The non-toxic, physiologically acceptable salts are preferred.

[0180] The salts can be formed by conventional means such as by reacting the free acid or free base forms of the product with one or more equivalents of the appropriate acid or base in a solvent or medium in which the salt is insoluble, or in a solvent such as water which is then removed in vacuo or by freeze-drying, or by exchanging the cations of an existing salt for another cation on a suitable ion exchange resin. An alkyl group is a straight chained or branched non-aromatic hydrocarbon that is completely saturated. Typically, a straight chained or branched alkyl group has from 1 to about 20 carbon atoms, preferably from 1 to about 10. Examples of straight chained and branched alkyl groups include methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, tertbutyl, pentyl, hexyl, pentyl and octyl. A C 1-C4 straight chained or branched alkyl group is also referred to as a “lower alkyl” group.

[0181] An alkenyl group is a straight chained or branched non-aromatic hydrocarbon that is includes one or more double bonds. Typically, a straight chained or branched alkenyl group has from 2 to about 20 carbon atoms, preferably from 2 to about 10. Examples of straight chained and branched alkenyl groups include ethenyl, n-propenyl, andn-butenyl. Aromatic (aryl) groups include carbocyclic aromatic groups such as phenyl, naphthyl, and anthracyl, and heteroaryl groups such as imidazolyl, thienyl, furyl, pyridyl, pyrimidyl, pyranyl, pyrazolyl, pyrrolyl, pyrazinyl, thiazolyl, oxazolyl, and tetrazolyl. Aromatic groups also include fused polycyclic aromatic ring systems in which a carbocyclic aromatic ring or heteroaryl ring is fused to one or more other heteroaryl rings. Examples include benzothienyl, benzofuryl, indolyl, quinolinyl, benzothiazole, benzoxazole, benzimidazole, quinolinyl, isoquinolinyl and isoindolyl.

[0182] An aralkyl group is an alkyl group substituted by an aryl group.

[0183] Formulations and Dosing

[0184] In accordance with the methods of the invention, an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist as described herein of the invention can be administered to a subject alone (e.g., as a purified peptide or compound), or as a component of a composition (in particular a pharmaceutical composition) or medicament (e.g., in the manufacture of a medicament for the treatment of the disease), as described herein or otherwise known in the art. The compositions can be formulated with a physiologically acceptable carrier or excipient to prepare a pharmaceutical composition. The carrier and composition can be sterile. The formulation should suit the mode of administration, for example intravenous or subcutaneous administration. Methods of formulating compositions are known in the art (see, e.g., Remington's Pharmaceuticals Sciences, 17th Edition, Mack Publishing Co., (Alfonso R. Gennaro, editor) (1989)).

[0185] Suitable pharmaceutically acceptable carriers include but are not limited to water, salt solutions (e.g., NaCl), saline, buffered saline, alcohols, glycerol, ethanol, gum arabic, vegetable oils, benzyl alcohols, polyethylene glycols, gelatin, carbohydrates such as lactose, amylose or starch, sugars such as mannitol, sucrose, or others, dextrose, magnesium stearate, talc, silicic acid, viscous paraffin, perfume oil, fatty acid esters, hydroxymethylcellulose, polyvinyl pyrolidone, etc., as well as combinations thereof. The pharmaceutical preparations can, if desired, be mixed with auxiliary agents (e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring and / or aromatic substances and the like), which do not deleteriously react with the active compounds or interference with their activity. In a preferred embodiment, a water-soluble carrier suitable for intravenous administration is used.

[0186] The composition or medicament, if desired, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. The composition can be a liquid solution, suspension, emulsion, sustained release formulation, or powder. The composition can also be formulated as a suppository, with traditional binders and carriers such as triglycerides.

[0187] The composition or medicament can be formulated in accordance with the routine procedures as a pharmaceutical composition adapted for administration to human beings. For example, in some embodiments, a composition for intravenous administration typically is a solution in sterile isotonic aqueous buffer. Where necessary, the composition may also include a solubilizing agent and a local anesthetic to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or water free concentrate in a hermetically sealed container such as an ampule or sachette indicating the quantity of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water, saline or dextrose / water. Where the composition is administered by injection, an ampule of sterile water for injection or saline can be provided so that the ingredients may be mixed prior to administration.

[0188] An Ang-(l-7) peptide or angiotensin (1-7) receptor agonist as described herein can be formulated as neutral or salt forms. Pharmaceutically acceptable salts include those formed with free amino groups such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with free carboxyl groups such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

[0189] An Ang-( 1-7) peptide or angiotensin (1-7) receptor agonist as described herein (or a composition or medicament containing an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist described herein) is administered by any appropriate route. In some embodiments, an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist described herein is administered subcutaneously. As used herein, the term “subcutaneous tissue”, is defined as a layer of loose, irregular connective tissue immediately beneath the skin. For example, the subcutaneous administration may be performed by injecting a composition into areas including, but not limited to, thigh region, abdominal region, gluteal region, or scapular region. In some embodiments, an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist described herein is administered intravenously. Alternatively, an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist described herein (or a composition or medicament containing an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist described herein) can be administered by inhalation, parenterally, intradermally, transdermally, rectally, or transmucosally. In some embodiments, an Ang(l-7) peptide or angiotensin (1-7) receptor agonist is administered orally. More than one route can be used concurrently, if desired.

[0190] In some embodiments, a composition is administered in a therapeutically effective amount and / or according to a dosing regimen that is correlated with a particular desired outcome (e.g., with treating or reducing risk for optic neuropathy).

[0191] Particular doses or amounts to be administered in accordance with the present invention may vary, for example, depending on the nature and / or extent of the desired outcome, on particulars of route and / or timing of administration, and / or on one or more characteristics (e.g., weight, age, personal history, genetic characteristic, lifestyle parameter, severity of cardiac defect and / or level of risk of cardiac defect, etc., or combinations thereof). Such doses or amounts can be determined by those of ordinary skill. In some embodiments, an appropriate dose or amount is determined in accordance with standard clinical techniques. For example, in some embodiments, an appropriate dose or amount is a dose or amount sufficient to reduce a disease severity index score by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100% or more. For example, in some embodiments, an appropriate dose or amount is a dose or amount sufficient to reduce a disease severity index score by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100%. Alternatively or additionally, in some embodiments, an appropriate dose or amount is determined through use of one or more in vitro or in vivo assays to help identify desirable or optimal dosage ranges or amounts to be administered.

[0192] In various embodiments, an Ang-(l-7) peptide or angiotensin (1-7) receptor agonist is administered at a therapeutically effective amount. As used herein, the term “therapeutically effective amount” is largely determined based on the total amount of the therapeutic agent contained in the pharmaceutical compositions of the present invention. Generally, a therapeutically effective amount is sufficient to achieve a meaningful benefit to the subject (e.g., treating, modulating, curing, preventing and / or ameliorating the underlying disease or condition). In some particular embodiments, appropriate doses or amounts to be administered may be extrapolated from doseresponse curves derived from in vitro or animal model test systems.

[0193] Therapeutically effective dosage amounts of angiotensin (1-7) peptides or angiotensin (1-7) receptor agonists, including derivatives, analogs, and / or salts may be present in varying amounts in various embodiments. For example, in some embodiments, a therapeutically effective amount of an angiotensin (1-7) peptide may be an amount ranging from about 10-1,000 mg (e.g., about 20 mg - 1,000 mg, 30 mg - 1,000 mg, 40 mg - 1,000 mg, 50 mg - 1,000 mg, 60 mg - 1,000 mg, 70 mg - 1,000 mg, 80 mg - 1,000 mg, 90 mg - 1,000 mg, about 10-900 mg, 10-800 mg, 10-700 mg, 10-600 mg, 10-500 mg, 100-1,000 mg, 100-900 mg, 100-800 mg, 100-700 mg, 100-600 mg, 100- 500 mg, 100-400 mg, 100-300 mg, 200-1,000 mg, 200-900 mg, 200-800 mg, 200-700 mg, 200-600 mg, 200-500 mg, 200-400 mg, 300-1,000 mg, 300-900 mg, 300-800 mg, 300-700 mg, 300-600 mg, 300-500 mg, 400 mg - 1,000 mg, 500 mg - 1,000 mg, 100 mg - 900 mg, 200 mg - 800 mg, 300 mg - 700 mg, 400 mg - 700 mg, and 500 mg - 600 mg). In some embodiments, an angiotensin (1-7) peptide or angiotensin (1-7) receptor agonist is present in an amount of or greater than about 10 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg. In some embodiments, an angiotensin (1-7) peptide or angiotensin (1-7) receptor agonist is present in an amount of or less than about 1000 mg, 950 mg, 900 mg, 850 mg, 800 mg, 750 mg, 700 mg, 650 mg, 600 mg, 550 mg, 500 mg, 450 mg, 400 mg, 350 mg, 300 mg, 250 mg, 200 mg, 150 mg, or 100 mg. In some embodiments, the therapeutically effective amount described herein is provided in one dose. In some embodiments, the therapeutically effective amount described herein is provided in one day.

[0194] In other embodiments, a therapeutically effective dosage amount may be, for example, about 0.001 mg / kg weight to 500 mg / kg weight, e.g., from about 0.001 mg / kg weight to 400 mg / kg weight, from about 0.001 mg / kg weight to 300 mg / kg weight, from about 0.001 mg / kg weight to 200 mg / kg weight, from about 0.001 mg / kg weight to 100 mg / kg weight, from about 0.001 mg / kg weight to 90 mg / kg weight, from about 0.001 mg / kg weight to 80 mg / kg weight, from about 0.001 mg / kg weight to 70 mg / kg weight, from about 0.001 mg / kg weight to 60 mg / kg weight, from about 0.001 mg / kg weight to 50 mg / kg weight, from about 0.001 mg / kg weight to 40 mg / kg weight, from about 0.001 mg / kg weight to 30 mg / kg weight, from about 0.001 mg / kg weight to 25 mg / kg weight, from about 0.001 mg / kg weight to 20 mg / kg weight, from about 0.001 mg / kg weight to 15 mg / kg weight, from about 0.001 mg / kg weight to 10 mg / kg weight. In some embodiments, the therapeutically effective amount described herein is provided in one dose. In some embodiments, the therapeutically effective amount described herein is provided in one day.

[0195] In still other embodiments, a therapeutically effective dosage amount may be, for example, about O.001 mg / kg weight to about 1 mg / kgweight, e.g. from about O.OOl mg / kg weight to about 0.9 mg / kg weight, from about 0.001 mg / kg weight to about 0.8 mg / kg weight, from about 0.001 mg / kg weight to about 0.8 mg / kg weight, from about 0.001 mg / kg weight to about 0.7 mg / kg weight, from about 0.001 mg / kg weight to about 0.6 mg / kg weight, from about 0.001 mg / kg weight to about 0.5 mg / kg weight, from about 0.01 mg / kg weight to about 1 mg / kg weight, from about 0.01 mg / kg weight to about 0.9 mg / kg weight, from about 0.01 mg / kg weight to about 0.8 mg / kg weight, from about 0.01 mg / kg weight to about 0.7 mg / kg weight, from about 0.01 mg / kg weight to about 0.6 mg / kg weight, from about 0.01 mg / kg weight to about 0.5 mg / kg weight, from about 0.02 mg / kg weight to about 1 mg / kg weight, from about 0.02 mg / kg weight to about 0.9 mg / kg weight, from about 0.02 mg / kg weight to about 0.8 mg / kg weight, from about 0.02 mg / kg weight to about 0.7 mg / kg weight, from about 0.02 mg / kg weight to about 0.6 mg / kg weight, from about 0.02 mg / kg weight to about 0.5 mg / kg weight, from about 0.03 mg / kg weight to about 1 mg / kg weight, from about 0.03 mg / kg weight to about 0.9 mg / kg weight, from about 0.03 mg / kg weight to about 0.8 mg / kg weight, from about 0.03 mg / kg weight to about 0.7 mg / kg weight, from about 0.03 mg / kg weight to about 0.6 mg / kg weight, from about 0.03 mg / kg weight to about 0.5 mg / kg weight, from about 0.04 mg / kg weight to about 1 mg / kg weight, from about 0.04 mg / kg weight to about 0.9 mg / kg weight, from about 0.04 mg / kg weight to about 0.8 mg / kg weight, from about 0.04 mg / kg weight to about 0.7 mg / kg weight, from about 0.04 mg / kg weight to about 0.6 mg / kg weight, from about 0.04 mg / kg weight to about 0.5 mg / kg weight, from about 0.05 mg / kg weight to about 1 mg / kg weight, from about 0.05 mg / kg weight to about 0.9 mg / kg weight, from about 0.05 mg / kg weight to about 0.8 mg / kg weight, from about 0.05 mg / kg weight to about 0.7 mg / kg weight, from about 0.05 mg / kg weight to about 0.6 mg / kg weight, from about 0.05 mg / kg weight to about 0.5 mg / kg weight. In some embodiments, the therapeutically effective amount described herein is provided in one dose. In some embodiments, the therapeutically effective amount described herein is provided in one day.

[0196] In still other embodiments, a therapeutically effective dosage amount may be, for example, about 0.0001 mg / kg weight to 0.1 mg / kg weight, e.g. from about 0.0001 mg / kg weight to 0.09 mg / kg weight, from about 0.0001 mg / kg weight to 0.08 mg / kg weight, from about 0.0001 mg / kg weight to 0.07 mg / kg weight, from about 0.0001 mg / kg weight to 0.06 mg / kg weight, from about 0.0001 mg / kg weight to 0.05 mg / kg weight, from about 0.0001 mg / kg weight to about 0.04 mg / kg weight, from about 0.0001 mg / kg weight to 0.03 mg / kg weight, from about 0.0001 mg / kg weight to 0.02 mg / kg weight, from about 0.0001 mg / kg weight to 0.019 mg / kg weight, from about 0.0001 mg / kg weight to 0.018 mg / kg weight, from about 0.0001 mg / kg weight to 0.017 mg / kg weight, from about 0.0001 mg / kg weight to 0.016 mg / kg weight, from about 0.0001 mg / kg weight to 0.015 mg / kg weight, from about 0.0001 mg / kg weight to 0.014 mg / kg weight, from about 0.0001 mg / kg weight to 0.013 mg / kg weight, from about 0.0001 mg / kg weight to 0.012 mg / kg weight, from about 0.0001 mg / kg weightto 0.011 mg / kg weight, from about 0.0001 mg / kg weight to 0.01 mg / kg weight, from about 0.0001 mg / kg weight to 0.009 mg / kg weight, from about 0.0001 mg / kg weight to 0.008 mg / kg weight, from about 0.0001 mg / kg weight to 0.007 mg / kg weight, from about 0.0001 mg / kg weight to 0.006 mg / kg weight, from about 0.0001 mg / kg weight to 0.005 mg / kg weight, from about 0.0001 mg / kg weight to 0.004 mg / kg weight, from about 0.0001 mg / kg weight to 0.003 mg / kg weight, from about 0.0001 mg / kg weight to 0.002 mg / kg weight. In some embodiments, the therapeutically effective dose may be 0.0001 mg / kg weight, 0.0002 mg / kg weight, 0.0003 mg / kg weight, 0.0004 mg / kg weight, 0.0005 mg / kg weight, 0.0006 mg / kg weight, 0.0007 mg / kg weight, 0.0008 mg / kg weight, 0.0009 mg / kg weight, 0.001 mg / kg weight, 0.002 mg / kg weight, 0.003 mg / kg weight, 0.004 mg / kg weight, 0.005 mg / kg weight, 0.006 mg / kg weight, 0.007 mg / kg weight, 0.008 mg / kg weight, 0.009 mg / kg weight, 0.01 mg / kg weight, 0.02 mg / kg weight, 0.03 mg / kg weight, 0.04 mg / kg weight, 0.05 mg / kg weight, 0.06 mg / kg weight, 0.07 mg / kg weight, 0.08 mg / kg weight, 0.09 mg / kg weight, or 0.1 mg / kg weight. The effective dose for a particular individual can be varied (e.g., increased or decreased) over time, depending on the needs of the individual.

[0197] In some embodiments, the angiotensin (1-7) peptide or angiotensin (1-7) receptor agonist is administered at an effective dose ranging from about 1-1,000 pg / kg / day (e.g., ranging from about 1-900 pg / kg / day, 1-800 pg / kg / day, 1-700 pg / kg / day, 1-600 pg / kg / day, 1-500 pg / kg / day, 1-400 pg / kg / day, 1-300 pg / kg / day, 1-200 pg / kg / day, 1- 100 pg / kg / day, 1-90 pg / kg / day, 1-80 pg / kg / day, 1-70 pg / kg / day, 1-60 pg / kg / day, 1-50 pg / kg / day, 1-40 pg / kg / day, 1-30 pg / kg / day, 1-20 pg / kg / day, 1-10 pg / kg / day). In some embodiments, the angiotensin (1-7) peptide or angiotensin (1-7) receptor agonist is administered at an effective dose ranging from about 1-500 pg / kg / day. In some embodiments, it is administered at an effective dose ranging from about 1-100 pg / kg / day. In some embodiments, it is administered at an effective dose ranging from about 1-60 pg / kg / day. In some embodiments, it is administered at an effective dose selected from about 1, 2, 4, 6, 8, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, or 1,000 gg / kg / day.

[0198] In some embodiments, the angiotensin (1-7) peptide is administered at an effective dose ranging from about 1-1,000 gg / kg / day, from about 50-500 gg / kg / day, or from about 1-60 gg / kg / day.

[0199] Combination Therapies

[0200] In some embodiments, an angiotensin (1-7) peptide or an angiotensin (1-7) receptor agonist will be used as a part of a combination therapy. In some embodiments, the angiotensin (1-7) peptide and / or angiotensin (1-7) receptor agonist may be administered prior to, concurrently with, or subsequent to one or more additional therapies. It is contemplated that any known therapy or therapeutic for the treatment of optic neuropathy may be used with one or more angiotensin (1-7) peptides and / or angiotensin (1-7) receptor agonists as disclosed herein. Exemplary therapies that may be used with one or more angiotensin (1-7) peptides or angiotensin (1-7) receptor agonists include, but are not limited to, medications that can help control pain and itching, medications that address complications such as sepsis (e.g., antibiotics), medications that reduce inflammation (e.g., corticosteroid), surgery to correct abnormal motion (e.g., surgery to correct fusing of finger or toes or abnormal bends in the joints), surgery to improve the ability to eat a healthy diet (e.g., surgical dilation of the esophagus or placement of a feeding tube), skin grafts (e.g., OrCel composite cultured skin), gene therapy, bone marrow transplantation, protein replacement therapy, cell-based therapies, and / or combinations thereof, among others.

[0201] Prophylactic treatments

[0202] The invention further relates to a prophylactic treatment of a subject, preferably a subject at increased risk for optic neuropathy. Accordingly, the invention relates to a method of prophylactically treating optic neuropathy comprising administering an angiotensin-(l-7) receptor agonist to a subject, preferably a subject at increased risk for optic neuropathy. Similarly, the invention relates to an angiotensin-(l-7) receptor for use in prophylactically treating optic neuropathy in a subject, preferably a subject at increased risk for optic neuropathy.

[0203] The prophylactic treatment is not meant to necessarily prevent the onset optic neuropathy, but to prevent or to slow down progression after onset and / or to prevent or reduce symptoms that would occur without the prophylactic treatment. It can also be termed “preparatory treatment”, “preventative treatment” or “preventive treatment”.

[0204] A subject at increased risk for optic neuropathy is a subject having one or more risk factors for optic neuropathy, in particular one or more of the risk factors defined above. Preferably, the risk factor is having small optic nerve openings (‘disk at risk’).

[0205] It has recently been found that the risk of optic neuropathy, in particular NAION, is increased in subjects treated with a GLP-1 agonist (Hathaway et al., JAMA Ophthalmol. 2024; 142(8):732-739). Thus, in a preferred embodiment, the angiotensin (1- 7) receptor agonist (in particular the angiotensin (1-7) peptide) is used as a part of a combination therapy with a GLP-1 agonist. Accordingly, the subject at increased risk for optic neuropathy has the risk factor of being under a GLP-1 agonist therapy. So for example, the invention relates to a method of prophylactically treating optic neuropathy comprising administering an angiotensin-(l-7) receptor agonist to a subj ect under a GLP- 1 agonist therapy; or similarly to an angiotensin-(l-7) receptor for use in prophylactically treating optic neuropathy in a subject under a GLP-1 agonist therapy. The subject may be obese and / or diabetic.

[0206] The invention also relates to a method for treating obesity and / or diabetes mellitus comprising administering a GLP-1 agonist and an angiotensin-(l-7) receptor agonist to a subject; or similarly to a GLP-1 agonist for use in treating obesity and / or diabetes mellitus in a subject, the use comprising administering an angiotensin-(l-7) receptor agonist to the subject. Therein, the angiotensin-(l-7) receptor agonist is for use in prophylactically treating optic neuropathy.

[0207] In this context, the invention also relates to a kit or to a pharmaceutical composition comprising an angiotensin-(l-7) receptor agonist and a GLP-1 agonist, preferably for the above methods or uses. The GLP-1 agonist may be an antibody, an antigen-binding fragment thereof or a small compound. Exemplary GLP-1 agonists are exenatide, liraglutide, albiglutide, dulaglutide, lixisenatide, semaglutide and tirzepatide.

[0208] Kits

[0209] In some embodiments, the present invention further provides kits or other articles of manufacture which contains an angiotensin (1-7) peptide, an angiotensin (1-7) receptor agonist or a formulation containing the same and provides instructions for its reconstitution (if lyophilized) and / or use. Kits or other articles of manufacture may include a container, a syringe, vial and any other articles, devices or equipment useful in administration (e.g., subcutaneous, by inhalation). Suitable containers include, for example, bottles, vials, syringes (e.g., pre-filled syringes), ampules, cartridges, reservoirs, or lyo-jects. The container may be formed from a variety of materials such as glass or plastic. In some embodiments, a container is a pre-filled syringe. Suitable prefilled syringes include, but are not limited to, borosilicate glass syringes with baked silicone coating, borosilicate glass syringes with sprayed silicone, or plastic resin syringes without silicone.

[0210] Typically, the container may hold one or more formulations and a label on, or associated with, the container that may indicate directions for reconstitution and / or use. For example, the label may indicate that the formulation is reconstituted to concentrations as described above. The label may further indicate that the formulation is useful or intended for, for example, subcutaneous administration. In some embodiments, a container may contain a single dose of a stable formulation containing an angiotensin (1-7) peptide or angiotensin (1-7) receptor agonist. In various embodiments, a single dose of the stable formulation is present in a volume of less than about 15 ml, 10 ml, 5.0 ml, 4.0 ml, 3.5 ml, 3.0 ml, 2.5 ml, 2.0 ml, 1.5 ml, 1.0 ml, or 0.5 ml. Alternatively, a container holding the formulation may be a multi-use vial, which allows for repeat administrations (e.g., from 2-6 administrations) of the formulation. Kits or other articles of manufacture may further include a second container comprising a suitable diluent (e.g., BWFI, saline, buffered saline). Upon mixing of the diluent and the formulation, the final protein concentration in the reconstituted formulation will generally be at least 1 mg / ml (e.g, at least 5 mg / ml, at least 10 mg / ml, at least 20 mg / ml, at least 30 mg / ml, at least 40 mg / ml, at least 50 mg / ml, at least 75 mg / ml, at least 100 mg / ml). Kits or other articles of manufacture may further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, fdters, needles, syringes, and package inserts with instructions for use. In some embodiments, kits or other articles of manufacture may include an instruction for self-administration.

[0211] EXAMPLES

[0212] While certain compositions and methods of the present invention have been described with specificity in accordance with certain embodiments, the following example serves only to illustrate the compounds and methods of the invention, and it is not intended to limit the same.

[0213] Example 1. Angiotensin (1-7) promotes functional recovery of vision in a rat model of ischemic optic neuropathy

[0214] A rat model of ischemic optic neuropathy was tested for functional recovery of vision after administration of angiotensin (1-7). Various functional measures were evaluated, predominantly, optomotry, i.e., visual acuity measured by optokinetic nystagmus read by the motion of the rat’s head in the direction of an image of moving bars, and flash visual evoked potentials (fVEPs) that record the electrical response of the signal from the eye to the visual cortex (VI) from the superior colliculus and are directly related to the number of remaining axons. Electrical responses were evaluated using standard devices, for example, using a Diagnosys-Celeris combined device which can give near simultaneous readouts of el ectroreti nogram (ERG), fVEP, pattern-evoked ERG (pERG) and photopic negative response (phNR). The pERG and phNR provide a direct indication of retinal ganglion cell (RGC) responsiveness.

[0215] Since rats rely more on motor function / equilibrium, hearing, smell, and vibration than vision, profound visual debilitation is required in order to detect changes upon administration of angiotensin (1-7). Electrode implantation beneath the occipital calvarium was used, which generated about 10-fold greater amplitudes than sub-scalp skin electrodes to obtain electrophysiological responses without increased background. rNAION induction rNAION was induced in Long-Evans (LE) pigmented male rats by IV injection (2.5 mg / ml, 1 ml / kg). Animals were anesthetized with Ketamine / Xylazine (80mg / 4mg / kg). Eyes were topically anesthetized with 0.5% proparacaine. Thirty seconds post-injection, the intraocular optic nerve segment (optic nerve head, ONH) was illuminated with a 500um spot size, 532nm laser, at 53mW power (measured by power meter at the point of the eye) for 11 seconds. Animals then were allowed to recover. One day post-induction, animals were re-anesthetized, spectral domain-optical coherence tomography (SD-OCT) was performed on the posterior of the eye (fundus), and optic nerve head (ONH) edema was measured by taking the mean of ONH diameter, using the three largest values (normal value = ~300um). Only animals with an average diameter >500um were included in the study. Maximal ONH edema is reached by 2 days; but the difference between 1- and 2-day edema is typically an additional 10-20um. Animals were stratified by degree of edema (500-600um, 600-700um, 700-800um). The results are shown in FIG. 1A-FIG. ID, which shows images from analysis of L-E rat optic nerve head edema, using spectral domain optical coherence tomography (SD- OCT). FIG. 1A shows an image of the retina and optic nerve of a control eye. FIG. IB shows images of measurement of the optic nerve head diameter in an uninduced mouse, where arrowheads indicate the width of a single section of 322 pm. FIG. 1C shows an image of post-rNAION optic nerve head edema showing swelling of the intraocular optic nerve. Individual cross-sections that can be measured are shown as lines. FIG. ID shows a cross-section of optic nerve head edema post-rNAION where arrowheads indicated expansion to 552 pm. Three largest contiguous sections were used to estimate mean optic nerve head diameter. No statistically significant difference was seen between uninduced and induced for optic nerve head edema (two-tailed t-test; p=0.923). Effects of Angiotensin (1-7) administration

[0216] Angiotensin (1-7) was administered at a dose of 1 mg / kg / day for 28 days. An equivalent amount of vehicle (PBS) was administered as a control. Various parameters were analyzed as described below. Animals were administered angiotensin (1-7) via a surgically implanted subcutaneous osmotic pump (Alzet 2ML4) beginning 1 day after ON ischemic induction.

[0217] (i) Retinal Ganglion Cell (RGC) survival.

[0218] Retinal ganglion cell survival was evaluated. Results comparing vehicle-treated and angiotensin(l-7) administered mice suggested that there was no increase in RGC survival in angiotensin (l-7)-treated animals (mean RGC loss= 0.32 for vehicle; 0.38 for Ang-(l-7)).

[0219] (ii) Optomotry.

[0220] Optomotry (cerebral mechanics) was performed 3-4 weeks post-induction. L-E rats have a maximum visual acuity of around 20 / 400 (1 cycle / degree), about 8 lines less than humans, whose acuity is 60 cycles / degree. Visual acuity results were stratified by ONH edema severity (500-600um=mild; 600-700um=moderate; >700um=severe). FIG. 2 shows a graph of stratification of optomotry results showing a comparison of visual acuity with degree of edema. A visual acuity of 500-600 pm was categorized as mild edema, which corresponds to the least degree of ON damage, visual acuity of 600-700 pm was categorized as moderate edema and greater than 700 pm was categorized as severe edema.

[0221] Treatment with Ang-(l-7) was associated with a small improvement in visual acuity relative to control in cases of mild edema. Relative improvement in visual acuity upon Ang-(l-7) was greater relative to control in eyes with a moderate degree of edema. The greatest improvement upon Ang-(l-7) treatment relative to control was observed in eyes with the most severe amount of edema.

[0222] Overall, the results showed that angiotensin (1-7), also referred to as TXA127 herein, was most effective in animals with the most severe ONH edema (i.e., most severe ischemia). Animals treated with Ang-(l-7) had consistently greater acuity in all three groups. Comparison with a 2-way analysis of variance (ANOVA) that incorporated both edema and optomotry values revealed that Ang-(l-7) significantly improved visual acuity after optic nerve stroke when all animals were included (p=0.0378).

[0223] (Hi) Electrophysiology.

[0224] Animals were surgically implanted with two sterile transcranial stainless steel (A286) screw electrodes (0-80 thread, 0.096 dia X 3 / 16” long; McMaster-Carr) at the occipital skull area (7mm post-bregma, 3mm+ / - midline). Electrodes were implanted after drilling through the skull without penetrating the dura. Electrodes were immobilized, first with cyanoacrylate glue, and then with dental cement. The skin was sutured over the surgical area using 3-0 nylon sutures, following which the animals were treated with ethiqua-XR (sustained release buprenorphine) and allowed to recover for 1 week. Animals were dark-adapted overnight, and visual electrophysiology performed using the Diagnosys-Celeris device, which enables near-simultaneous analysis of ERG and fVEPs after optimizing ocular LED-electrode position using ERG response at lcd / m2 illumination. Following ERG-signal maximization to both eyes (this ensures maximum retinal / VEP stimulation), VEPs were obtained bilaterally (50 flashes / eye) and averaged. The Pl-Nl amplitude was determined for each eye and for each post (right post— left and right flash; left post— left and right flash). Animals were treated in pairs (vehicle- vs ang(l-7)-treated), and double-blind results were obtained from dark- adapted, ketamine / xylazine-anesthetized animals a minimum of 30d post-induction.

[0225] FIG. 3 shows graphs of visual electrophysiology as measured by flash visual evoked potential (VEP). Animals were administered Ang-(l-7) (Img / kg / d) for 28 days beginning Id post -induction. SD-OCTs were used to evaluate degree of ONH edema and presumptive severity of ON ischemia (Mild: 500-600 gm; Moderate: 600-700 gm; Severe: >700 gm). Progressively greater retention of residual VEP function was observed in Ang-(l-7)-treated animals with a more severe ischemic lesion than in vehicle-treated animals. Results showed that the mean Pl-Nl amplitude for vehicle control was 0.51 vs 0.66 for Ang-(l-7), showing a strong trend towards improvement for Ang-(l-7) treatment (p=0.0844, 2 way ANOVA, all animals). FIG. 4 shows a graph of VEP in animals suffering moderate and severe edema (600-850 gm) treated with Ang-(l-7) relative to control animals. Although improvement was seen in every subgroup i.e. mild and moderate edema, the greatest improvement was seen in animals with the most severe edema (p=0.0368, two-tailed t-test, unpaired).

[0226] Overall, the results showed that Ang-(l-7) is functionally neuroregenerative when administered one day post-rNAION induction, indicating that Ang-(l-7) treatment ameliorates symptoms of optic neuropathy and restores visual function.

[0227] Example 2. Angiotensin (1-7) provides potent long-term neurorepair / neuro- regeneration in a rat model of ischemic optic neuropathy (rNAION: rat optic nerve white matter stroke model)

[0228] Following the initial study of Example 1, the results were confirmed with larger groups of animals and extended to investigate cellular optic nerve head (ONH) inflammation.

[0229] Methods

[0230] All experiments followed the ARRIVE 2.0 guidelines / Essential 10 for animal research.

[0231] Animals: All animal protocols experiments were approved by the institutional animal care and use committee (IACUC) and followed the recommendations of the Declaration of Helsinki. Pigmented Long-Evans (LE) outbred male rats (250-275 g; 8-10 weeks old) were used, as albino strains such as Sprague-Dawley and Wistar have both diminished visual acuity compared with normally pigmented strains (Prusky et al., Behavioural Brain Research. 2002;136:339-348) and potential optic chiasm decussation anomalies. Animals used for osmotic pump implantation and cranial surgery were treated with both subcutaneous and sustained release buprenorphine (Ethiqa XR). rNAION induction: The induction procedure was previously described (Bernstein et al., Invest Ophthalmol Vis Sci. 2003;44:4153-4162). Briefly, animals were anesthetized with ketamine-xylazine and the pupils dilated with 1% tropicamide and topically anesthetized with 0.5% proparacaine. A custom-made contact lens (Micro-R; Nissel and Cantor, UK) was placed on the eye. Animal were injected intravenously with rose Bengal dye dissolved in normal saline (2.5mM; Iml / kg). Thirty seconds later, the intraocular portion of the optic nerve was illuminated with a 532nm laser light (Oculite GL; Iridex, Mountain View, CA) at 50mW power, 500um spot size, for 11 seconds. Optic nerve head imaging-optic nerve edema analysis: One day post-rNAION induction, animals were re-anesthetized with ketamine-xylazine, their pupils dilated, and their corneas topically anesthetized. Spectral domain-optical coherence tomography (OCT; Heidelberg-Spectralis; Heidelberg, Germany) with a 28-diopter correcting rodent lens was used to image the intraocular optic nerve and to assess the severity of ONH edema. Mean ONH edema was based on the mean cross-sectional diameter of three contiguous central scans, as previously described (Guo et a\., Pl.OS ONE. 2021 ;16:e0243186). The same Microt custom-made contact lens was used to generate the cross-sectional scans. Results were compared with mean maximum diameters of the contralateral (uninduced eye) obtained from each animal. Animals with mean ONH edema <450 pm (about 20% of induced animals) were eliminated from the study prior to randomization and treatment based on previous studies by Guo et al. that showed 2-day ONH edema >500 pm consistently resulted in RGC loss whereas less edema resulted in extremely variable RGC loss (Guo et al., PLOS ONE. 2021 ;16:e0243186).

[0232] ANG(l-7) and vehicle (control) treatments: A power analysis was utilized to predict the number of animals needed, where the expected response difference was 15 + / - 15% (sd) between the induced / control and induced / treated groups. Previous experiments have revealed an -55-60% mean RGC loss with the induction conditions used. Power analysis suggested a minimum of 16 animals / group, with an alpha of 0.05 and power of 80%. Anesthetized threshold animals with > 450 pm mean ONH diameter Id post-induction were immediately used for osmotic pump implantation. Animals were paired for equivalent edema levels, and randomized: one animal from each pair was used for either ANG(l-7) or vehicle implant group. Post-imaging, anesthetized animals were administered subcutaneously an initial loading dose of either ANG(l-7) (1 mg / kg SC) or equivalent volume of vehicle (sterile pH 7.4 PBS), then prepped aseptically for surgery using 10% povidone scrub followed by 10% povidone solution, alcohol rinsed, and dried. A midline incision between the shoulder blades was made, and an Alzet pump 2ML4 (Durect Corp, Cupertino CA) was inserted subcutaneously with additional local anesthesia (1% lidocaine). 2ML4 pumps deliver 2.5 pl / hr for 28 days, to yield 350ug / day per animal. Animal IDs were then masked to the individual responsible for electrode implantation and testing (JW). OptoMotry-based visual acuity assessment: Beginning 22 days post-induction, visual acuity (VA) was measured in unsedated animals using a virtual optokinetic system (OptoMotry: CerebralMechanics, Toronto, Canada), using a rat pedestal (Douglas et al., Vis Neurosci. 2005;22:677-684). Rats were placed on a stable elevated platform in the center of a square surrounded by four monitors that projected a continuously moving sine-wave black- and-white grating. Assessed was the the ability of a rat to resolve a given spatial frequency by identifying when the animal turns its head at a constant speed in either clockwise (left eye) or counterclockwise (right eye) direction. The VA of both eyes of each animal were evaluated by three independent trials, on separate days, with each eyes estimated maximum VA elicited as a single measure using the staircase parameter, with a starting step size of 0.050 cycles / degree, based on the maximum reversal number (8) from any individual trial, and a column speed of 20. After each reversal, step size was decreased by half. Three daily trials were performed on each animal, beginning on day 21 post-induction, and the result with the greatest level acuity was considered maximal acuity. Because a rare animal can refuse voluntary acuity tests, animals that met ONH edema requirements but failed acuity in the uninduced eye were used for fVEP analysis, but were not included in acuity grouping.

[0233] Transcranial electrode implantation: Bilateral screw electrodes were implanted over the occipital prominence a day post-OptoMotry, which occurred at around 25 days after rNAION induction. Animals were anesthetized using ketamine / xylazine, then topically anesthetized subcutaneously using 1% lidocaine, with buprenorphine analgesia. The animal then was placed into a stereotactic instrument on a heating pad. The skull skin was incised at the midline and retracted, following which bilateral skull burr holes were made at stereotactic locations of VI visual cortex using published parameters (Bennion et al., Exp Physiol. 2018;103:916-923) using a motorized surgical drill with care not to penetrate the dura. Two stainless steel pan-head screws (5 / 16” length; catalog #11197; Minitaps, Seattle WA) were embedded, one over each visual cortex, and fixed in place with dental cement. The skin then was closed using stainless steel sutures. Following long-term analgesia administration (Ethiqua-XR), animals were placed on a warming pad until recovery from anesthesia and allowed to recover from surgery for 3 days prior to testing.

[0234] Visual Evoked potentials: At 28-30d post-induction, animals were dark adapted overnight and then re-anesthetized using inhaled isoflurane followed by ketamine-xyl azine in dim red light. Pupils were dilated using 1% cyclopentolate and 2.5% neosynephrine. Animals were placed on the prewarmed Celeris electrophysiology testing platform (Diagnosys LLC: Lowell, MA). Combined corneal electrode and light emitting diode stimulators were placed on the eyes using Systane gel drops (Alcon, Ft Worth, TX). Alligator microclips were hooked onto the two skull electrodes. Fifty simultaneous electroretinography (ERG) / fVEP flashes from each eye were averaged, and visual function of the rNAION-induced eye compared with the contralateral healthy eye was calculated from fVEP results. fVEP measurements were repeated twice for each eye within the same session. After the procedure, the eyes of each animal were covered with ophthalmic triple antibiotic ointment with dexamethasone, and the animal was placed in a warm cage and allowed to recover from anesthesia. ERG results were used to identify and exclude rats with possible retinal ischemia. Rats were excluded if the maximum ERGb-wave amplitude of the rNAION induced eye was less than 50% of that of the contralateral non-induced eye.

[0235] Tissue isolation: Following all testing, animals were euthanized using CO2 inhalation and then decapitated. Eyes and optic nerves were rapidly isolated, and the corneas were incised with a 26ga needle and placed in 4% paraformaldehyde-phosphate buffered saline pH 7.4 (PFA-PBS). Tissues were post-fixed overnight, and the retina then was dissected and removed for whole mount immunohistology. The ONH along with 1.5mm of the adjacent anterior optic nerve was dissected from the sclera, and a more distal portion of the optic nerve also was removed. Both were then cryopreserved in 30% sucrose-PBS, embedded in OCT, and flash frozen at -80°C. ONH and optic nerves were cross-sectioned at 10pm thickness and stored at -20°C until use.

[0236] Immunohistochemistry: Whole retinae were immunoreacted using polyclonal rabbit anti-Brn3a (Pou4fl) (Cat # 411003, Synaptic systems, Goettingen Germany) at 1 : 1000 dilution. Retinae were immunostained using Cy3-labeled donkey anti-rabbit polyclonal antibody 1 :500 (Affinipure, Cat 711-165-152; Jackson Immunoresearch, West Grove, PA). Optic nerve sections were evaluated using IBA1 (for inflammatory cells), CD206 (for M2 macrophage type immune response), iNOS (for Ml immune response), CD68 (for extrinsic macrophages, and GFAP (for ON glial scarring). Following primary antibody binding and wash, sections were reacted with the appropriate fluorescent-labeled secondary antibodies (Jackson Immunoresearch). Confocal microscopy was performed using a Leica 300 confocal microscope, and standard fluorescent microscopy was performed using a Keyence BZX-710 computerized microscope.

[0237] Statistical Calculations: Differences between ONH edema, OptoMotry-based visual acuity, fVEP based visual function, and RGC survival between the ANG(l-7) and vehicle- treated groups were compared with unpaired student’s t-test. It should be noted that not all animals were used for all functionality tests. For example, while optic nerve edema values could be obtained for all animals (50 animals included), optomotry values could not be obtained for a number of animals in each group, due to an animal’s refusal of acuity testing even through multiple trial attempts (n=3). This was confirmed by acuity testing of the contralateral (uninduced) eye. Thus, the total number of animals used for optomotry testing was 22 (vehicle) and 21 (treated), within the entire group. While the entire group received subcranial electrode implantation, a small number of implants failed, resulting in complete fVEP results on 18 vehicle- and 22-treated animals.

[0238] Additionally, analysis of covariance (ANCOVA) was used to compare the three dependent variables - visual acuity / optomotry, fVEP -based visual function, and RGC survival - with treatment status, ANG(l-7) vs vehicle, as a categorical independent variable, and ONH edema as a covariate. Python and GraphPad Prism (GraphPad Software; Boston, MA) were used to perform statistical analysis. Values are presented as mean±standard error of the mean (SEM) unless otherwise specified.

[0239] Results

[0240] A. Optic nerve edema following rNAION and randomization.

[0241] Mean ONH edema post-rNAION for randomized vehicle and treated groups are shown in Fig. 5. Edema values were 660±22.9 pm (N=26) for vehicle and 649±26 pm (N=24) for treated animals, respectively (p=0.735). This difference was due to elimination of two animals on determination by one of us (SLB) of suboptimal edema measurement on retrospective analysis. The mean ONH diameter in the uninduced (contralateral) eye of animals was 323±5.8 pm (n=16 animals). B. ANG(l-7) improves visual acuity following rNAION : OptoMotry

[0242] Visual acuity in the rNAION-induced eye was compared with the uninduced eye as a ratio of visual acuity measured in cycles per degree (c / d) in the rNAION induced eye to the visual acuity in the control eye. Overall results for both vehicle and ANGfl-7) treated animals are shown in Fig. 6A. ANCOVA analysis showed that ANG(l-7)-treated animals performed significantly better in the rNAION-induced eye compared with the vehicle-treated animals with ONH edema as a covariate [F(l, 40) = 7.68, p = 0.0084],

[0243] ANCOVA assumptions testing was performed to evaluate both homogeneity of variance, and distribution type. A Levene’s test revealed homogeneity of variance between the two groups with [F(l, 44) = 1.51, p = 0.225], A Shapiro Wilk test also was performed and showed that residuals between the observed and modeled values were normally distributed [p = 0.507],

[0244] An unpaired, two tailed t-testwas performed for all OptoMotry data. This revealed that ANG(l-7) treatment resulted in a mean 10% overall group improvement, compared with vehicle [t(44)=1.994, p=0.0523]. In animals with either moderate (500-599 um) or severe ONH edema (>600 um), the increase in VA compared with vehicle- treated animals with similar amounts of edema was even more profound (Fig. 6B), with an increase of -0.20% [t(25)=2.753, p=0.0108]. Thus, ANG(l-7) treatment improves VA in rNAION when administered 1 day post-rNAION induction. rNAION-affected eyes were segregated into the relative severity of ONH edema (mild: < 600pm; moderate: >600pm-699pm; severe: 700pm-850pm) and compared the effects of ANG(l-7) for each subgroup against the equivalent vehicle treatment subgroup (Fig. 6C). ANG(l-7)’s relative treatment efficacy improved the greater the severity of ONH edema, with 3.6% improvement in the mild group, 13% improvement in the moderate group, and 18% improvement in the severe group. Relative efficacy in improving VA was greatest for eyes with severe ONH edema.

[0245] C. ANG(I-7) improves visual function after rNAION: jVEP amplitude analysis fVEP is widely used to evaluate changes in visual function following various conditions. ANCOVA assumptions testing was performed to evaluate both homogeneity of variance and distribution type. A Levene’s test revealed homogeneity of variance between the two groups with [F(l, 41) = 0.233, p = 0.632] . A Shapiro Wilk test also was performed and showed that residuals between the observed and modeled values were normally distributed [p = 0.991],

[0246] ANCOVA analysis of fVEP-based visual function revealed that ANG(l-7)-treated animals demonstrated improved visual function with respect to waveform amplitudes measured at the visual cortex, compared with vehicle-treated animals (Fig. 7A) [F(l, 37) = 4.64, p = 0.0378], ANG(l-7) treatment therefore resulted not only in significant improvement of visual acuity but also in significant improvement of the fVEP in an overall significant improvement in visual function. fVEP improvement was compared using a two tailed unpaired t-test for VEP data from animals with ONH edema >600um (Fig. 7B). This revealed that ANG(l-7) treatment in animals with moderate-to-severe edema resulted in a 20% improvement in amplitudes compared with vehicle treatment [t(24)=2.703, p=0.0124]. Overall changes in fVEP amplitude vs severity of ONH edema were stratified into comparative fVEP results from animals with mild, moderate, and severe ONH edema (Fig. 7C), with 8% improvement in mild group, 21% improvement in moderate group, and 19% improvement in the severe group. ANG(l-7)’s effect was found to be greatest in eyes with the most severe disease.

[0247] D. ANG(l-7) effects are not explained by increasing RGC survival

[0248] Previous studies using RGC neuroprotectives have not been found to improve overall RGC survival when administered 1 day or later after the insult. Neuroregeneration is not based on neuroprotection and not hypothesized to be directly responsible for overall functional change. Nevertheless, RGC stereology was performed on a randomized subset of eyes in both groups, with sufficient power for analysis. Fig. 8 shows overall RGC comparisons. Mean RGC loss for vehicle-treated animals was 74.29±5.39 % (n=17 randomly selected animals), whereas mean RGC loss for ANG(l-7)-treated animals was 63.09±7.17 % (n=14 animals). This difference was not significant [t(29)=1.257, p=0.219]

[0249] E. ANG(l-7) treatment reduces cellular ONH inflammation after rNAION induction.

[0250] It was previously determined that early cellular invasion (extrinsic macrophages) occurs within 3 days post-rNAION induction. By 30d post-induction, the majority of inflammatory cells in the ONH presumably represent microglia. Immunohistochemical analysis was performed on a subgroup of vehicle- and ANG(l-7)-treated animals with similar ONH edema values, 30d post-induction, using ED1+ (activated macrophages and microglia) and IBA1(+) (macrophage / microglia specific) in more than 30d post-rNAION induced ONHs (n=5 / group). Analysis revealed that ONHs from ANG(l-7)-treated animals had lower IBA1(+) and ED1(+) signal than vehicle-animals, but this was not significant (Fig. 9: Naive ON is shown in A; Compare with D (ANG(l-7) treated) and C (vehicle treated) and graph in B). IBA1 densitometric analysis indicated that ONHs from ANG(l-7)-treated animals had signal intensity of 8.54±1.0 % vs 10.8±0.7 % for vehicle-treated animals. This suggests that ANG(l-7) treatment suppresses post-ON infarct cellular inflammation, even when animals have similar amounts of RGC loss.

[0251] Conclusions

[0252] In the rNAION-white matter stroke model, ANG(l-7) (TXA127) represents the first compound that is functionally effective when administered at least 1 day after axon ischemic induction. Until now, NAION clinical treatment trials employing a double-blind treatment approach have not demonstrated a statistically successful outcome in improving visual function. NAION treatment trials incorporate individuals who typically present at least a day after onset. Similarly, previous reports using the rNAION model have failed to identify effective treatments when treatment is delayed more than 1 day post-induction.

[0253] This study showed overall improvement in visual function as assessed by OptoMotry as well as IVEP amplitudes in ANG(l-7)-treated rats compared with vehicle-treated rats across all edema levels. As occurs in patients with spontaneous NAION, rNAION-affected animals exhibit varying levels of ONH edema, with the animals with the greatest severity in ONH edema having poorer visual function. Analysis of the primate NAION (pNAION) model also has revealed a close correlation between the severity of ONH edema and the severity of visual function loss as assessed by fVEP amplitudes. In the rNAION model, the severity of ONH edema correlates with the severity of the rNAION lesion. These differences in rNAION-induced ONH edema were used to segregate animals into mild (>450pm- 599pm), moderate (600pm-699pm) and severe (700pm-850pm) groups. ANG(l-7) exhibited its greatest relative neuroregenerative / neuroreparative effects in animals with the most severe ONH edema (see Fig. 7 and Fig. 8), with lesser relative improvement effects on animals with mild edema.

[0254] This effect is not explained entirely by differences in either RGC survival (10% improvement in RGC survival in ANG(l-7) vs vehicle) or post-rNAION ON inflammation, as measured by Iba-1 (~2%). This suggests that improved visual function associated with ANG(l-7) treatment may be due to either a combination of factors or unmeasured factors, such as enhanced function in residual (post-rNAION) neurons, changes in ONH scarring, alteration in inflammatory responses (Ml-neurodegenerative to M2-neuroprotective, or changes in other cell responses resulting in increased ON efficiency. Treatment with ANG(l-7) and other Angl-7 type agonists represents a new approach to effective clinical treatment of NAION and other forms of white matter stroke. The direct translation of this agent into the clinics is also supported by ANG(l-7)’s already proven margin of safety in various clinical trials. Even though effects in the ‘mild group’ were less pronounced than in the other treatment groups and the effect in the ‘moderate group’ was less pronounced than in the ‘severe group’, the treatment of mild and moderate forms of optic neuropathy is extremely useful as it would prevent or at least slow down progression to more severe forms of optic neuropathy (e.g. prevent or ameliorate further or stronger symptoms), or at the very least treat the severe form as soon as it has developed from the mild / moderate form.

[0255] EQUIVALENTS AND SCOPE

[0256] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. The scope of the present invention is not intended to be limited to the above Description, but rather is as set forth in the following claims:

Claims

CLAIMS1. A method of treating optic neuropathy comprising administering an angiotensin- (1-7) receptor agonist to a subject suffering from optic neuropathy.

2. A method of prophylactically treating optic neuropathy comprising administering an angiotensin-(l-7) receptor agonist to a subject, preferably a subject at increased risk for optic neuropathy.

3. The method of claim 1 or 2, wherein the administration is parenteral, rectal, intranasal, oral, or a combination thereof, wherein the parenteral administration is intravenous, subcutaneous, inhalation, intradermal, transdermal, and / or transmucosal administration.

4. The method of any one of claims 1-3, wherein the optic neuropathy is an ischemic optic neuropathy, preferably selected from the group consisting of anterior ischemic optic neuropathy (AION), or posterior ischemic optic neuropathy (PION), and combinations thereof.

5. The method of claim 4, wherein the anterior ischemic optic neuropathy is arteritic anterior ischemic optic neuropathy (A-AION), or non-arteritic anterior ischemic neuropathy (NA-AION).

6. The method of claim 5, wherein the arteritic anterior ischemic optic neuropathy is associated with autoimmune inflammation and / or giant cell arteritis; and / or wherein the posterior ischemic optic neuropathy is arteritic posterior ischemic optic neuropathy (A- PION), non-arteritic posterior ischemic optic neuropathy (NA-PION), trauma-related posterior ischemic optic neuropathy (T-PION) or perioperative posterior ischemic optic neuropathy (P-PION).

7. The method of any one of claims 1-6, wherein the angiotensin (1-7) receptor agonist is administered at an effective dose periodically at an administration interval such that at least one symptom or feature of optic neuropathy is reduced in intensity, severity, duration, or frequency or has delayed onset.

8. The method of claim 7, wherein the one or more symptoms of optic neuropathy is selected from vision loss or impairment characterized by blurring, blind spots, shadows obscuring upper or lower visual fields, impaired visual contrast, light sensitivity, decreased color vision, increased intraocular pressure, and combinations thereof, preferably wherein the vision loss or impairment is bilateral.

9. The method of any one of claims 1-8, wherein the optic neuropathy is associated with one or more of high blood pressure, anemia, diabetes mellitus, high cholesterol, cardiac disease, arteriosclerosis, vasculitis, liver disease, kidney disease, erectile dysfunction, GLP- 1 agonist therapy and combinations thereof.

10. The method of any one of claims 1-9, wherein the angiotensin (1-7) receptor agonist is an angiotensin (1-7) peptide.

11. The method of claim 10, wherein the angiotensin (1-7) peptide is angiotensin (1- 7) having the amino acid sequence of Asp1-Arg2-Val3-Tyr4-Ile5- His6-Pro7(SEQ ID NO: 1), a functional variant and / or a precursor thereof.

12. The method of claims 11, wherein the angiotensin (1-7) peptide has the amino acid sequence of Asp1- Arg2-Val3-Ser4-Ile5-His6-Cys7(SEQ ID NO: 2) or of Ala1- Arg2-Val3- Ser4-Ile5-His6-Cys7(SEQ ID NO: 3).

13. The method of any one of claims 1-12, wherein the angiotensin (1-7) peptide comprises one or more chemical modifications to increase protease resistance, serum stability and / or bioavailability, preferably wherein the one or more chemical modifications comprise pegylation.

14. The method of any one of claims 1-9, wherein the angiotensin (1-7) receptor agonist is a non-peptidic angiotensin(l-7) receptor agonist.

15. The method of claim 15, wherein the non-peptidic angiotensin(l-7) receptor agonist is a compound with the following structure:or a pharmaceutically acceptable salt thereof.

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