Monomeric annexin A5 for use in the treatment of macular edema or retinal vein occlusion

Abstract

The present invention relates to a novel method, use, and composition of a molecule consisting of monomeric annexin A5 protein for the prevention or treatment of macular edema and one or more diseases, disorders, or conditions associated with and / or caused by macular edema. In a preferred embodiment, annexin A5 is administered systemically, for example, by intravenous administration.

Description

Technical Field

[0001] The present invention relates to novel methods, uses, and compositions for the prevention or treatment of macular edema and related conditions.

Background Art

[0002] Macular edema is a pathological accumulation of fluid at the center of the retina. This is a complication of many retinal diseases, including, inter alia, diabetic retinopathy (DR), retinal vascular occlusion (RVO), and uveitis (UV) (Haydinger et al, 2023, Frontiers in Medicine, 10:1128811). Macular edema causes a decrease in visual acuity and, if chronic or refractory, can lead to severe permanent visual impairment and blindness. In combination with the underlying pathology, the edema can progress to cause irreversible tissue damage with death of retinal cells and permanent visual impairment.

[0003] In most cases, macular edema develops due to dysregulation of the blood-retinal barrier (BRB), a series of structures that tightly regulate the passage of proteins, salts, metabolites, and other solutes between the blood and retinal tissue in a healthy state. Dysregulation of the BRB allows infiltration of the retinal tissue by proteins and other solutes that are normally retained in the blood (Haydinger et al, 2023, (supra)). An increase in osmotic pressure in the tissue drives fluid accumulation.

[0004] Current treatments target blood-retinal barrier (BRB) dysfunction and include vascular endothelial growth factor (VEGF) inhibitors, corticosteroids, and nonsteroidal anti-inflammatory drugs (Haydinger et al, 2023 (supra), Ip & Hendrick, 2018, Asia-Pacific Journal of Ophthalmology, 7(1):40-45). These treatments target vasoactive and inflammatory mediators that cause disruption of the blood-retinal barrier. Other treatments include targeted laser photocoagulation (Haydinger et al, 2023 (supra), Ip & Hendrick, 2018 (supra)).

[0005] The first-line treatment for macular edema associated with diabetic retinopathy (DR) or retinal vein occlusion (RVO), and the treatment of RVO itself, is anti-VEGF therapy, which may be enhanced with intravitreal corticosteroids and / or retinal laser photocoagulation (Ehlers et al. 2022, Ophthalmology, 129:88-99, Ehlers et al, 2017, Ophthalmology, 124:1412-23, and Yeh et al, 2015, Ophthalmology, 122:769-78). Treatment of macular edema secondary to neovascular AMD also relies heavily on the use of anti-VEGF therapy (Flaxel et al, 2020, Ophthalmology, 127:P1-P65).

[0006] The established standard treatment for macular edema and / or RVO, and indeed for multiple retinal diseases, is by direct delivery of therapeutic agents to the eye via intravitreal injection (Haydinger et al, 2023 (supra), Ip & Hendrick, 2018 (supra), Cox et al., 2021, J.Clin.Med., 10(5):981). Since the retina is located at the back of the eye and is surrounded by multiple protective layers including the cornea, iris, and lens, such drugs are injected directly into the eye. These layers can make it difficult for the drugs to reach the retina if they are administered by other routes. When drugs are injected into the eye, they are delivered directly to the retina and can act locally to target the cause of the problem. For example, anti-VEGF drugs are injected into the eye to block the action of VEGF-A, a protein that promotes abnormal blood vessel growth in the retina. By reducing the level of VEGF-A, these drugs can help prevent the formation of new blood vessels and reduce eye inflammation and swelling. Injections into the eye are typically performed in an ophthalmologist's examination room or outpatient surgical center. After the injection, patients are typically monitored for a short period to ensure there are no adverse effects from the drug.

[0007] Targeting VEGF by intravitreal injection of monoclonal antibodies or high-affinity decoy receptors is an effective means for the treatment of most diseases with macular edema, including exudative age-related macular degeneration (Heier et al., 2012, Ophthalmology, 119:2537-48), diabetic macular edema (Nguyen et al., 2012, Ophthalmology, 119:789-801, Korobelnik et al, 2014, Ophthalmology, 121:2247-54), and macular edema caused by central retinal vein occlusion (CRVO) (Brown et al., 2010, Ophthalmology, 117:1124-33), branch retinal vein occlusion (BRVO) (Campochiaro et al., 2010, Ophthalmology, 117:1102-12), and uveitis (Mackensen et al., 2008, Retina, 28:41-5).

[0008] However, there are concerns about ocular and systemic toxicity with anti-VEGF therapy, and long-term frequent reinjections are required (Nguyen et al., 2010, Ophthalmology, 117:2146-51, Campochiaro et al., 2010, Ophthalmology, 117:2387-94, Saint-Geniez et al, 2008, PLoS One., 3:e3554). Potential complications associated with intravitreal injection of anti-VEGF therapy include infectious endophthalmitis, sterile intraocular inflammation, and particularly retinal vasculitis due to brolucizumab (Cox et al., 2021 (aforementioned)), and transient intraocular pressure (IOP) spikes (Levin et al., 2021, J Glaucoma, 30:1019-26).

[0009] Infectious endophthalmitis remains one of the most serious complications associated with anti-VEGF therapy. The reported incidence after injection varies from 0.008% to 0.092%, and despite its low incidence, high-volume repeated anti-VEGF injections mean that they account for a large proportion of cases of infectious endophthalmitis (Cox et al., 2021 (supra)).

[0010] Sterile Intraocular Inflammation (SII), also known as pseudophthalmitis, is characterized by acute-onset intraocular inflammation without infection that resolves without antibiotic treatment. The reported incidence of SII associated with anti-VEGF varies from 0.02% to 0.37% depending on the study, and symptoms usually occur between 24 hours and 7 days after the stimulating injection. Symptoms can consist of blurred vision, floaters, pain, and photophobia. Pain is present in up to 46% of patients and its presence is significantly associated with severe vitreous or anterior chamber (AC) inflammation. Photophobia occurs in 4 - 19% of patients. In addition, visual acuity drops significantly from its baseline at the time of symptoms but often returns to pre-injection levels after the inflammation has resolved. Intraocular inflammation is present at the time of examination in virtually all cases. This can consist of vitritis, AC reaction, or most commonly both, and additional less common findings include hypopyon, fibrin, corneal deposits, corneal edema, conjunctival injection, and conjunctival edema (Cox et al., 2021 (supra)).

[0011] Continuous anti-VEGF therapy by intravitreal injection is the current standard of care in the treatment of macular edema (Ip & Hendrick, 2018 (supra)). In fact, the indications for intravitreal injection are expanding, and it is recognized that the number of patients who are likely to be affected by the acute and chronic effects of intravitreal injection is increasing (Levin et al., 2021 (supra)). However, current treatments for macular edema or RVO have not demonstrated a reliable method to directly improve perfusion. Instead, current strategies focus on minimizing the impact from the consequences of RVO and minimizing vision loss due to macular edema and neovascularization (Ip & Hendrick, 2018 (supra)).

[0012] Against this background, there is a need in the art for improved therapies for macular edema and related conditions, including RVO. In particular, there is a need for effective therapies that can be effectively administered to a subject without the need for direct injection (especially multiple repeated injections) into the eye of the subject. SUMMARY OF THE INVENTION

[0013] Surprisingly, against this background, it has been determined that monomeric annexin A5 protein can preferentially localize to the occlusion site in a subject having retinal vein occlusion (RVO) when systemically administered to the subject (e.g., by intravenous administration). Further, annexin A5 has also been determined to provide a biological activity that modulates factors contributing to breakdown of the blood-retinal barrier (BRB) in a subject having macular edema and / or RVO, as well as factors contributing to other therapeutic benefits in a subject having macular edema and / or RVO.

[0014] Accordingly, in a first aspect, there is provided a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of macular oedema or a related condition in a subject.

[0015] In an alternative embodiment of the first aspect, the present invention provides a method for treating or preventing macular edema or a condition associated therewith in a subject, the method comprising administering to the subject a molecule consisting of a therapeutically effective amount of monomeric annexin A5 protein.

[0016] In an alternative embodiment of the first aspect, the present invention provides the use of a molecule consisting of monomeric annexin A5 protein in the manufacture of a medicament for the prevention or treatment of macular edema or a symptom associated therewith in a subject.

[0017] Preferably, according to the first aspect of the present invention, the molecule consisting of monomeric annexin A5 protein is administered systemically to the subject, preferably by intravenous, subcutaneous, or intramuscular administration. When it is administered intravenously, the molecule consisting of monomeric annexin A5 protein, or a composition containing the molecule, can be used in the form of a sterile aqueous solution that may contain, for example, other substances, such as salts or glucose sufficient to make the solution isotonic with blood. The aqueous solution should be suitably buffered as necessary (preferably to a pH of 3 to 9). For example, optionally, the aqueous solution is buffered at a pH within a range selected from about pH 4 to about pH 8, about pH 5 to about pH 8, about pH 6 to about pH 8, about pH 6.5 to about pH 7.9, about pH 6.8 to about pH 7.7, about pH 7.0 to about pH 7.5, about pH 7.1 to about pH 7.4, about pH 7.1 to about pH 7.4, and optionally, about pH 7.1, 7.2, 7.3, or 7.4, most preferably about pH 7.2.

[0018] Preferably, according to the first aspect of the present invention, the dosage of the molecule consisting of monomeric annexin A5 protein is administered to the subject continuously for 2 days or more, for example, continuously for 3, 4, 5, 6, 7 days or more, and / or at least 2, 3, 4, 5, 6, or 7 days within a period of every 7 days (1 week), over a period of at least 1, 2, 3, 4 weeks or more.

[0019] Optionally, according to the first aspect of the present invention, the daily dose of the molecule consisting of monomeric annexin A5 protein administered to a subject is a) a total daily dose of about 5 mg, about 4 mg, about 3 mg, about 2 mg, or less than about 2 mg, 0.1 - 1.5 mg, or about 1 mg, and / or b) within a range selected from the group consisting of about 50 - 100 μg, about 40 - 80 μg, about 30 - 60 μg, about 20 - 40 μg, 1 μg - 30 μg, or less than about 20 μg per kg of the subject's body weight. As used herein, the term "about" may be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0020] In a second aspect of the present invention, there is provided a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of retinal vein occlusion (RVO) or a condition associated therewith in a subject, wherein the dose of the molecule consisting of monomeric annexin A5 protein is systemically administered to the subject and / or the molecule consisting of monomeric annexin A5 protein is administered to the subject daily for 2 or more consecutive days.

[0021] In an alternative embodiment of the second aspect, the present invention provides a method for the treatment or prevention of RVO or a condition associated therewith in a subject, the method comprising the step of administering to the subject a therapeutically effective dose of a molecule consisting of monomeric annexin A5 protein, wherein the dose is systemically administered to the subject and / or the dose is administered to the subject daily for 2 or more consecutive days.

[0022] In an alternative embodiment of the second aspect, the present invention provides the use of a molecule consisting of monomeric annexin A5 protein in the manufacture of a medicament for the prevention or treatment of RVO or a condition associated therewith in a subject, wherein the dose of the molecule consisting of monomeric annexin A5 protein is systemically administered to the subject and / or the molecule consisting of monomeric annexin A5 protein is administered to the subject daily for 2 or more consecutive days.

[0023] Preferably, according to the second aspect of the present invention, the mode of systemic administration of the molecule consisting of monomeric annexin A5 protein is selected from intravenous, subcutaneous, or intramuscular administration. When it is administered intravenously, the molecule consisting of monomeric annexin A5 protein, or a composition containing the molecule, can be used in the form of a sterile aqueous solution that may contain, for example, other substances such as salts or glucose sufficient to make the solution isotonic with blood. The aqueous solution should be suitably buffered as necessary (preferably to a pH of 3 to 9). For example, optionally, the aqueous solution has a pH selected from within the range of about pH 4 to about pH 8, about pH 5 to about pH 8, about pH 6 to about pH 8, about pH 6.5 to about pH 7.9, about pH 6.8 to about pH 7.7, about pH 7.0 to about pH 7.5, about pH 7.1 to about pH 7.4, about pH 7.1 to about pH 7.4, and optionally, about pH 7.1, 7.2, 7.3, or 7.4, most preferably about pH 7.2.

[0024] More preferably, the dose of the molecule consisting of monomeric annexin A5 protein is systemically administered to the subject daily (e.g., by intravenous, subcutaneous, or intramuscular administration, etc.) for 2 or more consecutive days.

[0025] In an embodiment of the second aspect of the present invention, the molecule consisting of monomeric annexin A5 protein is administered to the subject daily for 3, 4, 5, 6, 7 or more consecutive days and / or at least 2, 3, 4, 5, 6, or 7 days within a period of every 7 days (1 week), over a period of at least 1, 2, 3, 4 weeks or more.

[0026] Optionally, according to a second aspect of the invention, the daily dose of the molecule consisting of monomeric annexin A5 protein administered to a subject is a) a total daily dose of about 5 mg, about 4 mg, about 3 mg, about 2 mg, or less than about 2 mg, 0.1 - 1.5 mg, or about 1 mg, and / or b) within a range selected from the group consisting of about 50 - 100 μg, about 40 - 80 μg, about 30 - 60 μg, about 20 - 40 μg, 1 μg - 30 μg, or less than about 20 μg per kg of the subject's body weight. As used herein, the term "about" may be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0027] The inventors have also determined that the treatment and prevention of retinal vein occlusion and / or one or more conditions associated therewith can be achieved using a molecule consisting of monomeric annexin A5 protein at a significantly lower dose.

[0028] For example, a subject may be administered a molecule consisting of monomeric annexin A5 protein at a dose of about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg, and / or a subject may be administered the molecule at a dose of about 0.001 mg to about 1 mg, or 1 μg to about 100 μg. As used herein, the term "about" may be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0029] Optionally, according to a second aspect of the present invention, a molecule consisting of monomeric annexin A5 protein is for use in the prevention or treatment of a condition in a subject having, suspected of having, or at risk of having retinal vein occlusion (RVO), said condition being selected from the group consisting of macular edema, retinal ischemia, iris neovascularization (optionally iris neovascularization characterized by an increase in cytokines such as an increase in VEGF), retinal neovascularization (optionally retinal neovascularization characterized by an increase in cytokines such as an increase in VEGF), neovascular glaucoma, vitreous hemorrhage, rubeosis, and retinal detachment.

[0030] Optionally, according to the first and / or second aspects of the present invention, the retinal vein occlusion can be selected, for example, from central retinal vein occlusion (CRVO), branch retinal vein occlusion (BRVO), or hemi-retinal vein occlusion (HRVO).

[0031] Optionally, according to the first and / or second aspects of the present invention, the retinal vein occlusion can optionally be an acute retinal vein occlusion, for example, acute central retinal vein occlusion (CRVO), acute branch retinal vein occlusion (BRVO), or acute hemi-retinal vein occlusion (HRVO).

[0032] Optionally, according to an embodiment of the first and / or second aspects of the present invention, the use or method can be (alternatively) for the prevention or treatment of one or more diseases, disorders, or conditions associated with macular edema and / or retinal vein occlusion and / or caused (directly or indirectly) by macular edema and / or macular edema.

[0033] Optionally, according to the first and / or second aspects of the present invention, the subject can be characterized by having from about 1.0% to about 2% of annexin A5-positive red blood cells.

[0034] Optionally, according to the first and / or second aspects of the present invention, the subject may be characterized by having an average number of about 400 to about 1000 monomeric annexin A5 binding sites per red blood cell.

[0035] Optionally, according to the first and / or second aspects of the present invention, the subject is typically a human subject, such as a human male and / or a human female. The subject can be an adult or, less typically, a juvenile, child, or neonate.

[0036] Optionally, according to the first and / or second aspects of the present invention, the subject is a human adult and / or has a body weight of about 50 kg to about 150 kg. As used herein, the term "about" can be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0037] Optionally, according to the first and / or second aspects of the present invention, the subject may additionally or alternatively be characterized by one or more of the following: a. The subject is about 50 years of age or older. b. The subject does not have, or is not suspected of having, a cardiovascular disease (e.g., myocardial infarction, stroke, hypertension, hyperlipidemia, hyperhomocysteinemia, blood coagulation disorders, and / or peripheral arterial disease, and optionally excluding atherosclerosis), or a disease associated with a cardiovascular disease (e.g., sickle cell disease, glaucoma, diabetes, diabetes mellitus, polycythemia vera, and / or a systemic inflammatory disorder (e.g., Behcets disease, polyarteritis nodosa, sarcoidosis, Wegener's granulomatosis, and / or Goodpasture syndrome), but preferably excluding diabetes and diabetes mellitus). c. The subject has no medical history of cardiovascular diseases (such as myocardial infarction, stroke, hypertension, hyperlipidemia, hyperhomocysteinemia, blood coagulation disorders, and / or peripheral arterial diseases, and optionally excluding atherosclerosis), or diseases associated with cardiovascular diseases (such as sickle cell disease, glaucoma, diabetes, diabetes mellitus, polycythemia vera, and / or systemic inflammatory disorders (such as Behcets disease, polyarteritis nodosa, sarcoidosis, Wegener granulomatosis, and / or Goodpasture syndrome), but preferably excluding diabetes and diabetes mellitus). d. The subject has no family history of cardiovascular diseases (such as myocardial infarction, stroke, atherosclerosis, hypertension, hyperlipidemia, hyperhomocysteinemia, blood coagulation disorders, and / or peripheral arterial diseases, and optionally excluding atherosclerosis), or diseases associated with cardiovascular diseases (such as sickle cell disease, glaucoma, diabetes, diabetes mellitus, polycythemia vera, and / or systemic inflammatory disorders (such as Behcets disease, polyarteritis nodosa, sarcoidosis, Wegener granulomatosis, and / or Goodpasture syndrome), but preferably excluding diabetes and diabetes mellitus), and / or e. The subject does not have or is not suspected of having one or more conditions selected from oncological conditions, such as solid tumors, lymphomas and / or blood cancers, such as breast cancer, endometrial cancer, cervical cancer, ovarian cancer, lung cancer, head and / or neck cancer, brain cancer, thyroid cancer, esophageal cancer, gastric cancer, colorectal and / or rectal cancer, liver cancer, pancreatic cancer, skin cancer, kidney cancer, bladder cancer, prostate cancer, testicular cancer, bone cancer, lymphoma (such as Hodgkin lymphoma and / or non-Hodgkin lymphoma), and / or blood cancer (such as acute lymphoblastic leukemia, acute myeloid leukemia, and / or chronic myeloid leukemia).

[0038] In one embodiment of the first and / or second aspect of the present invention, the dose of the molecule consisting of monomeric annexin A5 protein is administered to the subject one or more times per day. The dose of the molecule consisting of monomeric annexin A5 protein can optionally be administered to the subject one or more times per day for 1, 2, 3, 4, 5, or 6 or more consecutive days.

[0039] In one embodiment of the first and / or second aspect of the present invention, the molecule consisting of monomeric annexin A5 protein is administered to the vascular system of the subject as a systemic administration such as intravenous administration. When it is administered intravenously, the molecule consisting of monomeric annexin A5 protein, or a composition containing the molecule, can be used in the form of a sterile aqueous solution that can contain, for example, other substances such as salts or glucose sufficient to make the solution isotonic with blood. The aqueous solution should be suitably buffered as necessary (preferably to a pH of 3 to 9). For example, optionally, the aqueous solution has a pH selected from the range of about pH 4 to about pH 8, about pH 5 to about pH 8, about pH 6 to about pH 8, about pH 6.5 to about pH 7.9, about pH 6.8 to about pH 7.7, about pH 7.0 to about pH 7.5, about pH 7.1 to about pH 7.4, about pH 7.1 to about pH 7.4, and optionally, about pH 7.1, 7.2, 7.3, or 7.4, most preferably about pH 7.2.

[0040] Although disclosed herein as a treatment option with respect to the first and / or second aspect of the present invention, it is not very preferable to directly administer the molecule consisting of monomeric annexin A5 protein to the eye. For example, the molecule consisting of monomeric annexin A5 protein can be administered by intravitreal injection. In another option, the molecule consisting of monomeric annexin A5 protein can be directly administered to the retinal vein of the subject (such as the central retinal vein, branch retinal vein, and / or hemiretinal vein, etc.) by, for example, the catheter method. However, this is preferably avoided.

[0041] The monomeric annexin A5 protein for use in the first and / or second aspect of the present invention can optionally be selected from one or more of the following: a. A protein consisting essentially of, or consisting of, the sequence of the human monomeric annexin A5 protein (optionally with or without an N-terminal methionine, the protein consisting of the sequence of SEQ ID NO: 1), b. A recombinant human monomeric annexin A5 protein, c. A mammalian ortholog of the human monomeric annexin A5 protein, d. An allele or genetic variant of a), b), or c), e. A protein consisting of an amino acid sequence that is identical to the human monomeric annexin A5 protein having the sequence defined by SEQ ID NO: 1, with or without an N-terminal methionine, and is more than 50%, 60%, 70%, over 75%, for example, over 80%, over 85%, over 90%, or more preferably over 95% or over 99% identical, a functional analog or variant of the monomeric annexin A5 protein, f. A biologically active fragment of any of a) - e).

[0042] Compositions comprising a molecule consisting of monomeric annexin A5 protein for use according to the first and / or second aspect of the present invention are also provided herein. The composition can be, for example, a unit dosage composition. The unit dosage composition can contain a total dosage of, for example, about 5 mg, about 4 mg, about 3 mg, about 2 mg, or less than about 2 mg, such as about 0.1 to 1.5 mg, about 1 mg or less. Optionally, the composition is a pharmaceutical composition, and further optionally, the composition further comprises one or more pharmaceutically acceptable carriers. In some embodiments, the composition consists of a molecule consisting of monomeric annexin A5 protein and one or more pharmaceutically acceptable carriers. Optionally, the composition is for use as combination therapy with another agent. Optionally, the molecule consisting of monomeric annexin A5 protein is for use as a first-line treatment for the prevention or treatment of one or more diseases, disorders, or conditions associated with and / or caused (directly or indirectly) by macular edema and / or retinal vein occlusion (RVO), and / or macular edema and / or retinal vein occlusion, with or without one or more other agents.

[0043] According to the first and / or second aspect of the present invention, in some embodiments, a molecule consisting of monomeric annexin A5 protein or a composition comprising the molecule is for use as combination therapy with another agent and / or is used such that the subject is administered the molecule consisting of monomeric annexin A5 protein simultaneously with, separately from, or sequentially with one or more other agents in a separate formulation containing one or more other agents, in the same composition as one or more other agents. In some embodiments, the other agent can be an anti-vascular endothelial growth factor (anti-VEGF) agent. Optionally, the molecule consisting of monomeric annexin A5 protein and the anti-VEGF agent are administered in combination, simultaneously in separate administrations, or sequentially in separate administrations, and preferably, when the administrations are separate and sequential, the period between sequential administrations is 28 days or less, such as 3 weeks or less, 2 weeks or less, 1 week or less, 6, 5, 4, 3, 2, or 1 day or less, such as 24, 28, 12, 6, 5, 4, 3, 2, or 1 hour or less.

[0044] In a third aspect, the present invention provides a composition (e.g., a unit dosage composition, etc.) comprising a molecule consisting of a monomeric annexin A5 protein, Optionally, the composition comprises a molecule consisting of a monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, e.g., about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of a monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, e.g., about 1 mg, about 0.1 to 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg, and Optionally, the composition is for use in the prevention or treatment of one or more diseases, disorders, or conditions associated with and / or caused (directly or indirectly) by macular edema and / or retinal vein occlusion (RVO), and / or macular edema and / or retinal vein occlusion in a subject. As used herein, the term "about" may be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0045] In an alternative embodiment of the third aspect, the present invention provides a method for the treatment or prevention of one or more diseases, disorders, or conditions associated with and / or caused (directly or indirectly) by macular edema and / or retinal vein occlusion (RVO), and / or macular edema and / or retinal vein occlusion in a subject (typically a human subject), the method comprising the step of administering a composition comprising a therapeutically effective amount of a molecule consisting of a monomeric annexin A5 protein, Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg. As used herein, the term "about" can be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0046] In one embodiment of the third aspect of the present invention, the composition is a unit dosage composition, and optionally, the treatment and / or prevention can include, or consist of, administering a complete unit dosage to the subject, for example, by a single administration.

[0047] The composition used according to the third aspect of the present invention can be a pharmaceutical composition.

[0048] Optionally, the composition of the third aspect of the present invention further comprises one or more pharmaceutically acceptable carriers.

[0049] For example, the composition can consist of a molecule consisting of monomeric annexin A5 protein and one or more pharmaceutically acceptable carriers.

[0050] In a fourth aspect, the present invention provides a sterile intravitreal composition (e.g., a unit dosage sterile intravitreal composition) comprising a molecule consisting of monomeric annexin A5 protein, Typically, the composition is presented in a sterile syringe, more preferably the syringe contains the composition in a volume of about 200 μL or less, preferably about 100 μL or less, more preferably the volume is about 10 to about 90 μL, about 20 to about 80 μL, about 30 to about 70 μL, about 40 to about 60 μL, or about 50 μL (the term "about" in this context refers to a volume that is ±4, 3, 2, or 1 μL of the stated value), Optionally, the composition contains a molecule comprising monomeric annexin A5 protein, (i) at 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or (ii) in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 to 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg, the term "about" as used in this context may be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2% or ±1% of the recited value, Optionally, the composition is for use in the prevention or treatment of macular edema and / or retinal vein occlusion (RVO) in a subject, and / or one or more diseases, disorders, or conditions associated with and / or caused (directly or indirectly) by macular edema and / or retinal vein occlusion.

[0051] The sterile intravitreal composition of the fourth aspect of the present invention may be presented in the form of a kit containing the composition and one or more additional components.

[0052] For example, the kit may include a composition presented in a sterile syringe and one or more sterile needles attached thereto or separately included in the kit. Preferably, the needle or each needle may be of a gauge selected from gauge 27, 28, 29, or more preferably 30 or 31. The needle or each needle may preferably have a length of about 0.5 - 0.62 inches (12.7 - 15.75 mm). Longer needles may increase the risk of retinal damage if the patient accidentally moves forward during an intravitreal injection procedure.

[0053] In another example, the kit may include one or more additional compositions, each containing one or more antibiotics, for administration to a patient for and / or after intravitreal injection.

[0054] This application also provides, with reference to this specification, a molecule consisting of a monomeric annexin A5 protein or a composition containing such a molecule, or a use, or a method, for use substantially as described herein.

[0055] Definitions As used herein, the term "molecule consisting of a monomeric annexin A5 protein" is further defined in Section B of the Detailed Description of the Invention. It further includes, optionally, a composition that contains, consists essentially of, or consists of a molecule consisting of a monomeric annexin A5 protein. The composition can be, for example, a pharmaceutically acceptable or veterinarily acceptable composition.

[0056] A "pharmaceutically acceptable" composition can be a composition that is safe for administration to a subject, such as a human subject, by injection, such as intravenous, subcutaneous, or intramuscular injection. Most preferably, the injection is an intravenous injection.

[0057] A "veterinarily acceptable" composition can be a composition that is safe for administration to an animal subject by injection, such as intravenous, subcutaneous, or intramuscular injection. Most preferably, the injection is an intravenous injection.

[0058] Accordingly, a pharmaceutical or veterinary composition according to the invention can comprise, consist essentially of, or consist of a molecule comprising a monomeric annexin A5 protein mixed with a pharmaceutically or veterinarily acceptable adjuvant, diluent, and / or carrier, which would typically be selected with respect to the intended route of administration and standard pharmaceutical practice. The composition can be in the form of an immediate release, delayed release, or controlled release application. The formulation can be a unit dosage, for example, a unit dosage containing a daily dose or unit of the active ingredient, a daily sub-dose, or an appropriate fraction thereof.

[0059] A "sterile injectable intravitreal" composition can be a composition that is safe for administration to a subject, particularly a human subject, such as by direct injection into the vitreous of the eye of the subject.

[0060] Accordingly, a "sterile injectable intravitreal composition" according to the invention can comprise, consist essentially of, or consist of a molecule comprising a monomeric annexin A5 protein mixed with an adjuvant, diluent, and / or carrier selected for compatibility and / or tolerance when introduced directly into the vitreous of the eye. The composition can be in the form of an immediate release, delayed release, or controlled release application. The formulation can be a unit dosage, for example, a unit dosage containing a daily dose or unit of the active ingredient.

[0061] The terms "pharmaceutically acceptable", "pharmacologically acceptable", and "veterinarily acceptable" refer to compositions that, for example when administered to animals such as humans as necessary, do not cause harmful, allergic, or other adverse reactions. The same considerations apply to intravitreal compositions for aseptic injection. The preparation of such compositions is known to those skilled in the art in light of the present disclosure, as exemplified by Remington’s Pharmaceutical Sciences, 18th Ed. Mack Printing Company, 1990, which is incorporated herein by reference. Further, in the case of administration to animals (e.g., humans), it will be understood that the formulation should meet the standards of sterility, pyrogenicity, general safety, and purity, such as those required by the FDA Office of Biological Standards.

[0062] As used herein, "pharmaceutically acceptable carriers" and "veterinarily acceptable carriers" include, as known to those skilled in the art, any and all solvents, dispersion media, coatings, surfactants, antioxidants, preservatives (e.g., antibacterial, antifungal agents), isotonic agents, salts, preservatives, drugs, drug stabilizers, excipients, disintegrants, similar substances, etc., and combinations thereof. Their use in therapeutic or pharmaceutical compositions is contemplated, except where any conventional carrier is incompatible with the active ingredient.

[0063] A molecule consisting of monomeric annexin A5 protein, or a composition comprising a molecule consisting of monomeric annexin A5 protein (e.g., mixed with one or more pharmaceutically acceptable carriers, or veterinarily acceptable carriers) can be administered, for example, parenterally to a subject, such as intravenously, intraarterially, intraperitoneally, subarachnoidally, intracerebroventricularly, intrasternal, intracranial, intramuscularly, or subcutaneously, or can be administered by infusion techniques.

[0064] According to one option, administration avoids direct administration to the eye, such as by intravitreal injection.

[0065] In another option, a molecule consisting of monomeric annexin A5 protein can be administered directly to the eye. For example, in an optional and preferred embodiment, a molecule consisting of monomeric annexin A5 protein can be administered to one or both eyes of a subject by intravitreal injection. According to this embodiment, the monomeric annexin A5 protein can be administered to a patient in the form of a sterile intravitreal composition, for example, as defined according to the fourth aspect of the present invention.

[0066] In another option, a molecule consisting of monomeric annexin A5 protein can be administered directly to one or both retinal veins (e.g., central retinal vein, branch retinal vein, and / or hemiretinal vein, etc.) of a subject's eye, for example, by catheterization.

[0067] Systemic administration such as intravenous administration is a preferred embodiment of the first, second, and third aspects of the present invention. A molecule consisting of monomeric annexin A5 protein, or a composition containing such a molecule, can be used, for example, in the form of a sterile aqueous solution that may contain other substances, such as salts or glucose sufficient to make the solution isotonic with blood. The aqueous solution should be suitably buffered as necessary (preferably to a pH of 3 to 9). For example, optionally, the aqueous solution has a pH selected from within the range of about pH 4 to about pH 8, about pH 5 to about pH 8, about pH 6 to about pH 8, about pH 6.5 to about pH 7.9, about pH 6.8 to about pH 7.7, about pH 7.0 to about pH 7.5, about pH 7.1 to about pH 7.4, about pH 7.1 to about pH 7.4, and optionally, about pH 7.1, 7.2, 7.3, or 7.4, most preferably about pH 7.2. Preparation of suitable parenteral formulations under aseptic conditions can be readily achieved by standard pharmaceutical techniques well known to those skilled in the art.

[0068] Formulations suitable for parenteral administration typically include aqueous and non-aqueous sterile injection solutions which may contain antioxidants, buffers, bacteriostats, and solutes which render the formulation isotonic with the blood of the intended recipient, as well as aqueous and non-aqueous sterile suspensions which may contain suspending and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition which requires only the addition of a sterile liquid carrier, for example, water for injection, immediately prior to use. Immediate injection solutions and suspensions may be prepared from sterile powders, granules, and tablets of the previously described types.

[0069] In one embodiment, a pharmaceutically or veterinarily acceptable composition (e.g., such as may be used in any of the first, second, and / or third aspects of the present invention), and / or an intravitreal composition for sterile injection (e.g., as defined according to the fourth aspect of the present invention) may be defined as safe for injection if it does not contain or substantially does not contain endotoxin. Endotoxin is often used synonymously with the term lipopolysaccharide, which is a major component of the cell outer wall of Gram-negative bacteria. It consists of a polysaccharide (sugar) chain and a lipid moiety known as lipid A, which is responsible for the toxic effects observed with endotoxin. The polysaccharide chain is highly variable between different bacteria, determines the serotype of the endotoxin, and the lipid component is also highly variable such that a single endotoxin sample can contain dozens to hundreds of different molecular species. Endotoxin is approximately 10 kDa in size but can form large aggregates up to 1000 kDa. Endotoxin is typically harmful and pyrogenic in therapeutic compositions, and regulatory authorities impose strict limits on the acceptable levels of endotoxin in pharmaceutical compositions. Therefore, the level of endotoxin in the compositions according to the present invention should be minimized and can be less than 100 endotoxin units (EU) per dose, for example, less than 90, 80, 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2, 1, or less EU / dose. The concentration of endotoxin in the compositions according to the present invention can be less than 200 EU / m3, for example, less than 150, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2, 1, or less EU / m3. Methods for measuring endotoxin levels, such as the limulus amoebocyte assay (LAL) method, are well known in the art.

[0070] A composition containing a molecule consisting of monomeric annexin A5 protein may optionally contain, as the sole active agent, a molecule consisting of monomeric annexin A5 protein. In other words, in one embodiment, the composition (or pharmaceutical composition) may consist of a molecule consisting of monomeric annexin A5 protein and one or more pharmaceutically acceptable carriers or one or more veterinarily acceptable carriers. For example, compositions in which annexin A5 is formulated with, conjugated to, or expressed as a fusion protein with one or more additional proteins or non-protein agents that can provide an additional therapeutic effect may optionally be excluded. The additional therapeutic effect may be in either the context of macular edema and / or retinal vein occlusion, or in the context of a therapeutic effect not related to the treatment of macular edema and / or retinal vein occlusion. To avoid misunderstanding, a molecule consisting of monomeric annexin A5 protein is not dimeric annexin A5 (commonly referred to as "diannexin" in the art) and is not PEGylated annexin A5.

[0071] Other definitions: The term "treatment" will be understood by those of ordinary skill in the medical arts. As used herein, the term "treatment" can include any treatment of a condition (e.g., where the condition is macular edema or retinal vein occlusion) in a subject, particularly a human or other mammal, and optionally, the following effects: (i) inhibiting the condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO), i.e., delaying, reducing, or halting the onset of the condition, (ii) reducing the condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO), i.e., causing regression of the condition in a subject having the condition, or (iii) curing a condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO), i.e., returning a subject having the condition to a healthy state where the condition can no longer be detected, including one or more of the above.

[0072] The term "prevention" will be understood by those of ordinary skill in the medical arts. As used herein, the term "prevention" can include any prophylactic treatment of a condition (e.g., where the condition is retinal vein occlusion) in a subject, particularly a human or other mammal, and optionally, the following effects: (i) preventing a condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO) from occurring in a subject (e.g., a subject who has a predisposition or risk of developing the condition but has not yet been diagnosed as having it), i.e., stopping the subject from developing the condition, (ii) delaying the onset of a condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO) in a subject, i.e., delaying the subject's onset of the condition until later in the subject's life, (iii) limiting the occurrence of a condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO) in a subject, e.g., reducing the extent to which the subject is affected by the condition, or (iv) preventing one or more symptoms of a condition (e.g., macular edema or retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema or RVO) in a subject, i.e., stopping the subject from developing one or more symptoms of the condition, including one or more of the above.

[0073] In the case of preventing macular edema or another disease, disorder, or condition associated with and / or caused by RVO, the subject may optionally already have, be suspected of having, and / or be diagnosed as having macular edema or RVO.

[0074] Terms such as "optionally", "preferably", "such as", "for example", etc. mean that the subsequent features, characteristics, events, or situations may or may not occur, and their description includes both the cases where the above features, characteristics, events, or situations occur and the cases where they do not occur. To avoid misunderstanding, in some embodiments of the present invention, it should be understood that any feature, characteristic, event, or situation described in the context of "optionally", "preferably", "such as", "for example", etc. may be excluded from the scope of the claims.

[0075] As used herein, the term "subject" includes the meaning of any living human or animal to which the treatment of the present invention is applicable. By way of non-limiting example, exemplary subjects are further defined in Section C of the "Modes for Carrying Out the Invention" below. More preferably, the subject is a human subject.

[0076] As used herein, the term "elevated" means that a substance (e.g., a cell type such as a red blood cell (i.e., erythrocyte), or a marker on a cell such as an annexin A5 binding site on a red blood cell) is present in a higher amount in a subject having macular edema and / or retinal vein occlusion compared to the amount present in a subject or group of subjects (e.g., at least 10,000 or 1000 subjects) not having macular edema and / or retinal vein occlusion.

[0077] As used herein, the term "distal" includes the meaning that the administration of a molecule consisting of monomeric annexin A5 protein is not a direct administration into or to an eye diagnosed with macular edema and / or retinal vein occlusion.

[0078] The term "therapeutically effective amount" refers to an amount of a drug, compound, protein (e.g., a molecule consisting of monomeric annexin A5 protein as defined herein), or pharmaceutical composition sufficient to provide a beneficial benefit or desirable result including clinical outcomes related to the treatment or prevention of retinal vein occlusion and / or macular edema, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema and / or retinal vein occlusion. The effective amount can be administered in one or more administrations. For the purposes of the present invention, an effective amount of a drug, compound, protein (e.g., a molecule consisting of monomeric annexin A5 protein as defined herein), pharmaceutical composition, or veterinary composition is an amount sufficient to treat, relieve, reduce the intensity of, and / or prevent macular edema and / or retinal vein occlusion, and / or other diseases, disorders, or symptoms associated with and / or caused by macular edema and / or retinal vein occlusion. As understood in the clinical context, an effective amount of a drug, compound, protein (e.g., a molecule consisting of monomeric annexin A5 protein as defined herein), or pharmaceutical composition may or may not be achieved when administered in combination with (such as separately, sequentially, or simultaneously) another drug, compound, protein, or pharmaceutical as described elsewhere in this application. Thus, "effective amount" can be considered in the context of administering one or more therapeutic agents, and a single agent may be considered to be administered in an effective amount if a desirable result can be provided or achieved in combination with one or more other agents.

Brief Description of the Drawings

[0079]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Figure 7

Mode for Carrying Out the Invention

[0080] Established standard treatments for macular edema and / or RVO and, in fact, in a plurality of other retinal diseases are by direct delivery of a therapeutic agent (e.g., an anti-VEGF therapeutic agent) to the eye, by intravitreal injection (Haydinger et al, 2023 (supra), Ip & Hendrick, 2018 (supra), Cox et al., 2021, J. Clin. Med., 10(5):981). Since the retina is located at the back of the eye and is surrounded by a plurality of protective layers including the cornea, iris, and lens, such drugs are injected directly into the eye. These layers can make it difficult for the drugs to reach the retina if they are administered by other routes. When drugs are injected into the eye, they are delivered directly to the retina and can act locally to target the cause of the problem. The present invention is based on the surprising determination that monomeric annexin A5 protein can preferentially localize to the occlusion site in a subject with retinal vein occlusion (RVO) when administered systemically to the subject (e.g., by intravenous administration).

[0081] In addition, annexin A5 has been determined to provide biological activity that modulates various factors that contribute to breakdown of the blood-retinal barrier (BRB) in subjects with macular edema and / or RVO and that contribute to other therapeutic benefits in subjects with macular edema and / or RVO. The inventors have also determined that the prevention and / or treatment of macular edema, retinal vein occlusion, and / or other diseases, disorders, or conditions associated with and / or caused by macular edema and / or retinal vein occlusion can be achieved using a range of doses of annexin A5, including (but not limited to) low doses of monomeric annexin A5.

[0082] A. Macular Edema Macular edema is the pathological accumulation of fluid in the macula, the central region essential for high-precision vision. This is a complication common to many eye diseases, such as diabetic retinopathy, retinal vascular occlusion, uveitis, post-cataract surgery inflammation (pseudophakic macular edema or air-bubble syndrome), retinal dystrophy, drug reactions, intraocular tumors, serous central chorioretinopathy, radiation retinopathy, and other retinal vascular abnormalities, including retinal arteriovenous aneurysms and retinal telangiectasias (Tranos et al., 2004, Surv Ophthalmol, 49:470-90). Subretinal neovascular membranes observed in conditions such as age-related macular degeneration (AMD) are also associated with intraretinal and subretinal fluid accumulation (Trichonas et al., 2014, Br J Ophthalmol. 98:ii24).

[0083] Symptoms of macular edema include metamorphopsia, micropsia, haze, central scotoma, and decreased contrast or color sensitivity (Haydinger et al., 2023).

[0084] Optionally, treatment of macular edema or a condition associated therewith is initiated within 10, 20, 30, 40, 50, or 60 minutes, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours, or 1, 2, 3, 4, 5, 6, or 7 days of the onset of one or more symptoms of macular edema, according to various aspects of the present invention.

[0085] Clinical diagnosis of macular edema can be difficult in mild cases or when impaired by mydriasis, cataracts, and other media opacities that reduce fundus visibility. Thus, fluorescein angiography and optical coherence tomography (OCT) are commonly used to make the diagnosis. The latter is preferred because of its non-invasive nature, accurate measurement of retinal thickness, and ability to identify other structural abnormalities such as epiretinal membranes and vitreomacular traction, and is the standard method for assessing macular edema in clinical trials.

[0086] Thus, in a preferred embodiment, a subject diagnosed as having macular edema is a subject in which macular edema is diagnosed by fluorescein angiography and / or optical coherence tomography.

[0087] Optionally, according to the present invention, treatment of macular edema or a condition associated therewith in a subject is initiated within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or 60 minutes, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours, or 1, 2, 3, 4, 5, 6, or 7 days of the diagnosis of macular edema.

[0088] The thickness of the retina is generally measured between the inner limiting membrane and Bruch's membrane. According to current spectral domain OCT technology, the average macular thickness and central (foveal) macular thickness in the normal adult eye are 334 and 226 μm, respectively, and are thicker in men than in women. Three patterns of macular edema have been described using OCT: cystoid macular edema, diffuse macular edema, and subretinal detachment. Diffuse retinal thickening is the main pattern in diabetic macular edema, whereas in retinal vascular occlusion, accumulation of subretinal fluid is common. In uveitis, cystoid macular edema and diffuse macular edema are typical forms of macular edema, whereas subretinal detachment is usually seen in less than one-third of cases, in combination with cystoid or diffuse edema. In some embodiments of the present invention, macular edema is selected from the group consisting of one or more of cystoid macular edema, diffuse macular edema, and subretinal detachment.

[0089] Distortion of the retinal structure caused by fluid accumulation in the macula results in visual loss and, if the condition becomes chronic, irreversible visual impairment due to scarring. Macular edema is the main cause of visual loss in patients with true diabetes, retinal vein occlusion, and uveitis. Population studies have shown that the average prevalence of diabetic macular edema is approximately 6% in type 1 diabetes and slightly higher in type 2 diabetes. Macular edema is very common in central retinal vein occlusion, whereas in branch retinal vein occlusion, macular edema occurs in 30% of cases. Approximately one-third of patients with uveitis develop macular edema, which is more frequent in intermediate uveitis and panuveitis and less common in anterior uveitis. The burden of macular edema is related not only to the reduction in quality of life experienced by the patient, but also to the economic costs associated with the chronicity of the condition and frequent visits to treatment and medical centers. In particular, in uveitic macular edema, the most affected population group is of working age, and the socio-economic impact of this condition is increasing.

[0090] In some embodiments of the present invention, a subject having, diagnosed as having, or suspected of having macular edema may be a subject having, diagnosed as having, or suspected of having one or more conditions selected from true diabetes, diabetic retinopathy, retinal vein occlusion (e.g., CRVO, BRVO, and / or HRVO), and / or uveitis (e.g., intermediate uveitis and / or panuveitis).

[0091] In some embodiments of the present invention, a subject having, diagnosed as having, or suspected of having macular edema may be a subject who has received, is receiving, or is scheduled to receive a treatment selected from the group consisting of related therapies for macular edema, such as anti-VEGF therapy (e.g., anti-VEGF therapy delivered to the eye by intravitreal injection), corticosteroids, NSAIDs, pars plana vitrectomy, retinal laser photocoagulation, or anti-VEGF therapy enhanced with intravitreal corticosteroids and / or retinal laser photocoagulation.

[0092] Exemplary anti-VEGF therapies include any one of the following: · Ranibizumab, an FDA-approved anti-VEGF drug and a recombinant humanized antibody fragment against VEGF, · Aflibercept, another FDA-approved anti-VEGF drug and a recombinant protein containing the ligand-binding domains of VEGF receptors 1 and 2 fused to the Fc portion of immunoglobulin G1, · Bevacizumab, a humanized monoclonal against VEGF, widely used off-label (Haydinger et al, 2023 (supra), Ip & Hendrick, 2018 (supra)), and / or · Brolucizumab and / or Faricimab, new anti-VEGF drugs.

[0093] Corticosteroids are commonly used in the management of macular edema, particularly in subjects with non-infectious uveitis. Exemplary corticosteroids include difluprednate, triamcinolone acetonide, triamcinolone acetonide, and dexamethasone, which can be used as monotherapy or in combination with systemic immunosuppressive therapy to achieve more rapid resolution of the edema. The most common complications of topical corticosteroid treatment are increased intraocular pressure and cataract. Systemic use of corticosteroids is mainly indicated for bilateral macular edema and should be used for a limited period because of its wide range of side effects (Foster et al., 2016, Surv Ophthalmol, 61:1-17).

[0094] Non-steroidal anti-inflammatory drugs (NSAIDs) have an adjunctive role in the treatment approach and are mainly used for pseudophakic macular edema. Acetazolamide, an inhibitor of carbonic anhydrase, may be used when corticosteroid therapy fails, but may also be used as primary therapy for pseudophakic macular edema or macular edema associated with retinal dystrophy.

[0095] Pars plana vitrectomy is mainly used to treat the organic factors contributing to macular edema, such as epiretinal membranes and vitreoretinal traction. It is the last resort for the treatment of macular edema without organic factors because of its inherent risks, including retinal breaks and detachments, vitreous hemorrhage, cataract, glaucoma, and hypotony, as well as limited data on outcomes.

[0096] Fluid can diffusely accumulate in the center of the retina, or within cysts typically located in the inner nuclear layer or Henle fiber layer, or in the subretinal space, disrupting and distorting the retinal structure and potentially reducing vision (1, 2). In combination with the underlying pathology, the edema can progress to cause irreversible tissue damage with death of retinal cells and permanent visual impairment. A common feature of most diseases causing macular edema is dysfunction of the blood-retinal barrier (BRB), a series of structures that tightly regulate the passage of proteins, salts, metabolites, and other solutes between the blood and retinal tissue in a healthy state. Vasoproliferative factors and inflammatory cytokines, including vascular endothelial growth factor-A (VEGF) and tumor necrosis factor (TNF)-α, are upregulated in the ocular fluid in macular edema and are involved in the breakdown of the BRB. Dysregulation of the regulation of solute influx and accumulation in the retina disrupts the balance of osmotic and hydrostatic pressures, leading to fluid influx when the mechanisms maintaining fluid homeostasis are breached. Current treatments target angiogenesis, inflammation, and proteins and processes involved in blood-retinal barrier (BRB) dysfunction. This review provides an overview of the clinical aspects of macular edema and considers the cellular and molecular mechanisms targeted by current treatments.

[0097] Conditions associated with macular edema include, but are not limited to, dysregulation of the blood-retinal barrier (BRB), such as dysregulation of the inner BRB and / or outer BRB, dysfunction of the barrier between the vitreous and the vessels of the ciliary body, ocular diseases associated with macular edema, such as diabetic retinopathy, retinal vascular occlusion, uveitis, postoperative inflammation after cataract surgery (pseudophakic macular edema or Irvine-Gass syndrome), retinal dystrophy, drug reactions, intraocular tumors, serous central chorioretinopathy, radiation retinopathy, and other retinal vascular abnormalities including retinal arteriovenous aneurysms and retinal capillary telangiectasia, changes in subretinal neovascular membranes observed in conditions such as age-related macular degeneration (AMD), metamorphopsia, micropsia, foggy vision, central scotoma, and any one or more of a decrease in contrast or color sensitivity.

[0098] Mechanisms of Macular Edema Macular edema is the result of a failure of the mechanisms that maintain fluid homeostasis in the macula (Haydinger et al, 2023 (supra)). The movement of fluid across capillaries throughout the body is classically explained by Starling forces. The hydrostatic pressure gradient between the intravascular space and the extracellular space of the tissue tends to drive fluid into the tissue, which is counteracted by the colloid osmotic (oncotic) pressure gradient that tends to drive fluid from the extracellular space into the blood vessels. A recent review by Cunha-Vaz considers the formation of macular edema in relation to classical Starling forces (Ophthalmologica, 2017, 237:1-10). An increase in the relative hydrostatic pressure of the blood vessels relative to the retinal tissue, and an increase in the relative colloid osmotic pressure in the retinal tissue relative to the blood vessels, are the two forces that drive edema. Of these, macular edema in most diseases is thought to be due to an increase in tissue colloid osmotic pressure resulting from abnormal accumulation of solutes in the extracellular space of the retina (Haydinger et al, 2023 (supra)). The compliant nature of the retinal tissue, which can expand unhindered into the vitreous, is particularly susceptible to the effects of edema because an increase in tissue colloid osmotic pressure readily leads to an increase in tissue volume.

[0099] Dysregulation of the influx of solutes into the retina occurs as a result of disruption of the blood-retinal barrier (BRB), which is the main focus of most efforts to understand the mechanisms of this condition (Haydinger et al, 2023 (supra)).

[0100] The BRB is a group of intraocular structures that separate the blood from the neural retina and the extracellular space of the retina. There are two components: the inner BRB and the outer BRB. The inner BRB is formed around the retinal endothelial cells (RECs) that line the retinal blood vessels, and the outer BRB is formed at the level of the retinal pigment epithelial (RPE) cells. Since there can be non-selective bidirectional diffusion of solutes between the extracellular space of the retina and the vitreous, the barrier process between the vitreous and the blood vessels in the ciliary body may also be considered part of the BRB, although this region is not classified as either the inner BRB or the outer BRB.

[0101] The walls of retinal blood vessels form the inner BRB. The luminal surface of retinal blood vessels is continuously covered by a monolayer of flat epithelial cells of RECs, and the membranes of adjacent RECs are held together by a continuous network of tight junctions. RECs are not fenestrated and contain few pinocytotic vesicles. RECs and their junctions form a barrier that facilitates the controlled passage of solutes between the bloodstream and the retina (Cunha-Vaz et al., 1966, Br J Ophthalmol., 50:441-53). The basement membrane completely covers the abluminal surface of RECs. Evidence collected in vivo indicates that the basement membrane does not directly constitute a gate, but some evidence indicates that it can impede the diffusion of molecular tracers in vitro. Larger blood vessels have additional layers of smooth muscle, basement membrane, and adventitia with structures that vary according to the size and type of the vessel.

[0102] The outer BRB is not located in the walls of blood vessels but rather is formed by a monolayer of RPE cells located behind the retinal pigment epithelium, photoreceptors, and immediately opposite Bruch's membrane, which separates it from the vascular bed of the choroidal capillary lamina (Haydinger et al, 2023 (supra)). In contrast to the inner BRB, the blood vessels of the choroid have thin and highly fenestrated endothelial cells and do not form a selective gate. Thus, the epithelium is involved in maintaining the controlled environment necessary for retinal function.

[0103] Dysfunction of the BRB underlies the development of macular edema (Haydinger et al, 2023 (supra)). Loss of selectivity in the molecular exchange between blood and retinal tissue at any level of the inner or outer BRB allows entry of proteins and other solutes into the retinal tissue. Leakage is usually evident well before the formation of macular edema. Edema is likely to occur when the severity of leakage exceeds solute and fluid clearance mechanisms. The rate of solute clearance from the subretinal space is inversely proportional to their size. Injection of differently sized tagged dextran tracers into patients with uveitic macular edema showed leakage rates approximately proportional to the size of the tracer. Sizes up to 20 kDa leaked into the macula, but 150 kDa did not (Atkinson et al., 1991, Eye, 5:440-6). These data indicate that infiltration of larger solutes is likely to pose a greater challenge to clearance, but that complete breakdown of the BRB, which allows non-selective entry of the largest solutes, is not necessary for the formation of edema.

[0104] This evidence suggests that several inflammatory cytokines act to disrupt BRB integrity (Haydinger et al, 2023 (supra), Park et al. 2019, J Diabetes Res, 2019:813741, Noma et al., 2020, J.Clin.Med, 9(1):3457).

[0105] VEGF plays a major role in driving the progression of retinal diseases and macular edema, as evidenced incontrovertibly by the widespread utility of anti-VEGF therapy (Haydinger et al, 2023 (supra), Park et al. 2019, (supra), Noma et al., 2020 (supra)).

[0106] The intravitreal levels of VEGF and IL-6 correlate with the severity of macular edema and the size of the non-perfused area (Noma et al., 2009, Ophthalmology, 116:87-93, Noma et al, 2006, Graefes Arch Clin Exp Ophthalmol., 244:309-15). Evidence indicates that IL-6 is involved in the diseases that cause macular edema (Haydinger et al, 2023 (supra)). It is elevated in patients with uveitis (van Kooij et al., 2006, Am J Ophthalmol. 142:192-4). It is elevated in diabetic macular edema (Funatsu et al., 2002, Am J Ophthalmol. 133:70-7) and correlates with the thickness of the macula (Oh et al., 2010, Curr Eye Res. 35:1116-27). IL-6 is also upregulated in patients with central retinal vein occlusion, particularly those with ischemia (Noma et al., 2009, Ophthalmology. 116:87-93). Its expression often correlates with the expression of VEGF (Haydinger et al, 2023 (supra)). Furthermore, VEGF is thought to increase vascular permeability and further increase the expression of inflammatory cytokines including MCP-1 and ICAM-1 that disrupt the BRB, thereby causing the onset and progression of macular edema (Noma et al., 2020 (supra)).

[0107] Tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are also involved in the mechanism of BRB dysfunction in macular edema (Haydinger et al, 2023 (supra)).

[0108] TNF-α is a major pro-inflammatory cytokine shown to increase the paracellular permeability of REC monolayers in vitro (Haydinger et al, 2023 (supra), Aveleira et al., 2010, Diabetes. 59:2872-82). When injected into the vitreous of rabbits, TNF-α increased the proportion of REC tight junctions that appeared open, and albumin was detected within these junctions, indicating leakage (Luna et al., 1997, J Neurosci Res. 49:268-80). Larger vesicles were also positively stained for albumin in the RECs of treated eyes but not in controls. These results indicate disruption of both the transcellular and paracellular pathways. Interestingly, 24 hours after injection, TNF-α-induced tight junction changes reversed, which is a promising sign for a potential therapeutic target. When injected into the vitreous of rats, it also induced paracellular leakage, and the effect of the injection recovered after 7 days (Bamforth et al., 1996, Acta Neuropathol. 91:624-32). Huang et al. used two mouse models of diabetes to evaluate the pathological role of TNF-α (Ophthalmol Vis Sci. (2011) 52:1336-44), and vascular leakage in the late stage of the disease was completely prevented by TNF-α knockout.

[0109] Along with TNF-α, another major inflammatory cytokine that has been gaining increasing interest is IL-1β. Similar to TNF-α, when injected into the vitreous of rabbits, IL-1β increases the number of open leaky tight junctions (140). It also increases the permeability of REC monolayers in vitro (Aveleira et al., 2010 (cited above)). In streptozotocin-diabetic rats, retinal IL-1β expression increases between 1.5 and 4.5 months of diabetes, and IL-1β mRNA is induced by high glucose in bovine REC in vitro (Liu et al., 2012, PLoS One. 7: e36949), which induces its own expression in REC, Müller cells, and astrocytes in vitro, and this is inhibited by inhibition of protein kinase C.

[0110] B. Retinal vein occlusion Types of retinal vein occlusion Aspects of the present invention relate to the prevention and treatment of retinal vein occlusion and / or one or more diseases, disorders, or conditions associated with and / or caused by retinal vein occlusion.

[0111] Retinal vein occlusion (sometimes referred to as eye stroke) is a common retinal vascular disorder and one of the most common causes of vision loss worldwide. Generally, retinal vein occlusion is thought to be caused by occlusion (i.e., blockage) of the retinal venous system, such as by thrombosis. Other possible causes of retinal vein occlusion are thought to be occlusion of the vein caused by external pressure on the retinal vein or diseases of the retinal vein wall, such as vasculitis ("Retinal Vein Occlusion (RVO) Guidelines" (cited above)).

[0112] Red blood cells can coagulate to form thrombi, which is known to be mediated by phosphatidylserine exposed on the surface of red blood cells. Without being bound by theory, Applicant has characterized that the function of annexin A5 to mask (or in other words, cover or shield) phosphatidylserine on the surface of these red blood cells can avoid and treat thrombus formation in retinal vein occlusion. By characterizing the function of exposed phosphatidylserine in thrombus formation in retinal vein occlusion, Applicant has confirmed that it is possible to treat patients with substantially less monomeric annexin A5 than previously considered effective in the art.

[0113] In one embodiment, retinal vein occlusion is caused by occlusion of a blood vessel. For example, occlusion of a retinal vein can be related to one or more of formation of a thrombus (singular or plural), observably enhanced red blood cell aggregation and / or blood hyperviscosity, external pressure on the vein, and disease of the vein wall (such as vasculitis, etc.). Most preferably, occlusion of the retinal vein is related to formation of a thrombus.

[0114] "Related" includes that retinal vein occlusion is caused by a particular mechanism (such as thrombus formation, etc.), and / or it is suspected that retinal vein occlusion is caused by a particular mechanism, and / or it is diagnosed that retinal vein occlusion is caused by a particular mechanism. As will be appreciated herein, methods for identifying whether retinal vein occlusion is related to a particular mechanism will be known to those of ordinary skill in the art.

[0115] As is known to those of ordinary skill in the art, thrombosis is generally considered to be a blood clot formed in a blood vessel (such as a vein, etc.) by aggregation of platelets and / or red blood cells that impede blood flow. Thus, "thrombus" includes a blood clot formed in a blood vessel (such as a vein, etc.) that impedes blood flow.

[0116] Subjects can be classified as having observably enhanced erythrocyte aggregation and / or blood hyperviscosity by any suitable method. This enhancement is compared to healthy subjects or to the mean for a group of healthy subjects (e.g., a group of 10, 50, or 100 healthy subjects). For example, the levels of erythrocyte aggregation and / or blood hyperviscosity can be measured by determining the level of erythrocyte adhesion to endothelial cells. Suitable methodologies are described, for example, in Wauter et al, 2011, J. Thrombosis and Hemostasis, 9:1049-1055, the content of which is incorporated herein by reference for the description of the erythrocyte adhesion method.

[0117] More specifically, in one option, the level of erythrocyte adhesion to endothelial cells can be determined under static conditions. This can involve using radiometric techniques as described, for example, in Kuypers et al, 1996, Blood, 87:1179-87 (the content of which is incorporated herein by reference). Human microvascular endothelial cells (HMEC-1) can be cultured and incubated with washed erythrocytes resuspended (e.g., at 25% hematocrit in Hanks Balanced Salt Solution (HBSS), preferably labeled (e.g., 51 with chromium) and added with 0.5% human serum albumin for (e.g., 30 minutes at 37°C). Unattached erythrocytes are removed by washing, and the remaining attached erythrocytes can be detected. Such detection can involve, for example, lysing the remaining attached erythrocytes (e.g., with distilled water) and measuring the radioactivity in the lysate. Using such techniques, adhesion can be calculated, for example, as the number of erythrocytes attached per 1 mm 2 area.

[0118] Under such static conditions, for example, in the case of healthy individuals, typically, 3700 ± 300 erythrocyte adhesions per 1 mm 2 area can be shown, but more generally, patients with CRVO can have 1 mm 2It can show 13,500 ± 700 red blood cells per mm. The subject to be treated according to the present invention is optionally, for example, when evaluated for adhesion to endothelial cells under static conditions as determined by techniques such as those described above, 1 mm 2 per mm can have more than 4,000, for example, about 4,500, about 5,000, about 5,500, about 6,000, about 6,500, about 7,000, about 7,500, about 8,000, about 8,500, about 9,000, about 9,500, about 10,000, about 10,500, about 11,000, about 11,500, about 12,000, about 12,500, about 13,000, about 13,500, about 14,000, about 14,500, about 15,000, about 15,500, about 16,000, about 16,500, about 17,000 red blood cells and can be classified as such. The term "about" in this context refers to a value of ±500, 400, 300, 200, 100, 50 or fewer red blood cells per mm 2 of the stated value.

[0119] In another option, the level of red blood cell adhesion to endothelial cells can be determined under flow conditions. This can involve, for example, the adhesion of red blood cells to HMEC under flow conditions determined according to Wautier et al, 2007, Blood, 110:894 - 901, the content of which is incorporated herein by reference. For example, HMEC are cultured (e.g., for 24 hours in a glass capillary coated with gelatin (2%)), then a microslide containing confluent HMEC is placed on the stage of a video microscope, and a red blood cell suspension (e.g., hematocrit 0.5%) is perfused through the microslide at a suitable flow rate (e.g., a rate corresponding to a wall shear stress of 0.03 Pa) for a suitable time (e.g., 10 minutes), followed by washing away non - adherent cells (e.g., for 10 minutes). Then, the wall shear stress can be increased step - by - step (e.g., from 0.03 to 0.3 Pa every 5 minutes). In each step, the attached red blood cells are recorded (e.g., by video imaging in 10 consecutive fields of view, with one field of view measured at 0.1 mm 2 ), counted (e.g., using a computer - processed image analysis system), and the results are per 1 mm 2It can be expressed as the number of red blood cells per hit.

[0120] Under such flow conditions, for example, when analyzing adhesion at 0.03 Pa and washout at 0.07 Pa, in the case of a healthy individual, typically, 1 mm 2 can show adhesion of 30 ± 4 red blood cells per hit, but more generally, patients with CRVO can show 190 ± 8 red blood cells per 1 mm 2 per hit. A subject treated according to the present invention is optionally, for example, when determined by techniques such as those described above, when evaluated for adhesion to endothelial cells under flow conditions, 1 mm 2 per hit can be classified as having 35, for example, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, about 100, about 105, about 110, about 115, about 120, about 125, about 130, about 135, about 140, about 145, about 150, about 155, about 160, about 165, about 170, about 175, about 180, about 185, about 190, about 195, about 200 or more red blood cells. The term "about" in this context refers to a value of ±5, 4, 3, 2, or 1 red blood cell per 1 mm 2 per hit.

[0121] Under such flow conditions, for example, when analyzing adhesion at 0.03 Pa and washout at 0.3 Pa, in the case of a healthy individual, typically, 1 mm 2 can show adhesion of less than 1 red blood cell per hit, but more generally, patients with CRVO can show 20 ± 8 red blood cells per 1 mm 2 per hit. A subject treated according to the present invention is optionally, for example, when determined by techniques such as those described above, when evaluated for adhesion to endothelial cells under such flow conditions, 1 mm 2Those having red blood cells showing more than one red blood cell per, for example, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30 or more can be classified. The term "about" in this context refers to a value of red blood cells that is ±50%, 40%, 30%, 20%, 10% or less of the stated value per 1 mm 2 per red blood cell.

[0122] The retina usually has three veins associated with retinal vein occlusion, which are the central retinal vein, the hemicentral retinal vein, or the retinal branch vein ("Retinal Vein Occlusion (RVO) Guidelines", The Royal College of Ophthalmologists, July 2015).

[0123] The central vein, hemicentral vein, and branch central vein provide names for subtypes of retinal vein occlusion, which are central retinal vein occlusion (CRVO), hemicentral retinal vein occlusion (HRVO), and retinal vein branch occlusion (BRVO).

[0124] Central retinal vein occlusion is caused by occlusion of the central retinal vein, particularly the part passing through the retinal lamina cribrosa. Hemicentral retinal vein occlusion is caused by occlusion of the retinal vein in the upper or inner hemisphere of the retina. Retinal vein branch occlusion is caused by occlusion at the arteriovenous crossing where the retinal vein and retinal artery share a common vascular sheath.

[0125] Thus, in one embodiment, the retinal vein in which RVO is present and / or the present invention is directed to prevention or treatment is one or more selected from the central vein, hemicentral vein, and branch central vein. In other words, in one embodiment, retinal vein occlusion is one or more of central retinal vein occlusion, hemicentral retinal vein occlusion, and retinal vein branch occlusion.

[0126] Preferably, the retinal vein occlusion is central retinal vein occlusion.

[0127] Optionally, in accordance with various aspects of the present invention, the treatment of RVO or macular edema in a subject having retinal vein occlusion (RVO) is initiated within 10, 20, 30, 40, 50, or 60 minutes, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours, or 1, 2, 3, 4, 5, 6, or 7 days of the onset of symptoms of retinal vein occlusion. More preferably, in accordance with the present invention, the treatment of retinal vein occlusion or macular edema in a subject having RVO is initiated within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or 60 minutes, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours, or 1, 2, 3, 4, 5, 6, or 7 days of the diagnosis of retinal vein occlusion.

[0128] Retinal vein occlusion can also be classified as either ischemic or non-ischemic retinal vein occlusion. This is generally a classification regarding areas of capillary non-perfusion, as discussed herein. Thus, in one embodiment, retinal vein occlusion is ischemic retinal vein occlusion (e.g., ischemic central retinal vein occlusion, etc.) or non-ischemic retinal vein occlusion (e.g., non-ischemic central retinal vein occlusion, etc.).

[0129] Retinal vein occlusion can be further classified as acute retinal vein occlusion. In acute retinal vein occlusion, RVO can occur due to severe ischemia within the retinal vein, and the onset of symptoms is rapid. Therefore, it is necessary to provide treatment as soon as possible after the onset and / or diagnosis of acute retinal vein occlusion. Thus, in one embodiment, the retinal vein occlusion is an acute retinal vein occlusion such as, for example, acute central retinal vein occlusion. Preferably, the treatment of acute retinal vein occlusion according to the present invention is initiated within 10, 20, 30, 40, 50, or 60 minutes, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours, or within 1, 2, 3, 4, 5, 6, or 7 days after the onset of symptoms of acute retinal vein occlusion. More preferably, the treatment of acute retinal vein occlusion according to the present invention is initiated within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or 60 minutes, or within 1, 2, 3, 4, 5, or 6 hours after the diagnosis of acute retinal vein occlusion.

[0130] Retinal vein occlusion can affect either one eye (or one retina) referred to as unilateral retinal vein occlusion, or both eyes (or both retinas) referred to as bilateral retinal vein occlusion. Thus, in one embodiment, the retinal vein occlusion is unilateral retinal vein occlusion (such as, for example, unilateral central retinal vein occlusion). In another embodiment, the retinal vein occlusion is bilateral retinal vein occlusion (such as, for example, bilateral central retinal vein occlusion).

[0131] Diagnosis and Characterization of Retinal Vein Occlusion Methods for diagnosing retinal vein occlusion, as well as subtypes of central retinal vein occlusion, hemicentral retinal vein occlusion, and branch retinal vein occlusion are known to those of ordinary skill in the medical arts. Examples of such diagnoses are considered in "Retinal Vein Occlusion (RVO) Guidelines" (supra). Generally, patients with retinal vein occlusion will present to a physician with changes in their visual field such as blurred vision, partial loss of vision, floating spots within vision, flashes of light within vision, reduced or decreased vision (i.e., a reduction or decrease in vision), loss of vision (i.e., blindness), etc. In some situations, the loss of vision is sudden. In addition, in some situations, the patient will show no pain.

[0132] Some of the characteristics of retinal vein occlusion that can be used for diagnosis are as follows: papilledema (i.e., swelling of the optic nerve head), increased dilation of the retinal veins, increased tortuosity of the retinal veins, eye hemorrhage, macular edema, hemorrhage, "cotton wool spots" (these are white spots on the retina and can be observed during fundus examination of the retina), retinal edema, electrodiagnostic test (ERG) results, concentric pupillary defect, capillary non-perfusion, arteriolar narrowing, venous occlusion, and retinal pigment epithelial changes. These characteristics can generally be observed by a physician through examination of the fundus (the inner surface of the eye on the opposite side of the lens) or the retina using ophthalmoscopy and / or fundus photography. Another diagnostic method used particularly to identify venous occlusion is angiography (such as angiography using a fluorescent dye (e.g., fluorescein)). In the case of fluorescein angiography, a dye is injected into the patient and the behavior of the dye in the eye's veins is used to identify whether there is an occlusion in the veins.

[0133] In one embodiment, the subject is diagnosed as having a retinal vein occlusion, such as central retinal vein occlusion, hemicentral retinal vein occlusion, and branch retinal vein occlusion, preferably central retinal vein occlusion. In an alternative embodiment, the subject is suspected of having a retinal vein occlusion, such as central retinal vein occlusion, hemicentral retinal vein occlusion, and branch retinal vein occlusion, preferably central retinal vein occlusion.

[0134] In one embodiment, the subject has one or more symptoms of retinal vein occlusion, such as central retinal vein occlusion, hemicentral retinal vein occlusion, and branch retinal vein occlusion, preferably central retinal vein occlusion.

[0135] In one embodiment, the retinal vein occlusion is characterized by a change in the subject's vision, such as one or more of blurred vision, partial loss of vision, floating spots in vision, flashes of light in vision, reduced or decreased vision, and loss of vision. Typically, retinal vein occlusion is characterized by reduced or lost vision, most typically sudden reduced or lost vision.

[0136] In another embodiment, the retinal vein occlusion is characterized by a significant change in the subject's vision that occurs within 7 days, such as within 6 days, within 5 days, within 4 days, within 3 days, within 2 days, or within 1 day. This can optionally be a characteristic of acute RVO.

[0137] Otherwise, typically, the loss of vision, or reduced or decreased vision, occurs over a period of about 7 days or more, such as about 8 days or more, about 9 days or more, about 10 days or more, about 11 days or more, about 12 days or more, about 13 days or more, about 14 days or more, about 15 days or more, about 16 days or more, about 17 days or more, about 18 days or more, about 19 days or more, about 20 days or more, about 21 days or more, about 22 days or more, about 23 days or more, about 24 days or more, about 25 days or more, about 26 days or more, about 27 days or more, about 28 days or more, about 29 days or more, about 30 days or more, about 31 days or more, about 1 month or more, about 2 months or more, about 3 months or more, about 4 months or more, about 5 months or more, about 6 months or more, about 7 months or more, about 8 months or more, about 9 months or more, about 10 months or more, about 11 months or more, about 12 months or more, about 13 months or more, about 14 months or more, about 15 months or more, about 16 months or more, about 17 months or more, about 18 months or more, about 19 months or more, about 20 months or more, about 21 months or more, about 22 months or more, about 23 months or more, about 24 months or more, about 36 months or more, about 3 years or more, about 4 years or more, about 5 years or more, about 6 years or more, about 7 years or more, about 8 years or more, about 9 years or more, or about 10 years or more.

[0138] During the immediately preceding period, vision may be reduced or decreased until vision loss (i.e., blindness) occurs or until vision reaches a certain percentage of normal vision, at which point a certain percentage of vision is maintained. Typically, the certain percentage of reduction or decrease in vision is 95% or less of normal vision, for example, 90% or less of normal vision, 85% or less, 80% or less, 75% or less, 70% or less, 69% or less, 68% or less, 67% or less, 66% or less, 65% or less, 64% or less, 63% or less, 62% or less, 61% or less, 60% or less, 59% or less, 58% or less, 57% or less, 56% or less, 55% or less, 54% or less, 53% or less, 52% or less, 51% or less, 50% or less, 49% or less, 48% or less, 47% or less, 46% or less, 45% or less, 44% or less, 43% or less, 42% or less, 41% or less, 40% or less, 39% or less, 38% or less, 37% or less, 36% or less, 35% or less, 34% or less, 33% or less, 32% or less, 31% or less, 30% or less, 25% or less, 20% or less, 15% or less, 10% or less, or 5% or less. Normal vision includes the vision of the subject before the onset of retinal vein occlusion or the normal vision of the general population. If the retinal vein occlusion is unilateral retinal vein occlusion, normal vision can be the eye that is not affected by retinal vein occlusion.

[0139] In one embodiment, the subject exhibits no pain in the eye and / or no pain in the retina.

[0140] In one embodiment, the retinal vein occlusion is characterized by one or more of the following: papilledema, increased dilation of retinal veins, increased tortuosity of retinal veins, hemorrhage, "cotton wool spots", retinal edema, electroretinogram (ERG), concentric pupillary defect, capillary non-perfusion, arteriolar narrowing, and retinal pigment epithelial changes.

[0141] In one embodiment, the retinal vein occlusion (e.g., central retinal vein occlusion, etc., preferably ischemic central retinal vein occlusion) is characterized by one or more of the following: · Poor vision (e.g., about 95% or less of normal vision, e.g., about 90% or less, about 85% or less, about 80% or less, about 75% or less, about 70% or less, about 69% or less, about 68% or less, about 67% or less, about 66% or less, about 65% or less, about 64% or less, about 63% or less, about 62% or less, about 61% or less, about 60% or less, about 59% or less, about 58% or less, about 57% or less, about 56% or less, about 55% or less, about 54% or less, about 53% or less, about 52% or less, about 51% or less, about 50% or less, about 49% or less, about 48% or less, about 47% or less, about 46% or less, about 45% or less, about 44% or less, about 43% or less, about 42% or less, about 41% or less, about 40% or less, about 39% or less, about 38% or less, about 37% or less, about 36% or less, about 35% or less, about 34% or less, about 33% or less, about 32% or less, about 31% or less, about 30% or less, about 25% or less, about 20% or less, about 15% or less, about 10% or less, or about 5% or less, preferably about 44% or less of normal vision), · Central pupillary disorder, · The presence of one or more retinal hemorrhages, e.g., one or more intraretinal hemorrhages (e.g., two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, ten or more, eleven or more, twelve or more, thirteen or more, fourteen or more, fifteen or more, sixteen or more, seventeen or more, eighteen or more, nineteen or more, twenty or more retinal hemorrhages), · The presence of one or more "cotton wool spots" (e.g., two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, ten or more, eleven or more, twelve or more, thirteen or more, fourteen or more, fifteen or more, sixteen or more, seventeen or more, eighteen or more, nineteen or more, twenty or more "cotton wool spots"), · Dilatation of retinal veins, · Tortuosity of retinal veins, · Angiography showing one or more areas of retinal capillary non-perfusion (e.g., two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, ten or more, eleven or more, twelve or more, thirteen or more, fourteen or more, fifteen or more, sixteen or more, seventeen or more, eighteen or more, nineteen or more, twenty or more areas of retinal capillary non-perfusion, preferably ten or more areas of retinal capillary non-perfusion), · In an electrodiagnostic test (ERG), there is a reduction in the amplitude of the b-wave, a reduction in the b-wave:a-wave ratio, a prolongation of the b-wave latency, or the presence of one or more of a reduction in the b-wave:a-wave ratio and a prolongation of the b-wave latency.

[0142] Subgroup of patients with retinal vein occlusion In addition to the aforementioned patients having different subtypes of retinal vein occlusion (e.g., central retinal vein occlusion), patients with retinal vein occlusion can also be identified by different characteristics.

[0143] Approximately 1% to approximately 2% of the red blood cells of patients with retinal vein occlusion are annexin A5 positive (i.e., have monomeric annexin A5 binding sites such as phosphatidylserine on the outer side of the red blood cell membrane), while it is known that approximately 0.7% of the red blood cells of "normal" patients (e.g., patients without retinal vein occlusion) are annexin A5 positive. In comparison, in patients with sickle cell disease, typically approximately 2.84% of the red blood cells are annexin A5 positive. Similarly, typically, in patients with retinal vein occlusion, there is an average number of approximately 400 to approximately 1000 monomeric annexin A5 binding sites on each annexin A5 positive red blood cell, while it is known that in "normal" patients, there is an average number of approximately 280 monomeric annexin A5 binding sites on each annexin A5 positive red blood cell. In comparison, in patients with sickle cell disease, typically, there is an average number of approximately 12,340 monomeric annexin A5 binding sites on each annexin A5 positive red blood cell.

[0144] There can also be a clear difference between subjects with respect to the average number of monomeric annexin A5 binding sites exposed on the surface of vascular endothelial cells.

[0145] Microparticles are submicron vesicles that are shed from various cell types (including, for example, red blood cells and / or vascular endothelial cells) and released into the extracellular environment (Hargett and Bauer, 2013, Pulm. Circ., 3(2):329-340). They can typically be detected in the circulating plasma of a subject. The average number of monomeric annexin A5 binding sites exposed on the surface of the microparticles can also vary between subjects, for example, based on the binding site levels in the red blood cells and / or vascular endothelial cells of the subject involved in microparticle release.

[0146] Thus, in one embodiment, the subject is characterized by having about 1% (or 1.0%) to about 2% cells (such as red blood cells (i.e., erythrocytes) and / or vascular endothelial cells), and / or microparticles that are annexin A5 positive. In some preferred cases, the subject is characterized by having about 1.7% to about 1.9% cells (such as red blood cells and / or vascular endothelial cells), and / or microparticles that are annexin A5 positive.

[0147] Thus, the subject may be characterized by having about 1%, about 1.1%, about 1.2%, about 1.3%, about 1.4%, about 1.5%, about 1.6%, about 1.7%, about 1.8%, or about 1.9%, or about 2% cells (such as red blood cells and / or vascular endothelial cells), and / or microparticles that are annexin A5 positive. In some preferred cases, about 1.8% cells (red blood cells and / or vascular endothelial cells), and / or microparticles are annexin A5 positive.

[0148] Additionally or alternatively, the subject may be characterized by having an average of about 400 to about 1000 monomeric annexin A5 binding sites per cell (such as red blood cells and / or vascular endothelial cells), and / or per microparticle.

[0149] For example, the subject may be characterized by having an average number of monomeric annexin A5 binding sites of about 410 to about 1000, about 420 to about 1000, about 430 to about 1000, about 440 to about 1000, about 450 to about 1000, about 460 to about 1000, about 470 to about 1000, about 480 to about 1000, about 490 to about 1000, about 500 to about 1000, about 510 to about 1000, about 520 to about 1000, about 530 to about 1000, about 540 to about 1000, about 550 to about 1000, about 560 to about 1000, about 570 to about 1000, about 580 to about 1000, about 590 to about 1000, about 600 to about 1000, about 610 to about 1000, about 620 to about 1000, about 630 to about 1000, about 640 to about 1000, about 650 to about 1000, about 660 to about 1000, about 670 to about 1000, about 680 to about 1000, about 690 to about 1000, about 700 to about 1000, about 710 to about 1000, about 720 to about 1000, about 730 to about 1000, about 740 to about 1000, about 750 to about 1000, about 760 to about 1000, about 770 to about 1000, about 780 to about 1000, about 790 to about 1000, about 800 to about 1000, about 810 to about 1000, about 820 to about 1000, about 830 to about 1000, about 840 to about 1000, about 850 to about 1000, about 860 to about 1000, about 870 to about 1000, about 890 to about 1000, about 900 to about 1000, about 910 to about 1000, about 920 to about 1000, about 930 to about 1000, about 940 to about 1000, about 950 to about 1000, about 960 to about 1000, about 970 to about 1000, about 980 to about 1000, about 990 to about 1000 per particle, and / or cells (e.g., red blood cells and / or vascular endothelial cells).

[0150] The subject may preferably be characterized by having an average number of monomeric annexin A5 binding sites of about 700 to about 800 or about 700 to about 740, most preferably about 710 to about 730, per particle, and / or cells (e.g., red blood cells and / or vascular endothelial cells).

[0151] In other words, the subject may be characterized by having, per cell (e.g., erythrocyte and / or vascular endothelial cell) and / or microparticle, an average number of about 410, about 420, about 430, about 440, about 450, about 460, about 470, about 480, about 490, about 500, about 510, about 520, about 530, about 540, about 550, about 560, about 570, about 580, about 590, about 600, about 610, about 620, about 630, about 640, about 650, about 660, about 670, about 680, about 690, about 700, about 710, about 720, about 730, about 740, about 750, about 760, about 770, about 780, about 790, about 800, about 810, about 820, about 830, about 840, about 850, about 860, about 870, about 890, about 900, about 910, about 920, about 930, about 940, about 950, about 960, about 970, about 980, about 990 monomeric annexin A5 binding sites, preferably per cell (e.g., erythrocyte and / or vascular endothelial cell) and / or microparticle, about 720 monomeric annexin A5 binding sites.

[0152] Included is that the binding sites on the cell (e.g., erythrocyte and / or vascular endothelial cell) and / or microparticle are outside the surface membrane (i.e., outer membrane) of the cell or microparticle. Preferably, in the embodiments outlined immediately above, the monomeric annexin A5 binding sites per cell (e.g., erythrocyte and / or vascular endothelial cell) and / or microparticle are on annexin A5-positive cells and / or microparticles.

[0153] For example, methods for identifying the number of annexin A5-positive cells and / or microparticles in a patient by determining the number of annexin A5-positive erythrocytes and / or microparticles in the patient's blood are known to those skilled in the art of molecular biology. Various methods are available that enable counting of cells and / or microparticles based on markers on the surface of the cells and / or microparticles (e.g., flow cytometry, etc.). See, for example, Hargett and Bauer, 2013, Pulm. Circ., 3(2):329-340, the contents of which are incorporated herein by reference. There are also numerous variants of annexin A5 conjugated to fluorescent markers that enable such analysis. A simple example of a possible method, as known to those skilled in the art, is to obtain a sample of the subject's blood, label it with fluorescent annexin A5, and use a flow cytometer to calculate the number of annexin A5-positive cells and / or microparticles. Thus, "annexin A5-positive" includes cells (e.g., erythrocytes, etc.) that can be bound by annexin A5 (particularly monomeric annexin A5).

[0154] In one embodiment, a cell (e.g., an erythrocyte and / or vascular endothelial cell), and / or a microparticle is considered annexin A5-positive if it has 290 or more monomeric annexin A5 binding sites, e.g., 300 or more, 310 or more, 320 or more, 330 or more, 340 or more, 350 or more, 360 or more, 370 or more, 380 or more, 390 or more, or 400 monomeric annexin A5 binding sites.

[0155] However, for the same sample, the average number of monomeric annexin A5 binding sites determined per cell and / or per particle can vary depending on the specific determination method used. Thus, in another embodiment, a cell (e.g., a red blood cell and / or a vascular endothelial cell), and / or a particle is considered annexin A5 positive if it has more than 100% of the average number of annexin A5 binding sites of the corresponding cell (such as a red blood cell and / or a vascular endothelial cell), and / or particle in a healthy subject. For example, the average number of monomeric annexin A5 binding sites per red blood cell characteristic of RVO can be in the range of about 1.4-fold to about 4-fold more (e.g., about 2-fold to about 3-fold more) than the average number of monomeric annexin A5 binding sites per red blood cell in a healthy human.

[0156] One molecule that can be bound by annexin A5 is phosphatidylserine. The monomeric annexin A5 binding site is about 30 to about 60 phosphatidylserine molecules, and the monomeric annexin A5 binding site is an area of about 3,100 angstroms (Å) to about 3,300 Å on the surface membrane of the cell and / or particle. In addition, the monomeric annexin A5 binding site is an area of about 40 to about 60 phospholipid heads on the surface membrane of the cell and / or particle.

[0157] In one embodiment, the term "annexin A5 binding site" refers to a region of about 30 to about 60 phosphatidylserine molecules on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, for example, about 31 to about 59, about 32 to about 58, about 33 to about 57, about 34 to about 56, about 35 to about 55, about 36 to about 54, about 37 to about 53, about 38 to about 52, about 39 to about 51, about 40 to about 50, about 41 to about 49, about 42 to about 48, about 43 to about 47, or about 44 to about 46 phosphatidylserine molecules on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, preferably about 35 to about 45 phosphatidylserine molecules on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, more preferably about 42 to about 57 phosphatidylserine molecules on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, and most preferably about 40 phosphatidylserine molecules on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles.

[0158] In additional or alternative embodiments, the term "annexin A5 binding site" refers to a region of about 3,100 Å to about 3,300 Å on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, for example, about 3110 Å to about 3290 Å, about 3120 Å to about 3280 Å, about 3130 Å to about 3270 Å, about 3140 Å to about 3260 Å, about 3150 Å to about 3250 Å, about 3160 Å to about 3240 Å, about 3170 Å to about 3230 Å, about 3180 Å to about 3220 Å, about 3190 Å to about 3210 Å on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, preferably a region of about 3,150 Å to about 3,250 Å on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles, more preferably a region of about 3,000 Å on the surface membrane of cells (e.g., red blood cells and / or vascular endothelial cells) and / or microparticles.

[0159] In a further embodiment, the term "annexin A5 binding site" refers to from about 40 to about 60 phospholipid heads on a region on the surface membrane of a cell (e.g., a red blood cell and / or a vascular endothelial cell), and / or a microparticle, for example, from about 41 to about 59, from about 42 to about 58, from about 43 to about 57, from about 44 to about 56, from about 45 to about 55, from about 46 to about 54, from about 47 to about 53, from about 48 to about 52, from about 49 to about 51 phospholipid heads on a region on the surface membrane of a cell (e.g., a red blood cell and / or a vascular endothelial cell), and / or a microparticle, preferably from about 45 to about 55 phospholipid heads on a region on the surface membrane of a cell (e.g., a red blood cell and / or a vascular endothelial cell), and / or a microparticle, more preferably about 50 phospholipid heads on a region on the surface membrane of a cell (e.g., a red blood cell and / or a vascular endothelial cell), and / or a microparticle.

[0160] In one embodiment of any of the first, second, third, or fourth aspects of the present invention, the subject is characterized by one or more of the following: a. The subject is about 50 years of age or older, for example, at least 55, 60, 65, 70, 75, or 80 years old (optionally, 90 years of age or younger, for example, about 89 years of age or younger, about 88 years of age or younger, about 87 years of age or younger, about 86 years of age or younger, about 85 years of age or younger, about 84 years of age or younger, about 83 years of age or younger, about 82 years of age or younger, about 81 years of age or younger, about 80 years of age or younger, about 79 years of age or younger, about 78 years of age or younger, about 77 years of age or younger, about 76 years of age or younger, about 75 years of age or younger, about 74 years of age or younger, about 73 years of age or younger, about 72 years of age or younger, about 71 years of age or younger, about 70 years of age or younger, about 69 years of age or younger, about 68 years of age or younger, about 67 years of age or younger, about 66 years of age or younger, about 65 years of age or younger, about 64 years of age or younger, about 63 years of age or younger, about 62 years of age or younger, about 61 years of age or younger, about 60 years of age or younger, about 59 years of age or younger, about 58 years of age or younger, about 57 years of age or younger, about 56 years of age or younger, about 55 years of age or younger, about 54 years of age or younger, about 53 years of age or younger, about 52 years of age or younger, about 51 years of age or younger), b. The subject does not have, or there is no suspicion that the subject has, a cardiovascular disease (such as myocardial infarction, stroke, hypertension, hyperlipidemia, hyperhomocysteinemia, blood coagulation disorder, and / or peripheral arterial disease, and optionally excluding atherosclerosis), or a disease associated with a cardiovascular disease (such as sickle cell disease, glaucoma, diabetes, diabetes mellitus, polycythemia vera, and / or a systemic inflammatory disorder (such as Behcets disease, polyarteritis nodosa, sarcoidosis, Wegener granulomatosis, and / or Goodpasture syndrome), but preferably excluding diabetes and diabetes mellitus). c. The subject does not have a medical history of a cardiovascular disease (such as myocardial infarction, stroke, hypertension, hyperlipidemia, hyperhomocysteinemia, blood coagulation disorder, and / or peripheral arterial disease, and optionally excluding atherosclerosis), or a disease associated with a cardiovascular disease (such as sickle cell disease, glaucoma, diabetes, diabetes mellitus, polycythemia vera, and / or a systemic inflammatory disorder (such as Behcets disease, polyarteritis nodosa, sarcoidosis, Wegener granulomatosis, and / or Goodpasture syndrome), but preferably excluding diabetes and diabetes mellitus). d. The subject does not have a family history of a cardiovascular disease (such as myocardial infarction, stroke, hypertension, hyperlipidemia, hyperhomocysteinemia, blood coagulation disorder, and / or peripheral arterial disease, and optionally excluding atherosclerosis), or a disease associated with a cardiovascular disease (such as sickle cell disease, glaucoma, diabetes, diabetes mellitus, polycythemia vera, and / or a systemic inflammatory disorder (such as Behcets disease, polyarteritis nodosa, sarcoidosis, Wegener granulomatosis, and / or Goodpasture syndrome), but preferably excluding diabetes and diabetes mellitus), and / or e. The subject does not have or is not suspected of having one or more conditions selected from oncological conditions such as solid tumors, lymphomas, and / or blood cancers such as breast cancer, endometrial cancer, cervical cancer, ovarian cancer, lung cancer, head and / or neck cancer, brain cancer, thyroid cancer, esophageal cancer, gastric cancer, colorectal and / or rectal cancer, liver cancer, pancreatic cancer, skin cancer, kidney cancer, bladder cancer, prostate cancer, testicular cancer, bone cancer, lymphoma (e.g., Hodgkin lymphoma and / or non-Hodgkin lymphoma), and / or blood cancer (e.g., acute lymphoblastic leukemia, acute myeloid leukemia, and / or chronic myeloid leukemia).

[0161] The "family history" of the subject can be readily determined by a person of ordinary skill in the art. In one embodiment, the family history includes the medical histories of one or more of the following blood relatives: parents (e.g., mother or father), grandparents (e.g., grandmother or grandfather), siblings (e.g., brother or sister), children (e.g., son or daughter), and / or grandchildren (e.g., grandson or granddaughter).

[0162] In one embodiment, if the subject does not exhibit one or more symptoms of a cardiovascular disease or a disease associated with a cardiovascular disease, the subject does not have or is not suspected of having a cardiovascular disease or a disease associated with a cardiovascular disease. The symptoms of these diseases are known to a person of ordinary skill in the art.

[0163] In one embodiment, the blood coagulation disorder is not a retinal vein occlusion such as central retinal vein occlusion, branch retinal vein occlusion, or hemiretinal vein occlusion.

[0164] Preferably, the subject is characterized as follows: a. being about 50 years of age or older, b. the subject does not have or is not suspected of having one or more conditions selected from hypertension, diabetes, and / or sickle cell disease, c. the subject does not have a medical history of having one or more conditions selected from hypertension, diabetes, and / or sickle cell disease, and d. The subject does not have a family history of having one or more conditions selected from hypertension, diabetes, and / or sickle cell disease.

[0165] In an alternative embodiment, the subject may be (i) 50 years of age or older and / or (ii) have a medical history and / or may have a family history of being suspected of having one or more conditions selected from hypertension, diabetes, and / or sickle cell disease.

[0166] In one embodiment, the subject does not have or is not suspected of having a condition characterized by having increased extracellular annexin A5. Preferably, the increased extracellular annexin A5 is increased when compared to a patient who does not have a condition characterized as having increased extracellular annexin A5. In one embodiment, the condition characterized as having increased extracellular annexin A5 is not retinal vein occlusion and / or a disease associated with retinal vein occlusion.

[0167] It has been demonstrated that a subset of patients with retinal vein occlusion may have a genetic predisposition to form retinal vein occlusion (i.e., have that genetic risk factor). For example, patients with a genetic predisposition may produce red blood cells that are prone to forming blood clots in the veins of the eye, particularly in the retina. The formation of retinal vein occlusion in patients with these genetic predispositions is less likely to be associated with other cardiovascular diseases.

[0168] A person of ordinary skill in the medical arts will know methods for identifying patients who have a genetic predisposition to represent a particular disease such as retinal vein occlusion.

[0169] Accordingly, in one embodiment, the subject has, or is suspected of having, a genetic predisposition to retinal vein occlusion. Preferably, the subject has a genetic predisposition to the formation of a thrombus (or thrombi) in the veins of the eye (particularly the retina). Most preferably, the subject has a genetic predisposition to produce red blood cells that are prone to forming thrombi (or thrombi) in the veins of the eye (particularly the retina). For example, from in vitro bone marrow culture studies in CRVO patients, it is known that approximately 27% of CRVO patients exhibited spontaneous proliferation of erythroid precursors in the absence of any detectable myeloproliferative disorder (Heron et al, 2007, Ophthalmology, 114:2155 - 61). Accordingly, in one embodiment, the subject can be a subject who exhibits spontaneous in vitro proliferation of erythroid precursors from the bone marrow in the absence of any detectable myeloproliferative disorder.

[0170] Typically, the subject is a human subject.

[0171] In one embodiment, the subject (particularly a human subject) has a body weight of about 10 kg to about 110 kg, such as about 20 kg to about 100 kg, about 30 kg to about 90 kg, about 40 kg to about 80 kg, or about 50 kg to about 70 kg. In a preferred embodiment, the subject has a body weight of about 40 kg to about 80 kg. In another preferred embodiment, the subject has a body weight of about 50 kg to about 70 kg. In a more preferred embodiment, the subject has a body weight of about 60 kg. As used in this context, the term "about" can be used herein to mean a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0172] Diseases, disorders, or conditions associated with and / or caused by retinal vein occlusion A variety of diseases, disorders, or conditions are associated with and / or caused by RVO and can also be treated by the present invention.

[0173] Accordingly, in one embodiment, the present invention provides for the prevention or treatment of one or more diseases, disorders, or conditions associated with and / or caused by RVO.

[0174] For example, when a retinal vein is blocked during retinal vein occlusion, blood is not properly drained and there is a risk that it may leak into the retina. If it leaks into the macula, macular edema (ME) can occur. The leakage is exacerbated by the severity of the occlusion, the number of veins involved, and the pressure within them.

[0175] Macular edema (ME) is the accumulation of fluid in the macula, which is the central area of the retina. The retina is the light-sensitive tissue at the back of the eye, and the macula is the part of the retina responsible for sharp, straight-ahead vision. The accumulation of fluid expands and thickens the macula, distorting vision. Typically, it occurs due to an increase in venous and / or capillary pressure that causes vascular permeability, resulting in leakage of fluid and blood into the retina. Visual reduction, decrease, or loss in retinal vein occlusion is often associated with macular edema.

[0176] Accordingly, in one embodiment, the present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of macular edema in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO.

[0177] Other diseases, disorders, or conditions associated with and / or caused by RVO include retinal ischemia, iris neovascularization, retinal neovascularization, neovascular glaucoma, vitreous hemorrhage, rubeosis, and retinal detachment.

[0178] That is, in a separate embodiment, the present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of retinal ischemia in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO, The present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of diabetic iris neovascularization in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO. The present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of retinal neovascularization in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO. The present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of neovascular glaucoma in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO. The present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of vitreous hemorrhage in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO. The present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of rubeosis in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO. The present invention provides a molecule consisting of monomeric annexin A5 protein for use in the prevention or treatment of retinal detachment in a subject, preferably a subject characterized as having, suspected of having, or at risk of having RVO.

[0179] Retinal ischemia is ischemia due to non-perfusion of retinal veins and capillaries caused by retinal vein occlusion.

[0180] Iris neovascularization and retinal neovascularization are the growth of new blood vessels in the eye. These neovascularizations cause an increase in cytokines (e.g., VEGF, etc.), which act in a positive feedback loop to further promote new blood vessel growth.

[0181] One or more of retinal ischemia, iris neovascularization, and retinal neovascularization can cause neovascular glaucoma (i.e., damage to the optic nerve), vitreous hemorrhage (i.e., leakage of blood into the vitreous of the eye), rubeosis (i.e., formation of new blood vessels in the iris), and / or retinal detachment (i.e., the retina detaches from its blood supply) ("Retinal Vein Occlusion (RVO) Guidelines" (supra)).

[0182] Accordingly, the present invention also provides for the prevention or treatment of one or more diseases, disorders, or conditions selected from retinal ischemia, iris neovascularization, retinal neovascularization, neovascular glaucoma, vitreous hemorrhage, rubeosis, and retinal detachment in a subject characterized as preferably having RVO, or suspected of having RVO, or at risk of having RVO.

[0183] In one embodiment, iris neovascularization and / or retinal neovascularization is characterized by an increase in cytokines such as an increase in VEGF. In one embodiment, the retinal vein occlusion is a central retinal vein occlusion, and the disease associated with the retinal vein occlusion is iris neovascularization. In some embodiments, the retinal vein occlusion is a branch retinal vein occlusion, and the disease associated with the retinal vein occlusion is retinal neovascularization. In one embodiment, the retinal vein occlusion is a hemicentral vein occlusion, and the disease associated with the retinal vein occlusion is rubeosis. Preferably, the disease associated with the retinal vein occlusion is macular edema.

[0184] In a fifth aspect, the present invention provides a molecule consisting of a monomeric annexin A5 protein for use in the prevention or treatment of a disease, disorder, or condition associated with and / or caused by RVO in a subject, Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg.

[0185] In an alternative embodiment of the fifth aspect, the present invention provides a method for treating or preventing a disease, disorder, or condition associated with and / or caused by RVO in a subject, the method comprising administering to the subject a therapeutically effective amount of a molecule consisting of monomeric annexin A5 protein, Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg.

[0186] In an alternative embodiment of the fourth aspect, the present invention provides the use of a molecule consisting of monomeric annexin A5 protein in the manufacture of a medicament for the prevention or treatment of a disease, disorder, or condition associated with and / or caused by RVO in a subject, Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg.

[0187] The embodiments of the invention discussed herein with respect to the first, second, third, and fourth aspects of the invention are applicable to the fifth aspect of the invention unless otherwise specified.

[0188] In a sixth aspect, the invention provides a composition comprising a molecule consisting of monomeric annexin A5 protein, Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg, and Optionally, the composition is for use in the prevention or treatment of one or more diseases, disorders, or conditions associated with and / or caused by RVO in a subject.

[0189] In an alternative embodiment of the fifth aspect, the present invention provides a method for treating or preventing one or more diseases, disorders, or conditions associated with and / or caused by RVO in a subject, the method comprising administering a composition comprising a molecule consisting of a therapeutically effective amount of monomeric annexin A5 protein, Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in an amount of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or Optionally, the composition comprises a molecule consisting of monomeric annexin A5 protein in a total amount of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg.

[0190] The embodiments of the first, second, third, fourth, and fifth aspects of the invention discussed herein are applicable to the sixth aspect of the invention unless otherwise stated.

[0191] C. Annexin A5 Molecules consisting of monomeric annexin A5 protein for use in all aspects of the present invention may comprise, consist essentially of, or consist of a protein consisting of the sequence of annexin A5, such as the sequence of human annexin A5.

[0192] To avoid misunderstanding, molecules consisting of monomeric annexin A5 protein are not dimeric annexin A5 (commonly referred to as "diannexin" in the art), nor are they PEGylated annexin A5.

[0193] Preferably, a molecule consisting of a human monomeric annexin A5 protein is a protein consisting of or essentially consisting of a polypeptide molecule having the amino acid sequence of SEQ ID NO: 1, either with or without an N-terminal methionine. Preferably, a molecule consisting of or essentially consisting of a monomeric annexin A5 protein is a recombinant human monomeric annexin A5 protein, for example, a protein encoded by a gene having the sequence of SEQ ID NO: 1, which is recombinantly expressed in a recombinant host cell organism, for example, a recombinant E. coli host, and preferably then the monomeric annexin A5 protein is prepared by recovering and purifying it therefrom.

[0194] The sequence of the protein encoded by the human annexin A5 gene is defined by SEQ ID NO: 1 as follows -

Table 1

[0195] In another embodiment, the monomeric annexin A5 protein may contain, consist essentially of, or consist of a variant or mutant of the annexin A5 sequence, such as the sequence of human annexin A5 (optionally with or without an N-terminal methionine and consisting of the sequence of SEQ ID NO: 1). For example, the sequence of the variant or mutant may differ from the sequence of SEQ ID NO: 1 (either with or without an N-terminal methionine) at any of one or more positions, for example, at positions 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, 110, 120, 130, 140, 150, 160 or more positions, or up to those positions.

[0196] In another embodiment, the monomeric annexin A5 protein can comprise, consist essentially of, or consist of a biologically active fragment of mature annexin A5, such as human annexin A5 (optionally with or without an N-terminal methionine, a protein consisting of the sequence of SEQ ID NO: 1), or a variant or mutant thereof.

[0197] Biologically active fragments of annexin A5 share functional or binding properties with full-length annexin A5. Epitope fragments of annexin A5 bind to monoclonal antibodies that bind to full-length annexin A5. The “activity” of annexin A5 means any binding function performed by the protein, and further examples thereof are further defined below.

[0198] Biologically active fragments of the monomeric annexin A5 protein that can be used in the present invention include, without limitation, at least about 50 consecutive amino acids of an annexin A5 protein, usually at least about 100 consecutive amino acids, at least about 150 consecutive amino acids, at least about 200 consecutive amino acids, at least about 250 consecutive amino acids, at least about 260 consecutive amino acids, at least about 270 consecutive amino acids, at least about 280 consecutive amino acids, at least about 290 consecutive amino acids, at least about 300 consecutive amino acids, at least about 310 consecutive amino acids, and up to at most about 320 consecutive amino acids, including a human annexin A5 protein (optionally with or without an N-terminal methionine, a protein consisting of the sequence of SEQ ID NO: 1) or a modification thereof, and may further include fusion polypeptides known in the art in addition to the provided sequences. The annexin A5 sequence can be derived from any mammalian or avian species, such as primate species, particularly humans, rodents including mice, rats, and hamsters, rabbits, horses, cows, dogs, cats, etc. Particularly important is the human annexin A5 protein (optionally with or without an N-terminal methionine, a protein consisting of the sequence of SEQ ID NO: 1).

[0199] In one embodiment, a biologically active fragment of monomeric annexin A5 protein is a polypeptide molecule consisting of the amino acid sequence of SEQ ID NO: 1, with or without an N-terminal methionine, and is effective in treating retinal vein occlusion or a disease associated with retinal vein occlusion, preferably at least 10% effective, for example, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90% effective.

[0200] In one embodiment, the monomeric annexin A5 protein consists of about 320 or fewer amino acids, for example, about 319 or fewer amino acids, about 318 or fewer amino acids, about 317 or fewer amino acids, about 316 or fewer amino acids, about 315 or fewer amino acids, about 314 or fewer amino acids, about 313 or fewer amino acids, about 312 or fewer amino acids, about 311 or fewer amino acids, about 310 or fewer amino acids, about 305 or fewer amino acids, about 300 or fewer amino acids, about 295 or fewer amino acids, about 290 or fewer amino acids, about 285 or fewer amino acids, about 280 or fewer amino acids, about 270 or fewer amino acids, about 260 or fewer amino acids, or about 250 or fewer amino acids.

[0201] In one embodiment, the recombinant monomeric annexin A5 protein consists of 319 amino acids, for example, about 318 or fewer amino acids, about 317 or fewer amino acids, about 316 or fewer amino acids, about 315 or fewer amino acids, about 314 or fewer amino acids, about 313 or fewer amino acids, about 312 or fewer amino acids, about 311 or fewer amino acids, about 310 or fewer amino acids, about 305 or fewer amino acids, about 300 or fewer amino acids, about 295 or fewer amino acids, about 290 or fewer amino acids, about 285 or fewer amino acids, about 280 or fewer amino acids, about 270 or fewer amino acids, about 260 or fewer amino acids, or about 250 or fewer amino acids.

[0202] The monomeric annexin A5 protein may include one or more of the following: a. A protein consisting essentially of or consisting of the sequence of the human monomeric annexin A5 protein (optionally with or without an N-terminal methionine, a protein consisting of the sequence of SEQ ID NO: 1), b. A recombinant human monomeric annexin A5 protein, c. A mammalian ortholog of the human monomeric annexin A5 protein, d. An allele or genetic variant of a), b), or c), e. A protein consisting of an amino acid sequence that is identical to the human monomeric annexin A5 protein having the sequence defined by SEQ ID NO: 1 with or without an N-terminal methionine and is more than 50%, 60%, 70%, more than 75%, for example, more than 80%, more than 85%, more than 90%, or even more preferably more than 95% or 99% identical, a functional analog or variant of the monomeric annexin A5 protein, f. A biologically active fragment of any of a) - e).

[0203] Molecules consisting of monomeric annexin A5 protein may also be referred to herein as "agents consisting of monomeric annexin A5 protein" or "therapeutic agents consisting of monomeric annexin A5 protein", and may also be referred to as "monomeric annexin A5". The purpose of the term "molecule consisting of monomeric annexin A5 protein" is to exclude molecules that are not monomeric annexin A5, such as PEGylated annexin A5, dimeric monomeric annexin A5 protein (also known as di-annexin), and annexin A5 fusion proteins, particularly annexin A5 fusion proteins that have a higher weight average molecular weight and / or plasma half-life compared to the human monomeric annexin A5 protein, which is a polypeptide molecule consisting of the amino acid sequence of SEQ ID NO: 1 with or without an N-terminal methionine.

[0204] Accordingly, a molecule consisting of monomeric annexin A5 protein is not a PEGylated annexin A5, a dimeric monomeric annexin A5 protein, or an annexin A5 fusion protein. To avoid misunderstanding, in one embodiment, a molecule consisting of monomeric annexin A5 protein is not a "modified annexin" as defined in U.S. Patent No. 9,463,217, the contents of which are hereby incorporated by reference in their entirety.

[0205] In certain embodiments, a functional analog, variant, or mutant of annexin A5 according to the invention is either with or without an N-terminal methionine and is identical to human annexin A5, SEQ ID NO: 1, by 50%, 60%, 70%, more than 75%, for example, more than 80%, 85%, 90%, or even more preferably more than 95% or 99%.

[0206] The percent identity between two amino acid sequences is determined as follows. First, the amino acid sequences are compared, for example, to SEQ ID NO:1 using the BLAST 2 Sequences (Bl2seq) program from the stand-alone version of BLASTZ, which includes BLASTN version 2.0.14 and BLASTP version 2.0.14. This stand-alone version of BLASTZ can be obtained from the website of the U.S. government's National Center for Biotechnology Information (ncbi.nlm.nih.gov). Instructions for using the Bl2seq program can be found in the readme file accompanying BLASTZ. Bl2Seq performs a comparison between two amino acid sequences using the BLASTP algorithm. To compare two amino acid sequences, the options for Bl2seq are set as follows: -i is set to the file containing the first amino acid sequence to be compared (e.g., C:\seq1.txt), -j is set to the file containing the second amino acid sequence to be compared (e.g., C:\seq2.txt), -p is set to blastp, -o is set to any desired file name (e.g., C:\output.txt), and all other options are left at their default settings. For example, the following command can be used to generate an output file containing the comparison between two amino acid sequences: C:\Bl2seq -i c:\seq1.txt -j c:\seq2.txt -p blastp -o c:\output.txt. If the two compared sequences share homology, the specified output file will present the regions of homology as aligned sequences. If the two compared sequences do not share homology, the specified output file will not present aligned sequences. Once aligned, the number of matches is determined by counting the number of positions where identical nucleotide or amino acid residues are present in both sequences.

[0207] The percent identity is determined by dividing the number of matches by the length of the sequence shown in the identified sequence and subsequently multiplying the resulting value by 100. For example, if a certain sequence is compared with the sequence shown in SEQ ID NO: 1 (the length of the sequence shown in SEQ ID NO: 1 is 320) and the number of matches is 288, then that sequence has a percent identity of 90 (i.e., 288÷320×100 = 90) with respect to the sequence shown in SEQ ID NO: 1.

[0208] Accordingly, a functional analog, variant or mutant of annexin A5 can be a protein having an amino acid insertion, deletion or substitution, either conservatively or non-conservatively, at one or more positions, provided that such changes result in a protein whose basic properties for functioning in a manner equivalent to annexin A5 are not significantly changed. In this context, "significantly" means that which one of ordinary skill in the art would say that while the properties of the variant may still be different, they are not apparent as compared to the properties of the original protein.

[0209] "Conservative substitutions" are intended to include combinations such as Gly, Ala; Val, Ile, Leu; Asp, Glu; Asn, Gln; Ser, Thr; Lys, Arg; and Phe, Tyr.

[0210] Such variants and mutants can be made using methods of protein engineering and site-directed mutagenesis well known in the art.

[0211] A functional analog, variant, mutant or biologically active fragment of annexin A5 can optionally consist of a sequence of human annexin A5 (such as SEQ ID NO: 1), with or without an N-terminal methionine, having 50, 40, 30, 20, 10, 5, 4, 3, 2, or no more than 1 additional continuous or non-continuous amino acid, and / or 50, 40, 30, 20, 10, 5, 4, 3, 2, or no more than 1 continuous or non-continuous amino acid deletion, and / or 50, 40, 30, 20, 10, 5, 4, 3, 2, or no more than 1 continuous or non-continuous amino acid substitution.

[0212] D. Subjects for Treatment The subject to be treated according to the present invention can be a human subject or a non - human subject. Preferably, the subject is characterized as having or suspected of having or at risk of having macular edema and / or RVO.

[0213] In a preferred embodiment, the subject is a human subject.

[0214] In another embodiment, the subject is a non - human subject and can be selected from the group consisting of mammalian subjects other than humans, birds, amphibians, and reptiles.

[0215] Mammalian subjects other than humans can be selected from the list consisting of horses (e.g., horses or donkeys), cows (e.g., female cows, buffalo, bison, or yaks), camels, pigs, llamas, alpacas, birds (e.g., ostriches, chickens, geese, ducks, turkeys, quails, or pigeons), sheep, goats, dogs, cats, reptiles (e.g., snakes, lizards, crocodiles, alligators, turtles, or sea turtles), rabbits, amphibians (e.g., frogs or newts), or rodents (e.g., mice, rats, chinchillas, guinea pigs, or squirrels).

[0216] E. Administration Regimen Dosage and Timing: According to all aspects of the present invention, the molecule consisting of monomeric annexin A5 protein is Optionally, at a dosage determined with respect to the weight of the subject, the subject can be administered at a dosage of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the molecule, and / or Optionally, regardless of the subject's weight, as defined above, it can be administered to the subject at a total dose of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg - about 1 mg, or about 0.001 mg - about 0.1 mg.

[0217] (a) Dose per body weight: To avoid misunderstanding, it should be understood that the term "mg / kg body weight" refers to the reference of mg of the molecule consisting of monomeric annexin A5 protein per 1 kg of the subject's body weight.

[0218] Preferred doses can be selected from one or more of about 50 μg / kg - about 100 μg / kg, about 40 μg / kg - about 80 μg / kg, about 30 μg / kg - about 60 μg / kg, about 20 μg / kg - about 40 μg / kg, about 1 μg / kg - 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg - about 10 μg / kg, or about 0.01 μg / kg - about 1 μg / kg, for example, about 0.01 μg / kg, about 0.02 μg / kg, about 0.03 μg / kg, about 0.04 μg / kg, about 0.05 μg / kg, about 0.06 μg / kg, about 0.07 μg / kg, about 0.08 μg / kg, about 0.09 μg / kg, and / or about 0.1 μg / kg.

[0219] In some embodiments, the dose can be selected from one or more of about 0.1 μg / kg, about 0.2 μg / kg, about 0.3 μg / kg, about 0.4 μg / kg, about 0.5 μg / kg, about 0.6 μg / kg, about 0.7 μg / kg, about 0.8 μg / kg, about 0.9 μg / kg, and / or about 1 μg / kg.

[0220] In other embodiments, the dose can be selected from one or more of about 1 μg / kg, about 2 μg / kg, about 3 μg / kg, about 4 μg / kg, about 5 μg / kg, about 6 μg / kg, about 7 μg / kg, about 8 μg / kg, about 9 μg / kg, and / or about 10 μg / kg.

[0221] A pharmaceutical or veterinary composition comprising a molecule consisting of monomeric annexin A5 protein can also be administered as defined above, and the composition is administered such that the amount of the molecule consisting of monomeric annexin A5 protein in the composition is about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg - about 1 mg, or about 0.001 mg - about 0.1 mg of the molecule.

[0222] In one embodiment, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of about 0.00001 mg / kg body weight to about "X" mg / kg body weight, where "X" is about 1 mg / kg, 0.5 mg / kg, 0.4 mg / kg, 0.3 mg / kg, 0.2 mg / kg, 0.1 mg / kg, 0.09 mg / kg, 0.08 mg / kg, 0.07 mg / kg, 0.06 mg / kg, 0.05 mg / kg, 0.04 mg / kg, 0.03 mg / kg, 0.02 mg / kg, 0.01 mg / kg, less than or equal to 0.009 mg / kg body weight, for example, less than or equal to about 0.008 mg / kg body weight, less than or equal to about 0.007 mg / kg body weight, less than or equal to about 0.006 mg / kg body weight, less than or equal to about 0.005 mg / kg body weight, less than or equal to about 0.004 mg / kg body weight, less than or equal to about 0.003 mg / kg body weight, less than or equal to about 0.002 mg / kg body weight, more preferably, less than or equal to about 0.001 mg / kg body weight, less than or equal to 0.0009 mg / kg body weight, for example, less than or equal to about 0.0008 mg / kg body weight, less than or equal to about 0.0007 mg / kg body weight, less than or equal to about 0.0006 mg / kg body weight, less than or equal to about 0.0005 mg / kg body weight, less than or equal to about 0.0004 mg / kg body weight, less than or equal to about 0.0003 mg / kg body weight, less than or equal to about 0.0002 mg / kg body weight, less than or equal to about 0.0001 mg / kg body weight, less than or equal to about 0.00009 mg / kg body weight, less than or equal to about 0.00008 mg / kg body weight, less than or equal to about 0.00007 mg / kg body weight, less than or equal to about 0.00006 mg / kg body weight, less than or equal to about 0.00005 mg / kg body weight, less than or equal to about 0.00004 mg / kg body weight, less than or equal to about 0.00003 mg / kg body weight, or less than or equal to about 0.00002 mg / kg body weight of the molecule consisting of monomeric annexin A5 protein.

[0223] In an additional or alternative embodiment, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of from about "Y" mg / kg body weight to about 0.01 mg / kg body weight, preferably from about "Y" mg / kg body weight to about 1 mg / kg, 0.5 mg / kg, 0.4 mg / kg, 0.3 mg / kg, 0.2 mg / kg, 0.1 mg / kg, 0.09 mg / kg, 0.08 mg / kg, 0.07 mg / kg, 0.06 mg / kg, 0.05 mg / kg, 0.04 mg / kg, 0.03 mg / kg, 0.02 mg / kg, or 0.001 mg / kg body weight, where "Y" is at least about 0.00002 mg / kg body weight, for example, at least about 0.00003 mg / kg body weight, at least about 0.00004 mg / kg body weight, at least about 0.00005 mg / kg body weight, at least about 0.00006 mg / kg body weight, at least about 0.00007 mg / kg body weight, at least about 0.00008 mg / kg body weight, at least about 0.00009 mg / kg body weight, at least about 0.0001 mg / kg body weight, at least about 0.0002 mg / kg body weight, at least about 0.0003 mg / kg body weight, at least about 0.0004 mg / kg body weight, at least about 0.0005 mg / kg body weight, at least about 0.0006 mg / kg body weight, at least about 0.0007 mg / kg body weight, at least about 0.0008 mg / kg body weight, or at least about 0.0009 mg / kg body weight, at least about 0.001 mg / kg body weight, at least about 0.002 mg / kg body weight, at least about 0.003 mg / kg body weight, at least about 0.004 mg / kg body weight, at least about 0.005 mg / kg body weight, at least about 0.006 mg / kg body weight, at least about 0.007 mg / kg body weight, at least about 0.008 mg / kg body weight, or at least about 0.009 mg / kg body weight of a molecule consisting of monomeric annexin A5 protein.

[0224] In an additional or alternative embodiment, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of from about "X" mg / kg body weight to about "Y" mg / kg body weight, where "X" and "Y" are the values defined above.

[0225] In one embodiment, the subject is administered an amount of the molecule selected from about 0.0004 mg / kg body weight to about 0.0002 mg / kg body weight, about 0.0005 mg / kg body weight to about 0.0001 mg / kg body weight, about 0.0006 mg / kg body weight to about 0.00009 mg / kg body weight, about 0.0007 mg / kg body weight to about 0.00008 mg / kg body weight, or about 0.0008 mg / kg body weight to about 0.00007 mg / kg body weight, about 0.0009 mg / kg body weight to about 0.00006 mg / kg body weight, or about 0.001 mg / kg body weight to about 0.00005 mg / kg body weight.

[0226] In another embodiment, the subject is administered the molecule in an amount of about 0.0003 mg / kg body weight to about 0.0002 mg / kg body weight, about 0.0004 mg / kg body weight to about 0.0001 mg / kg body weight, about 0.0005 mg / kg body weight to about 0.00009 mg / kg body weight, about 0.0006 mg / kg body weight to about 0.00008 mg / kg body weight, about 0.0007 mg / kg body weight to about 0.00007 mg / kg body weight, about 0.0008 mg / kg body weight to about 0.00006 mg / kg body weight, about 0.0009 mg / kg body weight to about 0.00005 mg / kg body weight, or about 0.001 mg / kg body weight to about 0.00004 mg / kg body weight.

[0227] In a further embodiment, the molecule is administered to the subject in an amount of from about 0.000257 mg / kg body weight to about 0.000255 mg / kg body weight, from about 0.000258 mg / kg body weight to about 0.000254 mg / kg body weight, from about 0.000259 mg / kg body weight to about 0.000253 mg / kg body weight, from about 0.00026 mg / kg body weight to about 0.000252 mg / kg body weight, from about 0.000261 mg / kg body weight to about 0.000251 mg / kg body weight, from about 0.000262 mg / kg body weight to about 0.00025 mg / kg body weight, from about 0.000263 mg / kg body weight to about 0.000249 mg / kg body weight, from about 0.000264 mg / kg body weight to about 0.000248 mg / kg body weight, from about 0.000265 mg / kg body weight to about 0.000247 mg / kg body weight, from about 0.000266 mg / kg body weight to about 0.000246 mg / kg body weight, from about 0.000267 mg / kg body weight to about 0.000245 mg / kg body weight, from about 0.000268 mg / kg body weight to about 0.000244 mg / kg body weight, from about 0.000269 mg / kg body weight to about 0.000243 mg / kg body weight, or from about 0.00027 mg / kg body weight to about 0.000242 mg / kg body weight.

[0228] In a preferred embodiment, the molecule is administered to the subject in an amount of from about 0.0004 mg / kg body weight to about 0.0002 mg / kg body weight. In another preferred embodiment, the molecule is administered to the subject in an amount of from about 0.0003 mg / kg body weight to about 0.0002 mg / kg body weight. In a more preferred embodiment, the molecule is administered to the subject in an amount of from about 0.000257 mg / kg body weight to about 0.000255 mg / kg body weight. In an even more preferred embodiment, the molecule is administered to the subject in an amount of about 0.0003 mg / kg body weight. In another even more preferred embodiment, the molecule is administered to the subject in an amount of about 0.000256 mg / kg body weight.

[0229] The foregoing dosages per body weight may be particularly suitable for human subjects.

[0230] (b) Total dosage: Additionally or alternatively, the dosage may be determined independently of the subject's body weight.

[0231] Preferred dosages can be a total dosage selected from one or more of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg - about 1 mg, or about 0.001 mg - about 0.1 mg. Optionally, the dosage is about 1 μg, about 2 μg, about 3 μg, about 4 μg, about 5 μg, about 6 μg, about 7 μg, about 8 μg, about 9 μg, and / or 10 μg, about 10 μg, about 20 μg, about 30 μg, about 40 μg, about 50 μg, about 60 μg, about 70 μg, about 80 μg, about 90 μg, and / or about 100 μg, or about 100 μg, about 200 μg, about 300 μg, about 400 μg, about 500 μg, about 600 μg, about 700 μg, about 800 μg, about 900 μg, and / or about 1 mg.

[0232] The aforementioned total dosage can be particularly suitable for human subjects, especially adult human subjects, and / or human subjects having a body weight of about 30 kg - about 200 kg, for example, about 40 kg - about 150 kg, or 50 kg - about 120 kg, for example, about 60 kg - about 100 kg.

[0233] In additional or alternative embodiments, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of about 0.001 mg - about "A" mg, where "A" is about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.9 mg or less, for example, about 0.8 mg or less, about 0.7 mg or less, about 0.6 mg or less, about 0.5 mg or less, about 0.4 mg or less, about 0.3 mg or less, about 0.2 mg or less, more preferably about 0.1 mg or less, about 0.09 mg or less, for example, about 0.08 mg or less, about 0.07 mg or less, about 0.06 mg or less, about 0.05 mg or less, about 0.04 mg or less, about 0.03 mg or less, about 0.02 mg or less, about 0.01 mg or less, about 0.009 mg or less, about 0.008 mg or less, about 0.007 mg or less, about 0.006 mg or less, about 0.005 mg or less, about 0.004 mg or less, about 0.003 mg or less, or about 0.002 mg or less of a molecule consisting of monomeric annexin A5 protein.

[0234] In an additional or alternative embodiment, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of from about "B" mg to about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, e.g., about 1 mg, or about 0.1 mg, where "B" is a molecule consisting of monomeric annexin A5 protein of at least about 0.002 mg, e.g., at least about 0.003 mg, at least about 0.004 mg, at least about 0.005 mg, at least about 0.006 mg, at least about 0.007 mg, at least about 0.008 mg, at least about 0.009 mg, at least about 0.01 mg, at least about 0.02 mg, at least about 0.03 mg, at least about 0.04 mg, at least about 0.05 mg, at least about 0.06 mg, at least about 0.07 mg, at least about 0.08 mg, or at least about 0.09 mg.

[0235] In an additional or alternative embodiment, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of from about "B" mg to about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, e.g., about 1 mg, where "B" is a molecule consisting of monomeric annexin A5 protein of at least about 0.002 mg, e.g., at least about 0.003 mg, at least about 0.004 mg, at least about 0.005 mg, at least about 0.006 mg, at least about 0.007 mg, at least about 0.008 mg, at least about 0.009 mg, at least about 0.01 mg, at least about 0.02 mg, at least about 0.03 mg, at least about 0.04 mg, at least about 0.05 mg, at least about 0.06 mg, at least about 0.07 mg, at least about 0.08 mg, at least about 0.09 mg, at least about 0.1 mg, at least about 0.2 mg, at least about 0.3 mg, at least about 0.4 mg, at least about 0.5 mg, at least about 0.6 mg, at least about 0.7 mg, at least about 0.8 mg, or at least about 0.9 mg.

[0236] In an additional or alternative embodiment, the subject is administered a molecule consisting of monomeric annexin A5 protein in an amount of from about "A" mg to about "B" mg, where "A" and "B" are the values defined above.

[0237] In an alternative embodiment, the subject is administered a molecule of from about 0.03 mg to about 0.02 mg, or from about 0.04 mg to about 0.01 mg, or from about 0.05 mg to about 0.009 mg, or from about 0.06 mg to about 0.008 mg, or from about 0.07 mg to about 0.007 mg, or from about 0.08 mg to about 0.006 mg, or from about 0.09 mg to about 0.005 mg, or from about 0.1 mg to about 0.004 mg.

[0238] In another embodiment, the subject is administered a molecule of from about 0.03 mg to about 0.01 mg, or from about 0.04 mg to about 0.009 mg, or from about 0.05 mg to about 0.008 mg, or from about 0.06 mg to about 0.007 mg, or from about 0.07 mg to about 0.006 mg, or from about 0.08 mg to about 0.005 mg, or from about 0.09 mg to about 0.004 mg, or from about 0.1 mg to about 0.003 mg.

[0239] In a further embodiment, the subject is administered a molecule of from about 0.016 mg to about 0.014 mg, or from about 0.017 mg to about 0.013 mg, or from about 0.018 mg to about 0.012 mg, or from about 0.019 mg to about 0.011 mg, or from about 0.02 mg to about 0.01 mg, or from about 0.021 mg to about 0.0099 mg, or from about 0.022 mg to about 0.0098 mg, or from about 0.023 mg to about 0.0097 mg, or from about 0.024 mg to about 0.0096 mg, or from about 0.025 mg to about 0.0095 mg, or from about 0.026 mg to about 0.0094 mg, or from about 0.027 mg to about 0.0093 mg, or from about 0.028 mg to about 0.0092 mg, or from about 0.029 mg to about 0.0091 mg, or from about 0.03 mg to about 0.009 mg.

[0240] In a preferred embodiment, the subject is administered a molecule of from about 0.03 mg to about 0.02 mg. In another preferred embodiment, the subject is administered a molecule of from about 0.03 mg to about 0.01 mg. In a more preferred embodiment, the subject is administered a molecule of from about 0.016 mg to about 0.014 mg. In a further preferred embodiment, the subject is administered a molecule of from about 0.01 mg to about 0.001 mg. In an even more preferred embodiment, the subject is administered a molecule of about 0.015 mg. In another even more preferred embodiment, the subject is administered a molecule of about 0.02 mg. In an even more preferred embodiment, the subject is administered a molecule of about 0.01 mg.

[0241] The embodiments of the invention discussed herein with respect to all other aspects of the invention are applicable to the sixth aspect of the invention unless otherwise stated.

[0242] Route of administration: In a preferred embodiment, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered distally from the eye. Distal administration has the advantage of avoiding administration of the molecule consisting of monomeric annexin A5 protein to the subject's eye, which can be uncomfortable for the subject. In particular, there are a number of potential complications associated with intravitreal injection (e.g., using established anti-VEGF therapies for the treatment of macular edema and / or RVO), including infectious endophthalmitis, sterile endophthalmitis, and retinal vasculitis, particularly retinal vasculitis due to brolucizumab (Cox et al., 2021 (supra)), and transient intraocular pressure (IOP) spikes (Levin et al., 2021, J Glaucoma, 30:1019-26).

[0243] According to one preferred option, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered systemically. Preferably, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered to the subject's vascular system. Most preferably, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered into the subject's blood.

[0244] In one embodiment, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered by injection, transfusion, or infusion. For example, it is intravenous or intra-arterial (in particular, intravenous injection or intra-arterial injection).

[0245] In a preferred embodiment, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered by infusion.

[0246] In an alternative preferred embodiment, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered by intravenous injection.

[0247] In a more preferred embodiment, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered into the blood of a subject by infusion, for example, by intravenous injection.

[0248] In another embodiment, a molecule consisting of monomeric annexin A5 protein or a composition thereof is administered parenterally, intrathecally, intraperitoneally, intra-articularly, intramuscularly, subcutaneously, or topically (however, as described above, in one embodiment, direct administration to the eye can optionally be avoided).

[0249] In another option, although less preferred, a molecule consisting of monomeric annexin A5 protein can be administered directly to the eye. For example, in a preferred embodiment optionally, a molecule consisting of monomeric annexin A5 protein can be administered to one or both eyes of a subject by intravitreal injection. According to this embodiment, the monomeric annexin A5 protein can be administered to a patient in the form of a sterile intravitreal composition, for example, as defined according to the fourth aspect of the present invention.

[0250] The composition for intravitreal injection can be present, for example, in a sterile syringe in a form that can be injected immediately (i.e., without the need for reconstitution with a carrier or diluent). Preferably, the syringe contains the composition in a volume of about 200 μL or less, preferably about 100 μL or less, and more preferably the volume is about 10 to about 90 μL, about 20 to about 80 μL, about 30 to about 70 μL, about 40 to about 60 μL, or about 50 μL (the term "about" in this context refers to a volume of ±4, 3, 2, or 1 μL of the stated value).

[0251] The composition for intravitreal injection can be provided, for example, in the form of a kit containing the composition and one or more additional components.

[0252] For example, the kit can include the composition provided in a sterile syringe and one or more sterile needles attached thereto or separately included in the kit. Similarly, the uses and methods of various aspects of the present invention can include intravitreal injection of a composition containing a molecule consisting of an effective amount of monomeric annexin A5 protein into one or both eyes of a subject through a sterile needle.

[0253] Preferably, the needle or each needle can be of a gauge selected from gauge 27, 28, 29, or more preferably 30 or 31. The needle or each needle may preferably have a length of about 0.5 to 0.62 inches (12.7 to 15.75 mm). Longer needles can increase the risk of retinal damage if the patient accidentally moves forward during the intravitreal injection procedure.

[0254] In a less preferred embodiment where the composition is administered to a subject by intravitreal injection, the procedure can include one or more pre-injection risk management steps selected from the following: 1) applying a local anesthetic, 2) applying an agent suitable for reducing the risk of bacterial colony formation (povidone iodine eye drops and / or periorbital povidone iodine eyelid preparations may be particularly suitable), 3) inserting a sterile eyelid speculum to separate the eyelids, and 4) optionally, re-applying an agent suitable for reducing the risk of bacterial colony formation (e.g., povidone iodine) immediately above the injection site prior to injection.

[0255] In embodiments where intravitreal injection is performed with local anesthesia, a topical anesthetic eye drop can be conveniently used.

[0256] Any agent suitable for reducing the risk of bacterial colony formation can be used in the eye as part of a pre-injection risk management protocol. Povidone iodine is one preferred option. Povidone iodine has been shown to reduce the risk of bacterial colony formation and endophthalmitis. Prior to intravitreal injection, preferably also prior to introducing a sterile eyelid speculum, application of povidone iodine to the conjunctival surface, eyelids, and eyelashes is recommended. (The eyelid speculum prevents the tip of the needle from touching the eyelids or eyelashes prior to needle insertion.) Povidone iodine can optionally be used in a 5% - 10% solution. According to studies, a 5 percent povidone iodine solution is as effective as 10 percent and has been found to be less irritating to the eye. Optionally, an agent suitable for reducing the risk of bacterial colony formation (e.g., povidone iodine) is reapplied immediately above the injection site prior to injection.

[0257] Antibiotics can be used before, during, and / or after intravitreal injection. Any suitable antibiotic can be used. One exemplary embodiment is a fourth-generation fluoroquinolone.

[0258] In another example, the kit can include one or more additional compositions, each containing one or more antibiotics, for administration to a patient for and / or after intravitreal injection.

[0259] One of ordinary skill in the art will be able to select a suitable intravitreal injection site. For example, in one exemplary embodiment, the patient is instructed to look away from the needle entry site and the injection is placed approximately 3 - approximately 3.5 mm posterior to the limbus (e.g., in the case of an aphakic or pseudophakic eye), or approximately 3.5 mm - approximately 4 mm posterior to the limbus (e.g., in the case of a phakic eye). Injections in the inferotemporal quadrant are common, but any quadrant (e.g., the superotemporal quadrant) may be used.

[0260] In another alternative, a molecule consisting of monomeric annexin A5 protein can be directly administered to one or both retinal veins (e.g., central retinal vein, branch retinal vein, and / or hemiretinal vein, etc.) of the subject's eye, for example, by catheterization.

[0261] Administrative schedule: In light of the disclosure of the present invention and applying the common general knowledge thereto, a person skilled in the art can optionally determine a suitable dosage in the range of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg, and / or about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 to 1.5 mg, 0.001 mg to about 1 mg, or about 0.001 mg to about 0.1 mg of the molecule to achieve the desired therapeutic effect. For example, a suitable dosage will typically provide and maintain an amount of the molecule consisting of monomeric annexin A5 protein appropriate for treating or preventing macular edema and / or retinal vein occlusion, or related conditions. A suitable dosage may be aimed at achieving and / or maintaining the level of annexin A5 protein in the subject's plasma at a level higher than the naturally occurring physiological level of annexin A5 in the plasma.

[0262] In other embodiments, the administration of the dosage is over a long period of time. For example, the daily dosage is delivered over a period of 30 minutes or more, for example, 1 hour or more, 2 hours or more, 3 hours or more, 4 hours or more, 5 hours or more, 6 hours or more, 7 hours or more, 8 hours or more, 9 hours or more, 10 hours or more, 11 hours or more, 12 hours or more, 13 hours or more, 14 hours or more, 15 hours or more, 16 hours or more, 17 hours or more, 18 hours or more, or up to a maximum of 24 hours. In a preferred embodiment, the daily dosage is delivered by infusion over a certain period of time.

[0263] The treatment regimen can include, for example, continuous infusion of annexin A5 protein to the subject, or administration one or more times of a dosage consisting of an amount of a molecule composed of monomeric annexin A5 protein as defined for the foregoing embodiments, for example, once, twice, three times, four or more times a day.

[0264] In some cases, the therapeutically effective dosage is administered to the subject as a gradually increasing concentration (i.e., gradually increasing dosage) of two or more dosages, where (i) all dosages are therapeutically effective dosages, or (ii) dosages below the therapeutic dosage (or two or more dosages below the therapeutic dosage) are administered first, and the therapeutic dosage can be achieved by the above gradual increase. As one non-limiting example for illustration of the gradually increasing concentration (i.e., increase in dosage), the therapeutically effective dosage can be administered daily starting from a dosage below the therapeutic dosage, and each subsequent dosage can increase in a specific increment or a variable increment until the therapeutic dosage is reached, and at the time when the therapeutic dosage is reached, the administration can be discontinued or continued (for example, continuous therapeutic dosage). In some embodiments, the administration of the therapeutically effective dosage can be by continuous infusion, and the dosage can vary over time (for example, increase gradually). In some embodiments, combination therapy is also administered.

[0265] In any case, the physician determines the actual dosage that would be most suitable for any individual patient, which will vary depending on the age, weight, and response of the specific patient. The above dosages are illustrative of average cases. Of course, there can be individual cases where higher or lower dosage ranges are appropriate, and they are within the scope of the present invention.

[0266] For veterinary use, the annexin A5 protein is administered as a formulation that is appropriately acceptable according to normal veterinary practice, and the veterinarian determines the dosage regimen and route of administration that would be most appropriate for a particular animal.

[0267] Treatment duration: The treatment regimen may continue for a therapeutically beneficial period.

[0268] In one embodiment, the treatment can consist of a single administration of the annexin A5 composition.

[0269] In another embodiment, multiple administrations to the subject can be performed. For example, the dosage consisting of the amount of the molecule composed of the monomeric annexin A5 protein defined with respect to the foregoing embodiment can be administered once or more per day, for example, twice or more, three times or more, four times or more, five times or more, six times or more, seven times or more, eight times or more, nine times or more, or ten times or more per day, preferably once per day.

[0270] In one embodiment, the dosage of the molecule composed of the monomeric annexin A5 protein is administered once or more every two days, once or more every three days, once or more every four days, once or more every five days, once or more every six days, once or more every seven days, once or more per week, once or more every two weeks, once or more every three weeks, once or more every four weeks, once or more per month, once or more every two months, once or more every three months, once or more every four months, once or more every five months, once or more every six months, once or more every seven months, once or more every eight months, once or more every nine months, once or more every ten months, once or more every eleven months, once or more every twelve months, once or more every two years, once or more every three years, once or more every four years, once or more every five years, or once or more every ten years.

[0271] In one embodiment, the dosage of the molecule consisting of monomeric annexin A5 protein is administered daily over a period of 1 day or more, for example, continuously for 2 days or more, continuously for 3 days or more, continuously for 4 days or more, continuously for 5 days or more, continuously for 6 days or more, continuously for 7 days or more, continuously for 8 days or more, continuously for 9 days or more, continuously for 10 days or more, continuously for 11 days or more, continuously for 12 days or more, continuously for 13 days or more, continuously for 2 weeks or more, continuously for 3 weeks or more, continuously for 4 weeks or more, continuously for 2 months or more, continuously for 3 months or more, continuously for 4 months or more, continuously for 5 months or more, continuously for 6 months or more, continuously for 7 months or more, continuously for 8 months or more, continuously for 9 months or more, continuously for 10 months or more, continuously for 11 months or more, continuously for 12 months or more, continuously for 15 months or more, continuously for 18 months or more, continuously for 21 months or more, continuously for 24 months or more, continuously for 30 months or more, continuously for 3 years or more, continuously for 42 months or more, continuously for 4 years or more, continuously for 5 years or more, continuously for 6 years or more, continuously for 7 years or more, continuously for 8 years or more, continuously for 9 years or more, continuously for 10 years or more, continuously for 11 years or more, continuously for 12 years or more, continuously for 13 years or more, continuously for 14 years or more, continuously for 15 years or more, continuously for 16 years or more, continuously for 17 years or more, continuously for 18 years or more, continuously for 19 years or more, continuously for 20 years or more, continuously for 25 years or more, continuously for 30 years or more, continuously for 35 years or more, continuously for 40 years or more, or continuously for 50 years or more. In a preferred embodiment, the molecule consisting of monomeric annexin A5 protein is administered daily for 6 days or more continuously. In an alternative preferred embodiment, the molecule consisting of monomeric annexin A5 protein is administered daily for a period of 7 days or more continuously. In a more preferred embodiment, the annexin A5 protein is administered daily for a continuous period of 6 days or 7 days. In the most preferred embodiment, the annexin A5 protein is administered daily for a continuous period of 6 days. In an alternative most preferred embodiment, the annexin A5 protein is administered daily for a continuous period of 7 days.

[0272] In one embodiment, a molecule consisting of monomeric annexin A5 protein is administered more than twice a day for a period of 1 week or more, for example, 2 weeks or more, 3 weeks or more, 4 weeks or more, 2 months or more, 3 months or more, 4 months or more, 5 months or more, 6 months or more, 7 months or more, 8 months or more, 9 months or more, 10 months or more, 11 months or more, 12 months or more, 15 months or more, 18 months or more, 21 months or more, 24 months or more, 30 months or more, 3 years or more, 42 months or more, 4 years or more, 5 years or more, 6 years or more, 7 years or more, 8 years or more, 9 years or more, 10 years or more, 11 years or more, 12 years or more, 13 years or more, 14 years or more, 15 years or more, 16 years or more, 17 years or more, 18 years or more, 19 years or more, 20 years or more, 25 years or more, 30 years or more, 35 years or more, 40 years or more, or 50 years or more.

[0273] In one embodiment, a molecule consisting of monomeric annexin A5 protein is administered once a week for a period of 1 week or more, for example, 2 weeks or more, 3 weeks or more, 4 weeks or more, 2 months or more, 3 months or more, 4 months or more, 5 months or more, 6 months or more, 7 months or more, 8 months or more, 9 months or more, 10 months or more, 11 months or more, 12 months or more, 15 months or more, 18 months or more, 21 months or more, 24 months or more, 30 months or more, 3 years or more, 42 months or more, 4 years or more, 5 years or more, 6 years or more, 7 years or more, 8 years or more, 9 years or more, 10 years or more, 11 years or more, 12 years or more, 13 years or more, 14 years or more, 15 years or more, 16 years or more, 17 years or more, 18 years or more, 19 years or more, 20 years or more, 25 years or more, 30 years or more, 35 years or more, 40 years or more, or 50 years or more.

[0274] In one embodiment, a molecule consisting of monomeric annexin A5 protein is administered more than twice a week for a period of one week or more, for example, two weeks or more, three weeks or more, four weeks or more, two months or more, three months or more, four months or more, five months or more, six months or more, seven months or more, eight months or more, nine months or more, ten months or more, eleven months or more, twelve months or more, fifteen months or more, eighteen months or more, twenty-one months or more, twenty-four months or more, thirty months or more, three years or more, forty-two months or more, four years or more, five years or more, six years or more, seven years or more, eight years or more, nine years or more, ten years or more, eleven years or more, twelve years or more, thirteen years or more, fourteen years or more, fifteen years or more, sixteen years or more, seventeen years or more, eighteen years or more, nineteen years or more, twenty years or more, twenty-five years or more, thirty years or more, thirty-five years or more, forty years or more, or fifty years or more.

[0275] In one embodiment, a molecule consisting of monomeric annexin A5 protein is administered once a month for a period of one month or more, for example, two months or more, three months or more, four months or more, five months or more, six months or more, seven months or more, eight months or more, nine months or more, ten months or more, eleven months or more, twelve months or more, fifteen months or more, eighteen months or more, twenty-one months or more, twenty-four months or more, thirty months or more, three years or more, forty-two months or more, four years or more, five years or more, six years or more, seven years or more, eight years or more, nine years or more, ten years or more, eleven years or more, twelve years or more, thirteen years or more, fourteen years or more, fifteen years or more, sixteen years or more, seventeen years or more, eighteen years or more, nineteen years or more, twenty years or more, twenty-five years or more, thirty years or more, thirty-five years or more, forty years or more, or fifty years or more.

[0276] In one embodiment, a molecule consisting of monomeric annexin A5 protein is administered more than twice a month for a period of one month or more, for example, two months or more, three months or more, four months or more, five months or more, six months or more, seven months or more, eight months or more, nine months or more, ten months or more, eleven months or more, twelve months or more, fifteen months or more, eighteen months or more, twenty-one months or more, twenty-four months or more, thirty months or more, three years or more, forty-two months or more, four years or more, five years or more, six years or more, seven years or more, eight years or more, nine years or more, ten years or more, eleven years or more, twelve years or more, thirteen years or more, fourteen years or more, fifteen years or more, sixteen years or more, seventeen years or more, eighteen years or more, nineteen years or more, twenty years or more, twenty-five years or more, thirty years or more, thirty-five years or more, forty years or more, or fifty years or more.

[0277] Combination therapy: Subjects for treatment according to the present invention can be administered a molecule consisting of monomeric annexin A5 protein in an amount and number of times to provide an overall therapeutic effect. The molecule consisting of monomeric annexin A5 protein A5 can be administered alone (monotherapy) or in combination with other agents (combination therapy), mixed or separately, simultaneously or sequentially. To avoid misunderstanding, other agents are not agents containing the sequence of human annexin A5 or its biologically active fragment, and more specifically, are not dimeric annexin A5 (commonly referred to as "diannexin" in the art), and are not PEGylated annexin A5.

[0278] In the case of combination therapy, the dosage and frequency of administration can, for example, result in an additive or synergistic therapeutic effect. Furthermore, the administration of a molecule consisting of monomeric annexin A5 protein (with or without a second or additional agent) can be used as a primary, for example, as a first treatment regimen, or, for example, as a secondary treatment for subjects having an insufficient response to a previously administered therapy (i.e., a therapy other than a therapy involving annexin A5).

[0279] Thus, in one embodiment, a subject may be treated with a molecular therapy consisting of monomeric annexin A5 protein in addition to one or more therapeutic or prophylactic interventions. The subject can be administered the molecular therapy consisting of monomeric annexin A5 protein in the same composition alone, or in a separate formulation containing it, simultaneously or sequentially with one or more other therapeutic or prophylactic interventions, or separately from one or more additional therapeutic or prophylactic treatments.

[0280] In some aspects of the present invention, one or more other therapeutic or prophylactic interventions, or one or more additional therapeutic or prophylactic treatments, include or consist of one or more treatments selected from the group consisting of anti-VEGF therapy (e.g., anti-VEGF therapy delivered to the eye by intravitreal injection), corticosteroids, NSAIDs, pars plana vitrectomy, retinal laser photocoagulation, or anti-VEGF therapy enhanced with intravitreal corticosteroids and / or retinal laser photocoagulation.

[0281] Exemplary anti-VEGF therapies can include any one or more of ranibizumab, aflibercept, bevacizumab, brolucizumab, and / or faricimab.

[0282] Exemplary corticosteroids can include difluprednate, triamcinolone acetonide, triamcinolone acetonide, and dexamethasone, which can be used as monotherapy and may or may not be combined with immunomodulatory therapy to achieve more rapid resolution of edema.

[0283] Devices and kits for treatment: Pharmaceutical and veterinary compositions comprising a molecule consisting of monomeric annexin A5 protein can be administered using a medical device. The device is designed with features such as portability, storage at room temperature, and ease of use, so that in an emergency situation, it can be removed from medical facilities and other medical equipment and used, for example, by untrained subjects or on-site emergency personnel. The device can include, for example, one or more enclosures for storing a pharmaceutical formulation comprising a molecule consisting of monomeric annexin A5 protein and can be configured to deliver one or more unit doses of the molecule consisting of monomeric annexin A5 protein. The device can be further configured to administer a second agent, either as a single pharmaceutical composition that also includes the molecule consisting of monomeric annexin A5 protein or as two separate compositions.

[0284] Pharmaceutical and veterinary compositions can be administered by syringe. The pharmaceutical or veterinary composition can also be administered using a needleless subcutaneous injection device such as the devices disclosed in U.S. Patent Nos. 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824; or 4,596,556. Examples of well-known implants and modules include U.S. Patent No. 4,487,603, which discloses an implantable microinjection pump for dispensing a drug at a controlled rate; U.S. Patent No. 4,486,194, which discloses a therapeutic device for administering a drug through the skin; U.S. Patent No. 4,447,233, which discloses a drug infusion pump for delivering a drug at an accurate infusion rate; U.S. Patent No. 4,447,224, which discloses a variable flow rate implantable infusion device for continuous drug delivery; U.S. Patent No. 4,439,196, which discloses an osmotic drug delivery system having a multi-chamber compartment; and U.S. Patent No. 4,475,196, which discloses an osmotic drug delivery system. Many other devices, implants, delivery systems, and modules are also known.

[0285] A molecule consisting of a monomeric annexin A5 protein can be provided in a kit. In one embodiment, the kit includes (a) a container containing a composition comprising an annexin A5 protein, and optionally (b) information material. The information material can be descriptive, explanatory, marketing, or other material related to the methods described herein and / or the use of the agent for therapeutic benefit.

[0286] In one embodiment, the kit also includes a second agent for treating a disorder described herein. For example, the kit includes a first container containing a composition comprising a molecule consisting of a monomeric annexin A5 protein, and a second container containing the second agent.

[0287] The information materials of the kit are not limited to its form. In one embodiment, the information materials can include information regarding the production of the compound, the molecular weight, concentration, expiration date, batch, or production location of the compound, etc. In one embodiment, the information materials relate to a method of treating a subject in need thereof as described herein by administering a molecule consisting of monomeric annexin A5 protein at, for example, a suitable dosage, dosage form, or mode of administration (e.g., the dosage, dosage form, or mode of administration described herein). The information can be provided in various formats including printed text, computer-readable materials, video recordings, or audio recordings, or substantial materials such as information providing a link or address to the Internet. In one embodiment, the information materials provide instructions for administering to the subject at a dosage of about 50 μg / kg to about 100 μg / kg, about 40 μg / kg to about 80 μg / kg, about 30 μg / kg to about 60 μg / kg, about 20 μg / kg to about 40 μg / kg, about 1 μg / kg to 30 μg / kg, less than about 20 μg / kg, for example, about 0.01 μg / kg to about 10 μg / kg, or about 0.01 μg / kg to about 1 μg / kg of the subject's body weight, and / or administering a molecule consisting of monomeric annexin A5 protein at a dosage of about 5 mg, about 4 mg, about 3 mg, about 2 mg, less than about 2 mg, for example, about 1 mg, about 0.1 - 1.5 mg, 0.001 mg - 1 mg, or 0.001 mg - about 0.1 mg.

[0288] In addition to the molecule consisting of monomeric annexin A5 protein, the composition in the kit can include other components such as a solvent or buffer, stabilizer, or preservative. The molecule consisting of monomeric annexin A5 protein can be provided in any form, for example, liquid, dry, or lyophilized form, preferably in a substantially pure and / or sterile form. When the molecule consisting of monomeric annexin A5 protein is provided in solution, the solution is preferably an aqueous solution. When the molecule consisting of monomeric annexin A5 protein is provided in dry form, reconstitution is generally by the addition of a suitable solvent. The solvent, for example, sterile water or buffer, can optionally be provided in the kit.

[0289] The kit may include one or more containers for a single or multiple compositions containing the agent. In some embodiments, the kit includes separate containers, partitions, or compartments for the composition and the information material. For example, the composition can be contained in a bottle, vial, or syringe, and the information material can be contained in a plastic sleeve or packet. In other embodiments, the separate elements of the kit are contained within a single undivided container. For example, the composition is contained in a bottle, vial, or syringe with the information material attached in the form of a label. In some embodiments, the kit includes a plurality (e.g., packs) of individual containers, each containing an agent in one or more unit dosage forms (e.g., the dosage forms described herein). The container can include a combined unit dosage, for example, a unit containing both a molecule consisting of monomeric annexin A5 protein and a second agent, for example, in a desired ratio. For example, the kit includes a plurality of syringes, ampoules, foil packets, blister packs, or medical devices, for example, each containing a single combined unit dosage. The containers of the kit can be airtight, waterproof (e.g., impermeable to changes in moisture or evaporation), and / or light-shielding.

[0290] The kit optionally includes a device suitable for administering the composition, such as a syringe or other suitable delivery device. The device may be provided with one or both agents pre-loaded, or may be empty but suitable for loading.

[0291] F. Method for calculating the dosage of annexin A5 In a seventh aspect, the present invention provides a method for calculating the dosage of annexin A5 required to treat a condition such as, for example, macular edema and / or RVO, A. Determining the number of annexin A5-positive cells in a subject, B. Determining the number of annexin A5 binding sites on the annexin-positive cells in the subject, C. Determining the number of annexin A5 molecules required to bind to the annexin A5 binding sites, for example, by multiplying the value determined for A by the value determined for B. It includes a step of determining a dosage from the value determined for D.C.

[0292] The embodiments of the invention considered in this specification are applicable to the seventh aspect of the invention unless otherwise stated.

[0293] In one embodiment, the method further includes determining the molar concentration of the number of molecules required to bind to the annexin A5 binding site from the value determined at C’.C, and further includes determining a dosage for the value determined for D.C’.

[0294] Preferably, D further includes determining a dosage by multiplying the value determined for C’ by the molar mass of annexin A5. Preferably, the molar mass of annexin A5 used is about 35,700 Da. If annexin A5 protein variants with different molecular weights are used, the value used in calculation step D can be adjusted accordingly.

[0295] In one embodiment, annexin A5 is a molecule comprising monomeric annexin A5, for example, a monomeric annexin A5 protein as further defined herein.

[0296] In one embodiment, the method includes E. administering the dosage of annexin A5 to a subject.

[0297] Preferably, the method includes E. administering the dosage of annexin A5 to a subject to treat a condition.

[0298] In one embodiment, the method is an in vitro method. In another embodiment, steps A - D of the method are steps of an in vitro method.

[0299] In one embodiment, the condition is one or more selected from the list consisting of viral hemorrhagic fevers (e.g., Ebola virus viral hemorrhagic fever or dengue virus viral hemorrhagic fever, etc.), cardiovascular conditions (e.g., stroke, atherosclerosis, central retinal vein occlusion, myocardial infarction, stenosis, or restenosis), sickle cell disease, arthritis (e.g., osteoarthritis, etc.), and diabetes. Preferably, the condition is retinal vein occlusion, more preferably central retinal vein occlusion.

[0300] In an eighth aspect, the present invention provides a method for calculating the dosage of annexin A5 required to bind to an annexin A5 binding site in a subject, A. determining the number of annexin A5 positive cells in the subject; B. determining the number of annexin A5 binding sites on annexin positive cells in the subject; C. determining the number of annexin A5 molecules required to bind to the annexin A5 binding site, for example, by multiplying the value determined for A by the value determined for B; D. including determining the dosage from the value determined for C.

[0301] The embodiments of the present invention discussed herein are applicable to the eighth aspect of the present invention unless otherwise stated.

[0302] In one embodiment of the eighth aspect of the present invention, the method E. includes administering to the subject a dosage of annexin A5, preferably for the prevention of the condition.

[0303] Here, the present invention will be described with reference to one or more non-limiting examples.

Examples

[0304] The following examples are included to demonstrate specific embodiments of the invention. The techniques disclosed in the following examples represent techniques that the inventors have discovered to function well in the practice of the invention and thus can be considered to constitute a preferred mode for their implementation. However, those skilled in the art should understand that, in light of the present disclosure, many modifications can be made to the disclosed specific embodiments and still obtain the same or similar results without departing from the spirit and scope of the invention.

[0305] Example 1 Construction and verification of the probe: Purified recombinant monomeric human annexin A5 protein (Annexin A5, ANXA5) was labeled with 800CW, a molecular imaging and contrast agent, to form an ANXA5-800CW agent, where it was determined that approximately 40% of the ANXA5 present was labeled with 800CW (the remaining approximately 60% remained as unlabeled monomeric human annexin A5 protein).

[0306] ANXA5-800CW specifically binds to phosphatidylserine (PS), an important molecule in the etiology of RVO.

[0307] A 66-year-old woman known to have systemic hypertension was examined in the outpatient clinic. The woman had presented with reduced vision in her right eye over the past few weeks, which was determined to be caused by branch retinal vein occlusion (BRVO).

[0308] Using a PS assay performed on blood samples taken before and after systemic administration of ANXA5-800CW, the presence of PS on fresh red blood cells was determined by flow cytometry. These results showed the presence and increased percentage of PS availability compared to the negative control (Figure 1). This increase in PS externalization was observed between all blood sampling time points, indicating stable target availability of ANXA5-800CW.

[0309] Evaluation of the localization of intravenously administered annexin A5 to the retinal vein occlusion site: Next, the possibility of visualizing the binding of ANXA5-800CW at the retinal vein occlusion site was evaluated.

[0310] 1.0 mg of ANXA5-800CW was administered intravenously as a single bolus, and then near-infrared (NIR) fluorescence imaging using a Heidelberg Spectralis camera system was performed at predetermined time points (up to 4 hours after injection).

[0311] Figure 2 shows the results of fluorescein angiography (FA) performed as standard treatment. It shows the exact occlusion site (red arrow) and the damaged retinal area with bleeding (red ellipse). When performing NIR fluorescence imaging, the inventors observed a possible fluorescence signal of ANXA5-800CW in the blood vessels and optic nerve located at the site of the affected retinal area 5 minutes after administration. This intensity appears to increase over time, as shown by the image created at 229 minutes after injection.

[0312] No fluorescence signal could be detected in the surrounding retinal blood vessels identified by FA (see Figure 3). This indicates the absence of clear ANXA5-800CW uptake in blood vessels not affected by BRVO.

[0313] These observations show site-specific accumulation of ANXA5-800CW in retinal blood vessels and imply the binding of annexin A5 at the occlusion site. The inventors interpret these results as demonstrating the effectiveness of systemically administered annexin A5 (including the labeled form ANXA5-800CW) in preferentially accumulating at the exposed PS, its target, at the occlusion site in subjects with RVO.

[0314] Example 2 As discussed above, evidence indicates that multiple inflammatory cytokines, including IL-6, TNF-α, IL-1β, and ICAM-1, act to disrupt the integrity of the BRB and can contribute to the development of macular edema and / or retinal vein occlusion (Haydinger et al, 2023 (supra), Park et al., 2019 (supra), Noma et al., 2020 (supra)).

[0315] Here, the inventors report experiments on the potential of annexin A5 to block the effects of such inflammatory cytokines.

[0316] Methods: Chondrocytes were cultured with IL-1β (10 ng / mL) or TNF-α (10 ng / mL) with or without purified recombinant monomeric human annexin A5 (ANXA5) (100 ng / mL, 5 μg / mL, 20 μg / mL). Cells were harvested at 3, 6, 12, and 18 hours. Cell viability was determined by flow cytometry using propidium iodide (PI).

[0317] Expression of IL-6 was measured in IL-1β-stimulated cells. Cells were stimulated with 10 ng of IL-1β and treated with different concentrations of ANXA5 (0.1 μg, 5 μg, and 20 μg / mL) of ANXA5 for 6 hours. Gene expression was measured by qPCR using a SYBR green reaction kit. Gene expression was normalized to GAPDH, and the relative expression of the gene was measured using 2ΔΔCt.

[0318] Results: The results indicate that treatment with annexin A5 reduces cell death in this ex vivo study.

[0319] Figure 4 shows that cell death promoted by the application of TNFα in a human model is inhibited by annexin A5, and the viability of annexin A5-treated cells is similar to that of controls without TNFα treatment.

[0320] As shown in Figure 5, similar results are seen with IL-1β. Annexin A5 has been shown to inhibit cell death in a dose-dependent manner, and treatment at doses of 5 μg and 20 μg results in cell survival rates comparable to controls without IL-1β treatment. In addition, Figure 5 shows that treatment with annexin A5 protein alone does not affect cell survival rates when compared to controls without IL-1β treatment.

[0321] In addition to inhibiting cell death, treatment with annexin A5 also reduces important biomarkers.

[0322] As shown in Figure 6, treatment with annexin A5 reduces the expression of ICAM1 when compared to cells in which cell death and inflammation are promoted using TNFα and controls not treated with TNFα.

[0323] As shown in Figure 7, treatment with annexin A5 reduces the expression of IL-6 in a dose-dependent manner when compared to cells induced with IL-1β and controls not treated with IL-1β. In addition, Figure 7 shows that treatment with annexin A5 alone does not affect IL-6 expression.

[0324] Conclusion: Annexin A5 has been shown to inhibit the production and / or effects of multiple inflammatory cytokines, including IL-6, TNF-α, IL-1β, and ICAM-1, which are involved in the dysregulation of the integrity of the BRB and contribute to the development of macular edema and / or retinal vein occlusion.

[0325] Example 3 The inventors performed new calculations based on information not previously known and / or ignored in the prior art to determine the therapeutically effective dosage of monomeric annexin A5 protein for the treatment of retinal vein occlusions such as CRVO.

[0326] The calculations of the present inventors are based on various relevant parameters regarding the binding effect of monomeric annexin A5 and the pathological characteristics of retinal vein occlusion. By using these parameters, the present inventors calculated the dosage of monomeric annexin A5 that may be effective in treating retinal vein occlusion.

[0327] The calculations incorporate the following information: · The molecular weight of monomeric annexin A5 is 35.7 kilodaltons (kDa), and thus, 1 nanomole (nm) of monomeric annexin A5 is 0.036 micrograms per milliliter (μg / ml); · Under physiological conditions, a single monomeric annexin A5 protein binds to an area of approximately 3200 Å on the surface of red blood cells; in other words, a single monomeric annexin A5 protein binds to an area of approximately 50 phospholipid (「PL」) heads in the red blood cell membrane, and a single monomeric annexin A5 protein binds to approximately 40 phosphatidylserine (「PS」) molecules on the surface of red blood cells; · Approximately 1.8 ± (plus / minus) 0.1% of red blood cells in patients with retinal vein occlusion (especially central retinal vein occlusion) are annexin A5 positive, whereas approximately 0.7 ± (plus / minus) 0.1% of red blood cells in normal patients are annexin A5 positive; · Each of these annexin A5-positive red blood cells in patients with retinal vein occlusion has approximately 720 monomeric annexin A5 binding sites, whereas in normal patients, there are approximately 280 monomeric annexin binding sites per annexin A5-positive red blood cell; · In a 60 kilogram (kg) human patient, there is approximately 4 liters (L) of blood; · Typically, there are 5 × 10 12 red blood cells per liter of blood; · Since the amount of endogenous annexin A5 produced by the body is very low, its contribution to the binding to annexin A5 binding sites on annexin A5-positive red blood cells can be ignored; Typically, patients with CRVO do not have another disease (such as sickle cell disease, etc.) that results in an increase in the annexin A5 binding site.

[0328] To calculate the optimal dose of monomeric annexin A5 for patients with retinal vein occlusion, the inventors performed the following calculations: I. Calculate the number of annexin A5-positive red blood cells in the blood of a patient with retinal vein occlusion: (Percentage of annexin A5-positive red blood cells × Number of red blood cells) ÷ 100 = Number of annexin A5-positive red blood cells (AI); II. Calculate the number of monomeric annexin A5 protein molecules required to cover the annexin A5 binding sites on annexin A5-positive red blood cells in the blood of a patient with retinal vein occlusion: Number of annexin A5 binding sites on each annexin A5-positive red blood cell × Number of annexin A5-positive red blood cells (AI) = Number of monomeric annexin A5 protein molecules (AII); III. Calculate the number of moles (mole, M) of monomeric annexin A5 required to cover the annexin A5 binding sites on annexin A5-positive red blood cells in the blood of a patient with retinal vein occlusion: Number of annexin A5 protein molecules (AII) ÷ Avogadro's constant (6.022 × 10 23 ) = Moles of annexin A5 (AIII); and IV. Calculate the amount of annexin A5 required to cover the annexin A5 binding sites on annexin A5-positive red blood cells in the blood of a patient with retinal vein occlusion: Moles of annexin A5 (AIII) × Molar mass = Amount.

[0329] The information known to the applicant can be applied to the calculations as follows: · The calculation is based on the patient having an approximate world average human body weight of about 60 kg, having approximately 4 L of blood, and about 5 × 10 12 red blood cells per liter (i.e., a total of about 20 × 10 12It is based on the determination that it has (a certain number of red blood cells), and about 1.8% of the red blood cells in patients with CRVO are positive for annexin A5. Also, on average, it is based on the finding that there are about 720 monomeric annexin A5 binding sites per red blood cell in patients with retinal vein occlusion.

[0330] I. (1.8×20×10 12 )÷100 = 0.36×10 12 Annexin A5-positive red blood cells (AI) in the blood of a typical 60 kg patient with retinal vein occlusion; II. 720×0.36×10 12 = 259.2×10 12 Monomeric annexin A5 protein molecules (AII) required to cover the annexin A5 binding sites on annexin A5-positive red blood cells in the blood of a typical 60 kg patient with retinal vein occlusion; III. 259.2×10 12 ÷6.022×10 23 = 43.04218×10 -11 Moles of monomeric annexin A5 (AIII) required to cover the annexin A5 binding sites on annexin A5-positive red blood cells in the blood of a typical 60 kg patient with retinal vein occlusion; and IV. 43×10 -11 ×35,700 = 1,536,605×10 -11 Effective dose to cover the annexin A5 binding sites on annexin A5-positive red blood cells in the blood of a typical 60 kg patient with retinal vein occlusion.

[0331] This value is equal to a single dose of 0.01537 mg (i.e., 15.37 μg) of monomeric annexin A5. For practical purposes, the dose can be conveniently rounded up to a single therapeutic or prophylactic dose of 0.02 mg (i.e., 20 μg) of monomeric annexin A5.

[0332] 1,536,605×10 -11The value of g can also be converted to a numerical value with the unit of mg / kg body weight, which is 0.000256 mg / kg body weight (i.e., 0.256 μg / kg body weight) (in relation to considering the calculation is based on a 60 kg patient, the amount per 1 kg of the patient's weight can be used similarly in patients with other body weights and accordingly for the total amount calculated). For practical purposes, the dose can be rounded up to a maximum of 0.0003 mg / kg body weight (i.e., 0.3 μg).

[0333] Example 4 The following is an exemplary regimen for administering monomeric annexin A5 to patients with retinal vein occlusion: · Route of administration: injection or intravenous; · Dosage frequency: once a day; · Treatment period: 6 - 7 days; · Dosage per administration: The total dosage is in the range of 0.015 mg - 0.02 mg (i.e., a dosage of 15 - 20 μg), and / or is adjusted within the range of 0.000256 mg / kg body weight - 0.0003 mg / kg (i.e., 0.256 - 0.3 μg / kg) body weight considering the patient's weight.

[0334] In fact, the prophylactic and / or therapeutically effective dosage of monomeric annexin A5 can be increased or decreased from the values exemplified above, for example, depending on, for example, one or both of the following ranges, depending on, for example, RVO patients and / or specific characteristics of the selected monomeric annexin A5 protein: i. about 0.01 μg / kg body weight to about 10 μg / kg body weight, more preferably about 0.01 μg / kg body weight to about 1 μg / kg body weight, and / or ii. a total dosage of about 1 μg to about 1 mg, more preferably about 1 μg to about 100 μg.

[0335] For example, a patient may have a different total body weight than the exemplified subject, may have a greater or lesser total blood volume (e.g., more or less than a typical 60 kg patient having a total blood volume of about 4 L), may be characterized as having an atypical percentage of annexin A5-positive red blood cells (compared to the typical 1.8% ± 0.1% in most CRVO patients), may have an atypical number of annexin A5 binding sites per annexin A5-positive red blood cell (wherein it is typical for CRVO patients to exhibit about 720 sites per cell), may have a greater or lesser number of red blood cells per liter of blood than typical (i.e., the typical value is about 5×10 12 erythrocytes per liter), may contain higher or lower levels of endogenous annexin A5 protein than a typical patient, and / or may have a greater or lesser number of annexin A5 binding sites in non-erythrocyte cell types.

[0336] In addition, the selected monomeric annexin A5 protein (which may be a biologically active fragment of human annexin A5, for example) may be of a different molecular weight (compared to the naturally occurring protein having an MW of 35.7 kDa), and / or may have an altered ability to bind to annexin A5 binding sites on annexin A5-positive red blood cells.

[0337] Variations in any one or more of the foregoing properties or characteristics of the patient and / or the selected monomeric annexin A5 protein may affect the amount of monomeric annexin A5 protein for use in the prevention and / or treatment of one or more diseases, disorders, or conditions associated with and / or caused by retinal vein occlusion (RVO) and / or RVO.

[0338] It is contemplated that any of the uses, methods, or compositions described herein can be practiced with respect to any other use, method, or composition described herein.

[0339] The use of the word "a" or "an" when used in combination with the term "comprising" in the claims and / or this specification can mean "one", but is also consistent with the meaning of "one or more", "at least one", and "one or two or more".

[0340] Unless otherwise specified, as used herein, the term "about" can be used herein to mean within a range of ±50%, ±40%, ±30%, ±20%, ±10%, ±5%, ±4%, ±3%, ±2%, or ±1% of the recited value.

[0341] However, it should be understood that the above description, while showing various embodiments of the present invention and numerous specific details thereof, is given by way of illustration and not limitation. Without departing from the spirit of the present invention, many substitutions, modifications, additions, and / or rearrangements may be made within the scope of the present invention, and the present invention includes all such substitutions, modifications, additions, and / or rearrangements.

[0342] All compositions and / or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. Although the compositions and methods of the present invention have been described in terms of preferred embodiments, it will be apparent to those skilled in the art that changes can be made in the compositions and / or methods, and in the steps or the order of the steps of the methods, described herein without departing from the concept, spirit, and scope of the present invention. More specifically, it will be apparent that the same or similar results can be achieved by using certain chemically or physiologically related agents in place of the agents described herein. All such similar substitutions and modifications that are apparent to those skilled in the art are considered to be within the spirit, scope, and concept of the present invention as defined by the appended claims.

Claims

1. A pharmaceutical agent comprising a molecule consisting of monomeric annexin A5 protein, for use in the prevention or treatment of macular edema or related conditions in a subject.

2. The pharmaceutical product according to claim 1, wherein the dose of the molecule comprising the monomer annexin A5 protein is administered systemically to the subject, preferably intravenously.

3. The pharmaceutical product according to claim 1, wherein the dose of the molecule comprising the monomeric annexin A5 protein is administered to the subject daily for two or more consecutive days.

4. The pharmaceutical product according to claim 2, wherein the dose of the molecule comprising monomeric annexin A5 protein is administered to the subject daily for two or more consecutive days.

5. The pharmaceutical product according to claim 3, wherein the dose of the molecule comprising the monomeric annexin A5 protein is administered to the subject daily for 3, 4, 5, 6, or 7 days or more consecutively.

6. The pharmaceutical product according to claim 4, wherein the dose of the molecule comprising monomeric annexin A5 protein is administered to the subject daily for 3, 4, 5, 6, or 7 days or more consecutively.

7. The daily dose of the molecule consisting of the monomer annexin A5 protein administered to the subject is a) A total daily dose of approximately 5 mg, approximately 4 mg, approximately 3 mg, approximately 2 mg, or less than approximately 2 mg, b) The pharmaceutical product according to any one of claims 1 to 4, wherein the amount is within a range selected from the group consisting of approximately 50 to 100 μg, approximately 40 to 80 μg, approximately 30 to 60 μg, approximately 20 to 40 μg, or less than approximately 20 μg per kg of body weight of the subject.

8. The daily dose of the molecule consisting of the monomer annexin A5 protein administered to the subject is a) A total daily dose of 0.1 to 1.5 mg, or b) The pharmaceutical product according to any one of claims 1 to 4, wherein the amount is in the range of 1 μg to 30 μg per kg of body weight of the subject.

9. The pharmaceutical product according to claim 1, wherein the subject has, is suspected to have, or is at risk of having retinal vein occlusion (RVO).

10. A pharmaceutical agent comprising a molecule consisting of monomeric annexin A5 protein, for use in the prevention or treatment of retinal vein occlusion (RVO) or related conditions in a subject, A pharmaceutical product comprising a molecule consisting of the monomeric annexin A5 protein, administered systemically to the subject daily for two or more consecutive days.

11. The pharmaceutical product according to claim 10, wherein the aforementioned dose of the molecule comprising the monomeric annexin A5 protein is administered intravenously to the subject daily for two or more consecutive days.

12. The pharmaceutical product according to claim 10, wherein the dose of the molecule comprising the monomer annexin A5 protein is administered systemically, preferably intravenously, to the subject daily for 3, 4, 5, 6, or 7 days or more.

13. The pharmaceutical product according to claim 11, wherein the dose of the molecule comprising monomeric annexin A5 protein is administered systemically, preferably intravenously, to the subject daily for 3, 4, 5, 6, or 7 days or more.

14. The daily dose of the molecule consisting of the monomer annexin A5 protein, administered systemically, preferably intravenously, to the subject, a) A total daily dose of approximately 5 mg, approximately 4 mg, approximately 3 mg, approximately 2 mg, or less than approximately 2 mg, b) The pharmaceutical product according to any one of claims 10 to 13, wherein the amount is within a range selected from the group consisting of approximately 50 to 100 μg, approximately 40 to 80 μg, approximately 30 to 60 μg, approximately 20 to 40 μg, or less than approximately 20 μg per kg of body weight of the subject.

15. The daily dose of the molecule consisting of the monomer annexin A5 protein, administered systemically, preferably intravenously, to the subject, a) A total daily dose of 0.1 to 1.5 mg, or b) The pharmaceutical product according to any one of claims 10 to 13, wherein the amount is in the range of 1 μg to 30 μg per kg of body weight of the subject.

16. The use described above is for the prevention or treatment of a condition in a subject who has, is suspected of having, or is at risk of having retinal vein occlusion (RVO), The pharmaceutical product according to any one of claims 10 to 13, wherein the aforementioned condition is selected from the group consisting of macular edema, retinal ischemia, iris neovascularization (optionally, iris neovascularization characterized by an increase in cytokines such as an increase in VEGF), retinal neovascularization (optionally, retinal neovascularization characterized by an increase in cytokines such as an increase in VEGF), neovascular glaucoma, vitreous hemorrhage, rubeosis, and retinal detachment.

17. The pharmaceutical product according to any one of claims 1 to 6 or 9 to 13, wherein the subject is a human adult and / or has a body weight of about 50 kg to about 150 kg.

18. The pharmaceutical product according to any one of claims 9 to 13, wherein the RVO is selected from central retinal vein occlusion (CRVO), branch retinal vein occlusion (BRVO), or hemiretinal vein occlusion (HRVO).

19. The monomeric annexin A5 protein, a. A protein that is essentially derived from, or composed of, the sequence of human monomeric annexin A5 protein (optionally, a protein consisting of the sequence of SEQ ID NO: 1, with or without N-terminal methionine), b. Recombinant human monomer annexin A5 protein, c. Mammalian orthologues of human monomeric annexin A5 protein, d. Alleles or genetic variants of a), b), or c), e. A functional analog or variant of monomeric annexin A5 protein, wherein the amino acid sequence is identical by more than 50%, 60%, 70%, 75%, for example, more than 80%, 85%, more than 90%, or more more than 95% or 99%, to the human monomeric annexin A5 protein having the sequence defined by SEQ ID NO: 1, with or without an N-terminal methionine. f. A pharmaceutical product according to any one of claims 1 to 6 or 9 to 13, selected from one or more biologically active fragments among a) to e).

20. A composition comprising a molecule consisting of monomeric annexin A5 protein, for use in the prevention or treatment of macular edema or related conditions in a subject, or for use in the prevention or treatment of retinal vein occlusion (RVO) or related conditions in a subject.

21. The composition according to claim 20, wherein the composition is a unit dose composition.

22. The unit dose composition is a) A total dose of approximately 5 mg, approximately 4 mg, approximately 3 mg, approximately 2 mg, or less than approximately 2 mg, b) Total dose of 0.1 to 1.5 mg The composition according to claim 21, comprising:

23. The composition according to claim 20, wherein the composition is a pharmaceutical composition.

24. The composition according to claim 20, wherein the composition further comprises one or more pharmaceutically acceptable carriers.

25. The composition according to claim 24, wherein the composition comprises a molecule consisting of the monomeric annexin A5 protein and one or more pharmaceutically acceptable carriers.

26. A pharmaceutical product according to any one of claims 1 to 6 or 9 to 13, or a composition according to claim 20, wherein the use is as a combination therapy with another drug.

27. The pharmaceutical or composition according to claim 26, wherein the molecule comprising the monomeric annexin A5 protein is used as a first treatment strategy, with or without one or more other agents.

28. The pharmaceutical or composition according to claim 27, wherein the subject is administered a molecule comprising the monomer annexin A5 protein in the same composition as the one or more other drugs, separately from the one or more other drugs, simultaneously with a separate formulation containing the one or more other drugs, or sequentially with the one or more other drugs.

29. The pharmaceutical or composition according to claim 26, wherein the other drug is an anti-vascular endothelial growth factor (anti-VEGF) agent.

30. The pharmaceutically or composition according to claim 29, wherein the molecule comprising the monomeric annexin A5 protein and the anti-VEGF agent are administered together, simultaneously in separate doses, or sequentially in separate doses, and preferably, when the doses are separate and sequential, the interval between sequential doses is 28 days or less, for example, 3 weeks or less, 2 weeks or less, 1 week or less, 6, 5, 4, 3, 2, or 1 day or less, for example, 24, 28, 12, 6, 5, 4, 3, 2, or 1 hour or less.

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