Stable formulations of indocyanine green

Abstract

The present invention relates to an indocyanine green composition having improved stability, particularly improved storage stability. In particular, the present invention relates to a composition comprising indocyanine green and at least one stabilizer selected from ascorbic acid and its salts.

Description

[Technical field]

[0001] The present invention relates to the medical and pharmaceutical technical fields. The present invention relates to an indocyanine green composition having improved stability, in particular improved storage stability. Specifically, the present invention relates to an indocyanine green composition comprising at least one stabilizer selected from ascorbic acid and its salts. The present invention further relates to a method for producing and using the composition. [Background technology]

[0002] Indocyanine green (ICG) is a water-soluble tricarbocyanine fluorescent dye approved by the U.S. Food and Drug Administration for medical diagnostic use to assess liver function, cardiac output, hepatic blood flow, microcirculation in skin flaps, and in ophthalmic angiography. ICG is used for such medical applications because of its high absorbance in the spectral range (750-800 nm) through which human tissue is relatively transparent. Its extremely low toxicity also makes such medical applications possible. After intravenous administration, ICG is rapidly excreted in the bile by the liver and has an effective half-life of approximately 3-4 minutes, depending on liver function.

[0003] The above applications require that ICG be administered in the form of an aqueous solution. ICG is commercially available as a lyophilized powder that can be dissolved in sterile water and injected, often in kits. However, ICG has poor stability in aqueous environments and degrades rapidly, limiting its usefulness in some applications, such as time-sensitive surgical procedures. ICG package inserts vary among vendors, with some stating that the reconstituted ICG solution should be used immediately, others within 10 hours, and others that any unused portion should be discarded.

[0004] Studies have shown that ICG molecules in aqueous media undergo physicochemical changes, such as aggregation, especially at high concentrations. In water, ICG easily forms dimers and oligomers, whereas in human plasma, methanol, and dimethyl sulfoxide (DMSO), it maintains a monomeric state (Non-Patent Document 1). ICG also generates singlet oxygen to form dioxetanes, which can be thermally decomposed to multiple carbonyl compounds (Non-Patent Document 2). These decomposition products can reduce cell viability in vitro.

[0005] Physicochemical changes in ICG are more likely to occur when exposed to light, at high temperatures, and depending on the concentration of ICG itself (Non-Patent Document 3, Non-Patent Document 4, and Non-Patent Document 5). The type and intensity of light irradiated to ICG also affect its degradation. Physicochemical changes rapidly deteriorate the optical properties of ICG, causing discoloration, a decrease in fluorescence intensity, and a shift in absorbance peak. It has been reported that when ICG is stored in water under direct light for 10 hours, the absorbance of the spectral peak decreases by 10% (Non-Patent Document 4).

[0006] To overcome the above shortcomings, many attempts have been made to improve the stability of indocyanine green in aqueous solutions.

[0007] Engel et al. have reported that the chemical decomposition of ICG can be suppressed by adding sodium azide (NaN3), a singlet oxygen suppressor (Non-Patent Document 2).

[0008] Mindt et al. investigated the stability of ICG samples when stored in the dark and in clear vials at various temperatures. Absorbance and fluorescence measurements showed that ICG was stable for 3 days when stored in the dark at 4°C (although 20% of the fluorescence intensity was lost during this period). Based on these studies, it has been proposed that storing reconstituted ICG solutions in the dark at 4°C may allow the use of ICG for periods longer than 1 day (Non-Patent Document 6).

[0009] Other researchers have proposed to encapsulate ICG in nanometer-sized carriers to prevent the ICG molecules from decomposing. For example, CN103301482 (Patent Document 1) discloses a three-layer polypeptide-type ICG-loaded micelle consisting of a polyleucine core, a polylysine intermediate layer, and a polyethylene glycol shell, with indocyanine green dispersed in the core. The indocyanine green is encapsulated in the core-shell structure, which effectively prevents the aggregation of ICG molecules, thereby improving the stability of ICG.

[0010] Kirchherr et al. have disclosed the encapsulation of ICG in a micellar system of Solutol HS15 (polyoxyethylene esters of 12-hydroxystearic acid). The formulation showed reduced aggregation behavior compared to free ICG in aqueous solution and high stability in aqueous solution over a period of 4 weeks (Non-Patent Document 7).

[0011] US 6,944,493 (Patent Document 2) discloses an indocyanine green composition with a high indocyanine green concentration and improved stability. The invention is based on an indocyanine green liposome formulation that is stable for more than 24 hours after reconstitution. All formulations disclosed in the document are reconstituted with a diluent containing ethanol, polysorbate 80, etc.

[0012] WO2016 / 123864 (Patent Document 3) discloses a composition in which ICG is embedded in the lipid membrane of lipid nanoparticles. The fluorescence intensity of the nanoparticles is 4 to 5 times that of free ICG. After storage at 4°C for 0 to 300 days, the nanoparticles have a fluorescence intensity 4 to 100,000 times that of free ICG in an aqueous solution.

[0013] Dedora et al. have described complexation of ICG with highly soluble methyl β-CyD and FDA-approved sulfobutyl ether β-CyD (Captisol®) in aqueous solution. Such complexes reduce ICG aggregation and provide sustained fluorescence enhancement over 24 hours that is more than two-fold higher than free ICG in water. However, ICG complexed with methyl β-CyD has been shown to severely reduce the viability of some fibroblast cells (8).

[0014] The encapsulation or complexation of ICG using the above-mentioned systems may have disadvantages such as poor compatibility with cells or may reduce the ability of ICG molecules to interact with plasma proteins after entering the blood, which may result in a prolonged excretion kinetics from the vascular system.

[0015] WO2020 / 240514 (Patent Document 4) discloses a formulation of indocyanine green containing disodium hydrogen phosphate, sodium dihydrogen phosphate and sodium chloride. After reconstitution with an aqueous diluent, the solution is stable for up to 24 hours.

[0016] US 2010 / 0181535 (Patent Document 5) discloses a method for enhancing the fluorescence intensity of a fluorescent dye by mixing the fluorescent dye with a redox buffer. The fluorescent dye may be selected from rhodamines including indocyanine green and / or carborhodamines and / or oxazine dyes and / or cyanine dyes. The redox buffer comprises at least one reducing agent and / or at least one oxidizing agent and / or at least one reducing-oxidizing agent. Suitable reducing agents may be selected from the group consisting of thiophenol, thionaphthol, phenol sulfide, uric acid, urea, bilirubin, ascorbic acid and / or flavin. Single molecule fluorescence measurement experiments have shown that the addition of 1 mM ascorbic acid to a cyanine dye immobilized on DNA can extend the length of the dark state of the cyanine dye (i.e., the period during which the fluorescent dye is in a non-fluorescent state). On the other hand, the combination of the reducing agent 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox®) and the oxidizing agent methyl viologen in solution with the cyanine dye increased the photostability by up to 2.6 times compared to a solution of the fluorescent dye in phosphate-buffered saline. These experiments were carried out by illuminating the dye with a laser in a fluorescence detection device to activate the signal. The authors state that another advantage of using the described redox buffer is that the shelf life of a fluorescent dye solution containing it is increased by 10% to 20% compared to the same fluorescent dye solution without the redox buffer. However, the long-term storage stability of the composition is not mentioned in the book.

[0017] WO9423646 (Patent Document 6) discloses a storage-stable composition comprising a voltage-sensitive dye (in particular indocyanine green) for use as a contrast agent for optical imaging. The composition contains air-protective agents such as antioxidants and surfactants to inhibit the decomposition of ICG. The air-protective agents may be selected from sodium sulfite, sodium ascorbate, glutathione, dithiothreitol, EDTA, polysorbate 80 and carboxymethylcellulose. Storage away from air and light inhibited the amount of decomposition of ICG in the composition as measured by the decrease in maximum absorbance in the UV-visible spectrum from 600 to 900 nm. The best results reported herein show a 30% decrease in ICG content in 1 hour. When the experimental conditions of this document are reproduced, rapid degradation of the composition is observed, as demonstrated by the experimental results of the present application.

[0018] Although the above prior arts all report improved stability of ICG in aqueous solution, they do not provide an aqueous solution of ICG that is stable for more than 1-3 days. Thus, there is a demand for an aqueous composition of ICG that exhibits stability superior to that of the prior art formulations.

[0019] Some of the prior art formulations are prepared with expensive equipment and / or require long manufacturing process times and / or require special excipients, and some of these formulations contain excipients such as alcohol and polysorbates that make the injection of the ICG composition painful and difficult. [Prior art documents] [Patent documents]

[0020] [Patent Document 1] Chinese Patent No. 103301482 [Patent Document 2] U.S. Patent No. 6,944,493 [Patent Document 3] International Publication No. 2016 / 123864 [Patent Document 4] International Publication No. 2020 / 240514 [Patent Document 5] US Patent Application Publication No. 2010 / 181535 [Patent Document 6] International Publication No. 94 / 23646

Non-licensed documents

[0021]

Non-patent document 1

Non-patent document 2

Non-patent document 3

Non-patent document 4

[0022] In particular, there is a need for aqueous compositions of ICG that are stable at room temperature for periods of one or several months.

[0023] There is also a demand for an ICG composition for intravenous administration that is compatible with target tissues, is non-toxic, and is rapidly metabolically eliminated.

[0024] The present invention aims to provide a storage stable ICG composition that is simple to formulate, has readily available raw materials, and has a sufficient ICG concentration to be used in applications similar to those of prior art ICG formulations that have a short shelf life.

[0025] Surprisingly, it has been found that formulations of ICG containing a compound selected from ascorbic acid or a salt thereof exhibit increased stability compared to prior art compositions.

[0026] The formulations of the present invention contain ICG at concentrations comparable to or greater than those of the prior art. They can be formulated as lyophilized powders or aqueous solutions. They have improved storage stability and shelf life compared to the prior art.

[0027] Another object of the present invention is to provide a formulation which is stable, cost effective and easy to manufacture.It is yet another object of the present invention to provide a formulation which is comfortable to the patient upon administration. [Means for solving the problem]

[0028] The present invention relates to a composition comprising indocyanine green (a) and a compound (b) selected from ascorbic acid, a salt thereof, and a mixture thereof, wherein the amount of indocyanine green content in the composition is reduced by 10% or less when stored at room temperature for one month or more, as measured by HPLC area ratio (%) at 240 nm.

[0029] Advantageously, the content of ICG (Indocyanine Green) in the composition of the invention before storage, measured as HPLC area percentage (%) at 240 nm, is greater than or equal to 90%.

[0030] Preferably, the total amount of impurities in the ICG composition before storage, measured as an area percentage (%) at 240 nm by HPLC, is 10% or less.

[0031] Preferably, the increase in the amount of impurities in the composition of the present invention when stored at room temperature for one month or more, as measured as a change in area ratio (%) at 240 nm by HPLC, is 10% or less.

[0032] The present invention also relates to an aqueous composition comprising at least indocyanine green (a) and compound (b) selected from ascorbic acid, a salt thereof and a mixture thereof, wherein the weight ratio of compound (b) / indocyanine green (a) is in the range of 0.15 to 30, and the concentration of indocyanine green in the aqueous composition is 0.1 to 50 mg / ml, preferably 0.1 to 30 mg / ml.

[0033] Advantageously, the weight ratio of compound (b) to indocyanine green (a) ranges from 0.15 to 25, preferably from 0.15 to 20.

[0034] Advantageously, the weight ratio of compound (b) to indocyanine green (a) ranges from 0.15 to 15.

[0035] According to one embodiment, compound (b) is ascorbic acid.

[0036] According to another embodiment, compound (b) is a salt of ascorbic acid.

[0037] Preferably, according to this embodiment, compound (b) is selected from sodium ascorbate, potassium ascorbate, calcium ascorbate, magnesium ascorbate, and mixtures thereof. Preferably, according to this embodiment, compound (b) is sodium ascorbate.

[0038] Optionally, the indocyanine green composition according to the present invention further comprises at least one compound (c) selected from ethylenediaminetetraacetic acid (EDTA), D-α-tocopherol polyethylene glycol 1000 succinate (vitamin E-TPGS), thiomalic acid, isosorbide, PEG400, PEG1000, thioglycolic acid, thiourea, α and β cyclodextrin, thiolactic acid, reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0039] Preferably, compound (c) is selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0040] Preferably, the weight ratio of compound (c) to indocyanine green (a) ranges from 1:500 to 100:1, preferably from 1:100 to 10:1, more preferably from 1:100 to 4:1.

[0041] According to one embodiment, the composition according to the invention comprises up to 5% by weight of sodium iodide, based on the weight of indocyanine green.

[0042] Preferably, the concentration of indocyanine green in the aqueous composition is 1 to 25 mg / ml.

[0043] According to a preferred embodiment, the aqueous indocyanine green composition according to the present invention is filled in a vial or an ampoule.

[0044] The present invention also relates to an aqueous composition for use as a pharmaceutical or diagnostic agent, comprising indocyanine green (a) and compound (b) selected from ascorbic acid, a salt thereof and a mixture thereof, in which the weight ratio of compound (b) / indocyanine green (a) is 0.15 to 30, and the concentration of indocyanine green in the aqueous composition is 0.1 to 50 mg / ml, preferably 0.1 to 30 mg / ml.

[0045] The present invention also relates to a kit comprising, in separate compartments, at least 1) a lyophilized powder of the composition as defined above and herein, and 2) an aqueous diluent.

[0046] The present invention also relates to a kit that can be used for the preparation of an aqueous composition according to the present invention, comprising in separate compartments an aqueous solution comprising at least 1) indocyanine green (a) and 2) at least one compound (b) selected from ascorbic acid, its salts and mixtures thereof.

[0047] The present invention also relates to a method of diagnosis and / or treatment in a patient comprising the administration, in particular by the intravenous route, of an effective amount of an aqueous composition according to the invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0048] When one or more components or processes / steps are listed in the "..." portion of the phrase "consisting essentially of...", this means that the method or material of the invention may contain other components or processes / steps in addition to the specified components or processes / steps, which do not significantly affect the characteristics or properties of the invention.

[0049] The term "X to Y" is inclusive unless otherwise specified. It means that the range includes all values ​​from X to Y in addition to the values ​​X and Y themselves.

[0050] Reference herein to any method of treatment should be interpreted as a description of the use of the compounds, pharmaceutical compositions and medicaments of the invention in therapeutic or diagnostic methods of treatment of the human (or animal) body.

[0051] (Stability of the indocyanine green composition according to the present invention) The present invention aims to provide indocyanine green compositions that have improved stability compared to prior art compositions.

[0052] The terms "stable composition" and "stability" refer to a composition in which the content of indocyanine green remains stable even after a certain period of storage (e.g., 1 week, 1 month, 2 months, 6 months, 1 year, 2 years, etc.).

[0053] The stability of the composition is evaluated by measuring the change in indocyanine green content in the composition after long-term storage.

[0054] Preferably, the compositions of the present invention are advantageous in that they exhibit substantially no signs of aggregation, decomposition, or precipitation upon extended storage, and substantially no or minimal formation of impurities due to chemical modification of the indocyanine green. The compositions of the present invention are also advantageous in that they do not experience significant discoloration or significant loss in fluorescence intensity or peak absorbance after extended storage.

[0055] In particular, the present invention provides compositions containing indocyanine green that have improved stability compared to currently available formulations of indocyanine green.

[0056] Advantageously, the compositions of the present invention are at least 10 times more stable than currently approved formulations of indocyanine green, more preferably at least 30 times more stable, more preferably at least 50 times more stable, and even more preferably up to 100 times more stable.

[0057] In one embodiment, the composition of the present invention is stable when stored at room temperature for at least 1 month, preferably at least 3 months, more preferably at least 6 months, even more preferably at least 12 months, and most preferably at least 2 years.

[0058] "Room temperature" means room temperature controlled at 20 to 25°C (15 to 30°C with some variation) in the United States Pharmacopoeia, and 15 to 25°C in the European Pharmacopoeia.

[0059] Preferably, the composition of the present invention is stable at room temperature for 1 month to 3 years, preferably 3 months to 3 years, more preferably 1 year to 3 years, and even more preferably 1 year to 2 years.

[0060] Advantageously, the compositions according to the invention are stable for at least 1 month, preferably at least 3 months, more preferably at least 6 months and most preferably at least 2 years when stored in a container that protects against natural light and / or ultraviolet light and / or fluorescent light.

[0061] A container capable of protecting the composition from natural light and / or ultraviolet light and / or fluorescent light is one having walls with a maximum light transmittance of at least 50% at any wavelength. Typically, the container can be an opaque or amber colored container and / or an opaque secondary packaging member (e.g., carton, overwrap, etc.).

[0062] Advantageously, the compositions according to the invention are stable for at least 1 month, preferably at least 3 months, more preferably at least 6 months, and most preferably at least 2 years when stored in a sealed or airtight container.

[0063] The stability of the compositions of the present invention can be assessed visually for color and / or clarity and / or by other analytical techniques.

[0064] Analytical techniques used to assess the stability of the compositions are known to those of skill in the art and include nuclear magnetic resonance (NMR), high performance liquid chromatography (HPLC), size exclusion chromatography (SEC), liquid chromatography mass spectrometry (LC-MS), dynamic light scattering (DLS), differential scanning calorimetry (DSC), UV spectroscopy, Fourier transform infrared spectroscopy (FTIR), or a combination of these techniques.

[0065] Advantageously, the stability of the composition is evaluated by measuring the change in indocyanine green content in the composition before and after long-term storage by HPLC. More advantageously, the ICG content in the composition and the change in said content are evaluated by HPLC area percentage at 240 nm.

[0066] Preferably, the stability of the composition of the present invention corresponds to a decrease in the content of indocyanine green in the composition of 10% or less when the composition is stored at room temperature for one month or more, preferably three months, more preferably six months, even more preferably twelve months, and most preferably two years or more, as measured by HPLC area ratio (%) at 240 nm.

[0067] Preferably, the stability of the composition of the present invention is evidenced by a decrease in the amount of indocyanine green content in the composition, as measured by HPLC area ratio (%) at 240 nm, of 5% or less when the composition is stored at room temperature for one month or more, preferably three months, more preferably six months, even more preferably twelve months, and most preferably two years or more.

[0068] More preferably, the stability of the composition of the present invention is evidenced by a decrease in the amount of indocyanine green content in the composition, as measured by HPLC area ratio (%) at 240 nm, of 2% or less when the composition is stored at room temperature for one month or more, preferably three months, more preferably six months, even more preferably twelve months, and most preferably two years or more.

[0069] "Reduction in content" in the sense of the present application is to be understood as the difference (%) between the measured content of ICG in the composition before storage, measured as an area percentage (%) by HPLC at 240 nm, and the measured content of ICG in the same composition after a given storage period, measured as an area percentage (%) by HPLC at 240 nm.

[0070] Preferably, the content of ICG (indocyanine green) in the composition before storage, measured as an area percentage (%) at 240 nm by HPLC, is 90% or more, preferably 95% or more, more preferably 97% or more, and most preferably 99% or more.

[0071] The stability of the indocyanine green composition of the present invention may also correspond to the total amount of impurities contained in the composition of the present invention not increasing during storage, in particular not increasing to an extent that reduces the ability of the indocyanine green composition to be used in medical and / or diagnostic applications.

[0072] For example, when the composition is stored at room temperature for more than one month and then the impurities are measured by HPLC, only small amounts of impurities are found in the composition, and the amount does not increase significantly over time.

[0073] In the context of the present invention, the term "impurities" should be understood to refer to chemicals that are neither indocyanine green nor excipients or other additives to the indocyanine green composition of the present invention.

[0074] The impurities may be process-related impurities, such as by-products or intermediates that may be formed during the manufacture of the indocyanine green, or decomposition-related impurities that arise due to chemical changes in the indocyanine green during storage, such as products that may be formed during storage due to exposure to light, temperature, humidity, reactions with oxygen, etc.

[0075] In one embodiment, when the composition of the present invention is stored at room temperature for one month or more, preferably two months, more preferably six months, and most preferably two years or more, the increase in the amount of impurities in the composition, as measured as a change in area ratio (%) at 240 nm by HPLC, is 10% or less, preferably 5% or less, and more preferably 2% or less.

[0076] In one embodiment, the total amount of impurities in the ICG composition before storage, measured as HPLC area percentage (%) at 240 nm, is 10% or less, preferably 5% or less, more preferably 1% or less.

[0077] The term "total amount of impurities" includes both process-related impurities and degradation-related impurities as discussed above.

[0078] (Indocyanine Green Composition) The present invention provides a composition comprising indocyanine green (a) and at least one stabilizing compound (b) selected from ascorbic acid and its salts.

[0079] Optionally, the composition according to the invention may contain at least one additional compound (c) different from compound (b).

[0080] (Indocyanine Green (a)) The term indocyanine green refers to indocyanine green and its pharma- ceutically acceptable salts, solvates, hydrates, acid, anhydrate and free base forms, preferably indocyanine green.

[0081] [ka]

[0082] Indocyanine green can be prepared by any method known to those skilled in the art. For example, indocyanine green can be prepared according to the methods described in WO95 / 07888 and WO2017093889. Preferably, the preparation of indocyanine green includes a purification step of the obtained product.

[0083] Advantageously, the indocyanine green used in the composition according to the invention is substantially pure, and is preferably provided as a composition having a purity of more than 90.0%, preferably more than 95.0%, more preferably more than 97.0%, even more preferably more than 99.7%, as measured by HPLC.

[0084] As used herein, the term "purity" refers to the degree to which the raw indocyanine green is free from unwanted or degrading chemicals.

[0085] In the present invention, the purity and impurities of the indocyanine green substance are evaluated by area ratio (%) at 240 nm by HPLC.

[0086] Preferably, the indocyanine green used in the compositions of the present invention is provided in a composition having less than 5.0% impurities, preferably less than 1.0%, more preferably less than 0.5% impurities as measured by HPLC.

[0087] Preferably, the indocyanine green used in the compositions of the present invention is provided in a composition having less than 0.50%, preferably less than 0.40%, more preferably less than 0.20%, and most preferably less than 0.15% of any impurity as measured by HPLC.

[0088] The impurities contained in the indocyanine green may be any reaction by-products or intermediates generated during the manufacturing process. For example, as described in US2019 / 0337896, the impurities may include, but are not limited to, N-phenylacetamide, 4-(1,1,2-trimethyl-1H-benzo[e]indolium-3-yl)butane-1-sulfonate, 4-(1,1,2-dimethyl-2-((1E,3E,5E)-6-(N-phenylacetamide)hexa-1,3,5-trienyl)-1H-benzo[e]indolium-3-yl)butane-1-sulfonate, etc.

[0089] Preferably, the indocyanine green is provided as a composition having a metal contaminant content of less than 200 ppm by weight, preferably less than 100 ppm by weight, more preferably less than 50 ppm by weight, and even more preferably less than 20 ppm by weight of metal based on the weight of indocyanine green. The metal contaminant content in the indocyanine green can be measured by inductively coupled plasma mass spectrometry (ICP-MS).

[0090] "Metal contaminants" is to be understood as meaning the so-called "heavy" metals, in particular Al, As, Cd, Cr, Cu, Fe, Sn, Mn, Hg, Mo, Ni, Pb, Zn and their organic and inorganic derivatives.

[0091] More preferably, the indocyanine green is free of lead and arsenic or contains less than 2 ppm of lead and arsenic.

[0092] The indocyanine green used in the composition of the present invention is provided as a composition which may further contain sodium iodide, an additive which is often present in commercially available indocyanine green compositions. When present, the iodide content is preferably kept below 5% by weight based on the weight of indocyanine green. The sodium iodide content in the indocyanine green can be measured by potentiometry.

[0093] The indocyanine green used in the compositions of the present invention may be provided in any suitable form, such as a lyophilisate or crystalline powder.

[0094] (Stabilizing compound (b)) Compound (b) may be selected from (5R)-5-[(1S)-1,2-dihydroxyethyl]-3,4-dihydroxyfuran-2(5H)-one, 3-oxo-L-gulofuranolactone, ascorbic acid also known as L-ascorbic acid or vitamin C, salts of ascorbic acid and mixtures thereof.

[0095] By "mixtures" is meant mixtures of ascorbic acid and salts of ascorbic acid in all proportions.

[0096] Salts of ascorbic acid can include, but are not limited to, sodium ascorbate, potassium ascorbate, calcium ascorbate, and magnesium ascorbate.

[0097] According to a first embodiment of the invention, compound (b) is ascorbic acid.

[0098] According to a second embodiment of the invention, compound (b) is a salt of ascorbic acid.

[0099] Preferably, according to this embodiment, compound (b) is sodium ascorbate.

[0100] According to a third embodiment, the composition comprises indocyanine green, ascorbic acid, and at least one salt of ascorbic acid, preferably sodium ascorbate.

[0101] The present inventors have found that by associating compound (b) with indocyanine green (a), the stability of the indocyanine green composition in an aqueous medium is improved.

[0102] In a preferred embodiment, the weight ratio of compound (b) to indocyanine green (a) is in the range of 0.15 to 30, preferably 0.15 to 25, preferably 0.15 to 20, preferably 0.15 to 15, more preferably 0.20 to 15, and even more preferably 0.20 to 13.

[0103] In a preferred embodiment, the molar ratio of compound (b) to indocyanine green (a) ranges from 0.30 to 120, preferably from 0.30 to 60, more preferably from 0.40 to 60, and even more preferably from 0.40 to 55.

[0104] In a preferred embodiment of the invention, the composition consists essentially of indocyanine green (a) and at least one compound (b) selected from ascorbic acid, its salts and mixtures thereof.

[0105] In another preferred embodiment of the invention, the composition consists essentially of indocyanine green (a), at least one compound (b) selected from ascorbic acid, its salts and mixtures thereof, and further comprises iodide.

[0106] According to another aspect, the present invention relates to the use of ascorbic acid and / or its salts for improving the storage stability of an ICG composition.

[0107] (Compound (c)) In another embodiment of the invention, the composition may further comprise one or more compounds (c) selected from those known to the skilled artisan to be suitable for medical and / or diagnostic injections.

[0108] If present, compound (c) may be selected, for example, from ethylenediaminetetraacetic acid (EDTA), D-α-tocopherol polyethylene glycol 1000 succinate (vitamin E-TPGS), thiomalic acid, isosorbide, PEG400, PEG1000, thioglycolic acid, thiourea, α and β cyclodextrin, thiolactic acid, reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0109] Preferably, compound (c) may be selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0110] "Mixture" refers to any mixture of two or more of the following in any proportion: reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride-histidine, and dithiothreitol (DTT).

[0111] Preferably, the weight ratio of compound (c) to indocyanine green (a) ranges from 1:500 to 100:1, preferably from 1:500 to 10:1, more preferably from 1:100 to 10:1, most preferably from 1:100 to 4:1.

[0112] Preferably, the weight ratio of compound (c) to compound (b) is in the range of 1:100 to 100:1, preferably 1:10 to 10:1.

[0113] When present, compound (c) may be provided in any form, such as a solid, liquid, semi-solid, or may be in the form of an aqueous solution.

[0114] In a preferred embodiment of the invention, the composition consists essentially of indocyanine green (a), a stabilizing compound (b) selected from ascorbic acid, its salts and mixtures thereof, and a compound (c) selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0115] In another preferred embodiment of the invention, the composition consists essentially of indocyanine green (a), a stabilizing compound (b) selected from ascorbic acid, its salts and mixtures thereof, iodide, and a compound (c) selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0116] (Other additives) The composition of the present invention may further contain one or more pharma- ceutically acceptable excipients.

[0117] As used herein, the term "pharmacologically acceptable" refers to compounds, substances and compositions that are suitable for use in contact with the tissues of humans or animals without excessive toxicity, irritation or other problems or complications commensurate with a reasonable benefit-risk ratio.

[0118] The term "additives" includes components contained in the composition other than components a), b) and c) as defined above or sodium iodide. Examples of such additives include buffers such as phosphate buffers, pH adjusters, tonicity agents, surfactants, preservatives, tonicifying agents, antimicrobial agents, wetting agents, emulsifiers, etc.

[0119] For example, the compositions of the present invention may contain one or more additives selected from the group including: sodium hydroxide; potassium hydroxide; methylparaben; propylparaben; sodium acetate; sodium citrate; sodium carbonate; ammonium carbonate; sodium bicarbonate; benzoates or benzoates; acetates or acetates; boric acid; lactic acid; glutaric acid; malic acid; succinic acid; carbonic acid; alkali or alkaline earth salts of these acids; meglumine; hydrochloric acid; cyclodextrin derivatives; sorbitan monolaurate; triethanolamine acetate; triethanolamine oleate; citric acid; lactic acid; phosphoric acid; sulfuric acid; arginine; alanine; glycine; and lysine.

[0120] These additives are commonly available to one of ordinary skill in the art and may be in any form, such as solid, liquid, or semi-solid.

[0121] Preferably, the amount of additives in the composition of the present invention is within a range that does not significantly adversely affect the activity of the indocyanine green. Preferably, the composition of the present invention contains the minimum number and amount of additives necessary to provide a stable and effective composition.

[0122] (Form of the composition according to the present invention) The compositions of the present invention may be provided in any suitable form, but are preferably provided in the form of an aqueous composition or a lyophilized solid composition.

[0123] (aqueous composition) In a first embodiment, the composition according to the invention is in the form of an aqueous composition.

[0124] An "aqueous composition" is to be understood as meaning an aqueous solution, aqueous suspension, colloidal aqueous suspension, aqueous dispersion, preferably an aqueous solution.

[0125] The term "aqueous solution" refers to the aforementioned indocyanine green composition in which the components are dissolved in an aqueous solvent.

[0126] Advantageously, according to this variant, the composition comprises an aqueous diluent, indocyanine green (a), one or more compounds (b) as defined above, optionally one or more compounds (c) as defined above, and optionally one or more pharma- ceutically acceptable excipients.

[0127] The aqueous diluent is a pharma- ceutically acceptable solvent, safe and non-toxic for administration to humans or animals, and useful for preparing pharmaceutical formulations.

[0128] The aqueous diluent comprises water and may comprise one or more pharma- ceutically acceptable additives. In some variations of the methods disclosed herein, one or more compounds (b) as defined above, and optionally one or more compounds (c) as defined above, are introduced into the aqueous diluent prior to mixing with ICG. In such embodiments, one or more compounds (b), and optionally one or more compounds (c) may be considered as part of the aqueous diluent.

[0129] The suitable aqueous diluent may be any liquid that is biologically acceptable and in which the composition of the present invention can be completely dissolved. Water, particularly sterile water for injection (SWFI) and / or bacteriostatic water for injection (BWFI), is the preferred diluent since it does not contain salts or other compounds that may affect the stability of the ICG. However, other aqueous diluents may be used, such as sterile saline, Ringer's solution, dextrose solution, phosphate buffered saline (PBS) solution, glucose solution, etc. The aqueous diluent may also contain buffers, bacteriostatic agents, and solutes that make the composition isotonic with the blood of the recipient to which it is administered.

[0130] In one embodiment, the aqueous diluent is selected from the group consisting of water for injection, bacteriostatic water for injection, sterile saline, Ringer's solution, dextrose solution, phosphate buffered saline, and glucose solution.

[0131] A person skilled in the art can select the diluent depending on the desired application and in relation to the other compounds of the composition, and can also adjust the amount of the excipient depending on its solubility in the aqueous solution.

[0132] In a preferred embodiment, the aqueous diluent is water for injection, in particular sterile water for injection (SWFI) and / or bacteriostatic water for injection (BWFI).

[0133] Advantageously, the osmotic pressure of the aqueous solution is 1 mOsM or more, more advantageously, the osmotic pressure of the aqueous solution is 1 to 300 mOsM, more preferably, 1 to 200 mOsM.

[0134] The concentration of indocyanine green in the aqueous compositions of the present invention may be any concentration suitable for use in medical treatment or diagnosis, particularly any concentration suitable for producing angiographic images of satisfactory quality.

[0135] In a preferred embodiment, the concentration of indocyanine green in the aqueous composition according to the present invention is 0.1 to 50 mg / ml.

[0136] If the concentration of the ICG solution is less than 0.1 mg / ml, it will not be fluorescent enough for satisfactory detection in analytical instruments.

[0137] In a preferred embodiment, the concentration of indocyanine green in the aqueous composition according to the present invention is 0.1 to 25 mg / ml, preferably 1 to 25 mg / ml, and more preferably 1 to 10 mg / ml.

[0138] When one or more components of the compositions of the invention, such as compound (b) or, optionally, compound (c), are added as an aqueous solution, the amount of aqueous diluent can be adjusted to obtain the desired concentration of ICG.

[0139] In one embodiment of the present invention, the aqueous composition consists essentially of an aqueous diluent, indocyanine green (a), and a stabilizing compound (b) selected from ascorbic acid, salts thereof, and mixtures thereof.

[0140] In one embodiment of the invention, the aqueous composition consists essentially of an aqueous diluent, indocyanine green (a), a stabilizing compound (b) selected from ascorbic acid, its salts and mixtures thereof, and iodide.

[0141] In another embodiment, the aqueous composition consists essentially of an aqueous diluent, indocyanine green (a), a stabilizing compound (b) selected from ascorbic acid, its salts and mixtures thereof, and a compound (c) selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

[0142] In another embodiment, the aqueous composition consists essentially of an aqueous diluent, indocyanine green (a), a stabilizing compound (b) selected from ascorbic acid, its salts and mixtures thereof, and a compound (c) selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof, and iodide.

[0143] The preferred ratios of the compounds (ICG (a), stabilizing compound (b) and compound (c)) in the aqueous composition are the same as those expressed in the section "Indocyanine Green Compositions" above.

[0144] (lyophilized solid composition) In another preferred embodiment, the composition according to the invention is in the form of a lyophilized composition or a lyophilized powder.

[0145] The terms "lyophilized composition", "lyophilized powder" and "lyophilisate" refer to a solid composition according to the invention obtained by lyophilization of an aqueous solution.

[0146] The term "lyophilization" refers to a process in which the composition in an aqueous solvent is first frozen and then the frozen solvent is removed by sublimation in a vacuum environment.

[0147] When the compositions of the present invention are provided as lyophilizates, they can be converted to infusion solutions using the aqueous diluents described above prior to administration, and the reconstituted aqueous solutions (which are also compositions of the present invention) are stable at room temperature for at least one month, preferably at least two months, more preferably at least six months, and most preferably at least two years.

[0148] All the limitations regarding ICG concentration, pH, osmolality and stability mentioned above in relation to the aqueous diluent also apply to the aqueous solution resulting from reconstitution of the lyophilized composition according to the invention.

[0149] In one embodiment, the lyophilized composition of the present invention has a short reconstitution time of less than 2 minutes, preferably less than 1 minute, making it suitable for intravenous administration after dilution with an aqueous diluent.

[0150] By "reconstitution time" is meant the time required for the lyophilized composition to be rehydrated with an aqueous solution to a clear, particle-free solution.

[0151] Once the composition has been reconstituted in an aqueous solution for medical or diagnostic use, it is preferably stored in a container that protects it from light and oxygen.

[0152] (kit) In one preferred embodiment, the invention relates to a kit comprising in a first part the composition according to the invention in the form of a lyophilized powder and in a second part an aqueous diluent.

[0153] According to another preferred embodiment, the present invention relates to a kit comprising an aqueous solution comprising in a first part indocyanine green (a) and in a second part at least a stabilizing compound (b) selected from ascorbic acid and its salts. Optionally, the aqueous solution may also contain a compound (c) as defined above, and optionally one or more additives as explained above.

[0154] According to this embodiment, the indocyanine green is preferably in the form of a lyophilized powder.

[0155] All the preferred embodiments previously described for the ICG composition and the aqueous diluent apply to any of the examples of the kit.

[0156] In one embodiment, the kit of the present invention is composed of each component (powder (power) and diluent) in such amounts that the ICG concentration of an aqueous solution reconstituted from the kit is within the range of 0.1 to 50 mg / ml, preferably 0.1 to 30 mg / ml, more preferably 1 to 25 mg / ml, and most preferably 1 to 10 mg / ml.

[0157] In the above preferred embodiment, the kit comprises at least two separate compartments in which the lyophilized powder and the aqueous diluent are stored separately.

[0158] The kit may consist of two separate vials packaged with or without a syringe, or may consist of a pre-filled syringe with two compartments, such as a dual chamber bypass syringe.

[0159] Such a kit allows for mixing of the powder and the aqueous solution prior to administration.

[0160] Either of the kits allows for the immediate reconstitution of a stable aqueous solution of indocyanine green as described. They have the advantage that each component of the composition required for injection is available in the appropriate amount and under optimal storage conditions to obtain the desired concentration. In other words, the aqueous composition can be reconstituted without the need for dilution with a diluent other than that included in the kit. This packaging as a kit can avoid administration errors and the introduction of contaminants.

[0161] (Method of preparing the composition according to the present invention) The composition according to the present invention may be prepared by any known method and is not limited to any particular method.

[0162] For example, the composition can be prepared by mixing the indocyanine green (a) with a stabilizing compound (b), and optionally one or more compounds (c) described above, and optionally one or more pharma- ceutically acceptable excipients.

[0163] Preferably, the components of the composition of the present invention are mixed in the aqueous solvent described above. The components constituting the composition of the present invention may be added in various orders. For example, indocyanine green may be dissolved in the aqueous diluent, and then stabilizing compound (b), and optionally compound (c) and various additives may be added. Compounds (b) and (c) may be mixed and then added to the dissolved indocyanine green. Alternatively, indocyanine green may be added to an aqueous solution in which compound (b) and optionally compound(s) (c) have been dissolved in advance.

[0164] Preferably, mixing is carried out under an inert atmosphere, such as under an argon or nitrogen atmosphere.The aqueous diluent and / or liquid components are preferably degassed prior to use.

[0165] When the composition of the present invention is provided as a lyophilisate, lyophilisation can be carried out using standard equipment for lyophilisation or vacuum drying. The lyophilisation cycle can be modified by those skilled in the art to suit the equipment and facilities used.

[0166] Preferably, said pre-lyophilized aqueous solution is an aqueous solution of indocyanine green as described above. The pre-lyophilized aqueous solution according to the invention may be sterilized before lyophilization. Typically, this sterilization is carried out by filtering the solution through a suitable membrane.

[0167] Preferably, the aqueous solution is lyophilized shortly after preparation to prevent degradation of the composition. The lyophilized product can be used even after storage for several months. Preferably, the lyophilized product is stored under conditions protected from air and / or moisture and / or light.

[0168] In addition, when the indocyanine green composition according to the present invention is provided as an aqueous composition, it may be reconstituted using the lyophilized composition and the aqueous diluent of the above-mentioned kit. When reconstituting, the lyophilized composition may be introduced into a vial, and then the aqueous diluent may be added thereto.

[0169] Advantageously, the composition prepared in lyophilized or aqueous form is immediately after preparation placed in a light-protected, airtight, sealed container, which may be, for example, a bottle, an ampoule, a vial, a syringe, a tube, etc. The container may be, for example, made of glass, polymer, metal, etc. The container may be a single-dose or multi-dose container.

[0170] In a preferred embodiment of the present invention, the container is preferably a colored glass vial or a colored glass ampoule. Advantageously, the container is selected from a colored glass vial with a vacuum headspace or a colored glass ampoule. Preferably, the composition according to the present invention is stored in a colored glass ampoule.

[0171] Advantageously, the composition according to the invention is stored at room temperature.

[0172] (Use of the composition according to the present invention) The composition of the invention may be used in a method of diagnosis and / or (as a therapeutic agent) treatment of a patient, comprising administering an effective amount of an aqueous solution according to the invention. The administration may be by enteral route, parenteral route, in particular by intravenous injection or topical application. Preferably, the administration is by intravenous injection.

[0173] The term "effective amount" refers to an amount of indocyanine green that elicits the biological or medical response in a tissue, system, animal or human that is desired by, for example, a researcher, clinician, or the like.

[0174] Advantageously, the composition of the present invention may be used in a method of diagnosis and / or treatment as described above, wherein an aqueous ICG composition is administered to a patient at least one month, preferably at least six months, more preferably at least twelve months, and most preferably at least two years after preparation or reconstitution from any of the above-mentioned kits.

[0175] In particular, the composition according to the present invention can be used for hyperthermia, ICG-enhanced selective photocoagulation, photodynamic therapy (PDT), and photodynamic thermotherapy (PHT). More specifically, the composition according to the present invention can be used for PDT, in the treatment of infectious diseases, acne, macular surgery, and cancer treatment. The composition according to the present invention may be used in conjunction with other therapeutic techniques, such as immunotherapy, radiation therapy, ultrasound therapy, and chemotherapy.

[0176] The compositions of the present invention may also be used to obtain angiographic images of patient tissue, measure cardiac output, measure liver function and hepatic blood flow, and diagnose and treat age-related macular degeneration and associated choroidal neovascularization and tumors.

[0177] The compositions of the present invention are based on components that have undergone clinical trials and are approved for administration to humans and / or animals.

[0178] The amount of the ICG composition administered to a patient for diagnostic use should be sufficient to cause the dye to fluoresce when irradiated with a suitable wavelength, taking into account that the absorption and emission peaks of ICG are within the range of 800-850 nm. In the same manner, in the case of a treatment method using an ICG solution, a sufficient amount of dye should be administered so that the treatment can be performed efficiently. The dosage of the ICG composition can be easily determined by a person skilled in the art, and is preferably equal to or higher than the currently permitted concentration for use in ophthalmic angiography, for example, 2 ml of a 20 mg / mL ICG solution for diagnostic use. A person skilled in the art will understand that in any of the above diagnostic and treatment methods, a higher concentration of dye may be advantageous.

[0179] The increased stability of ICG in the solutions of the present invention allows for improved diagnosis / treatment of patients with the same injected dose of compound as in prior art ICG compositions, because the increased stability of ICG results in a stronger response to fluorescent illumination. EXAMPLES

[0180] I. Materials and Methods (1. Reagents and Chemicals) Indocyanine green was provided by Biophore India Pharmaceuticals and used without further purification. Ascorbic acid was provided by Fisher Scientific. Sodium ascorbate was provided by Aldrich. Other reagents were purchased from Aldrich and used without further purification.

[0181] (2.Equipment) The stability of the formulations of the examples was measured by high pressure liquid chromatography (HPLC).The equipment used for the analysis was an Agilent Infinity1260 HPLC / MS system equipped with a diode array detector (UV-visible light), a Waters XbridgeShield RP18 (4.6x100; 5μm) column, and an ESI type mass spectrometer (solution input volume: 5μL, flow rate: 1ml / min, column temperature: 30℃, detection: 240nm).

[0182] The elution is performed by gradient elution. The mobile phase consists of a mixture of ammonium acetate / acetic acid buffer (10 mM, pH: 5.5) (eluent A) and acetonitrile (eluent B). The composition of the mobile phase was changed continuously during the elution process according to Table I below:

[0183] [Table 1]

[0184] (3. Sample Preparation) To prepare the ICG formulation described below, weigh out the amount of stabilizing compound (b) and, optionally, one or more compounds (c) into a Type I clear glass vial. Then, taking into account the tare weight of the vial, weigh out 25 mg of indocyanine green into the vial. Close the vial with a bromobutyl rubber stopper (VWS) and purge with a stream of nitrogen.

[0185] Then, add 10 ml of pre-degassed WFI via syringe. The vial is gently stirred by hand until the solubility of the solute is reached. The vial is then stored at room temperature away from light.

[0186] II- Compositions Comprising Indocyanine Green (a) and a Stabilizing Compound (b) According to the Invention

[0187] [Table 2]

[0188] Table III below shows the stability over several months of an ICG composition containing 25 mg ICG and 10 mg sodium ascorbate (ICG concentration = 2.5 mg / ml, sodium ascorbate / ICG weight ratio = 0.4).

[0189] [Table 3]

[0190] The results shown in Tables II and III indicate that in the presence of ascorbic acid or sodium ascorbate, ICG does not decompose in aqueous solution and is stable over time.

[0191] (III-Compositions Comprising Indocyanine Green (a), a Stabilizing Compound (b) and an Additional Compound (c) According to the Invention)

[0192] [Table 4]

[0193] To prepare a solution consisting of sodium chloride, the amounts of ICG, compound (b) and methionine or histidine (compositions C8 and C10) are weighed into a vial, the vial is closed with a bromobutyl rubber stopper and then purged with a nitrogen stream. Then, 5 ml of degassed WFI and 5 ml of 0.9% NaCl degassed solution are added. Composition C9 is prepared according to the same protocol without methionine or histidine.

[0194] The results reported in Table IV show that aqueous indocyanine green compositions containing sodium ascorbate and one or more additional compounds (c) selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, or DTT are also stable over time. Compound (c) in Table IV does not significantly affect the stability of ICG compositions containing stabilizing compound (b) according to the present invention.

[0195] (IV-Effect of Varying Compound (b) / Indocyanine Green (a) Ratio)

[0196] [Table 5]

[0197] (V- Comparative composition) Composition C18 was prepared according to the protocol in § I-3. Composition C19 was prepared according to the following protocol: 1.0 mg of ICG was weighed and added to 250 ml of phosphate buffer (pH 7.41). Then, 3 ml of 0.0002 M sodium ascorbate solution was added. The solutions used to prepare this composition were degassed before use. The resulting ICG solution was stored at room temperature away from light. This composition is according to Example 7 of the prior art WO9423646.

[0198] Composition C19 was prepared according to the following protocol: 2.5 mg of ICG was weighed and added to 250 ml of phosphate buffer (pH 7.41). 1.5 ml of this ICG / phosphate buffer was combined with 1.5 ml of 0.10 M ascorbate solution. The solutions used to prepare this composition were degassed before use. The resulting ICG solution was stored at room temperature away from light. This composition is according to Example 8 of the prior art WO9423646.

[0199] [Table 6]

[0200] The results in Table VI show that when the concentration of ICG is very low, even when the weight ratio of sodium ascorbate / ICG is within the range of 0.15-30, the ICG solution is not storage stable.

[0201] [Table 7]

[0202] Composition C25 was prepared by weighing the amount of ICG in a vial, closing the vial with a bromobutyl rubber stopper and purging with a stream of nitrogen, followed by the addition of 5 ml of degassed WFI and 5 ml of 0.9% NaCl degassed solution.

[0203] Compositions C26 and C27 were prepared by weighing the amount of ICG and additional compound into a vial, closing the vial with a bromobutyl rubber stopper and purging with a stream of nitrogen. 10 ml of phosphate buffered saline was added.

[0204] The results in Table VII show that the content of ICG in aqueous solution changes considerably over a short period of time in the absence of stabilizer compound (b), and the solution is not stable in the presence of compound (c) alone.

Claims

1. The aqueous composition comprises at least indocyanine green (a) and a compound (b) selected from ascorbic acid, a salt thereof and a mixture thereof, the weight ratio of compound (b) / indocyanine green (a) being 0.15 to 30, and the concentration of indocyanine green in the aqueous composition being 0.1 to 30 mg / ml.

2. 2. The indocyanine green composition according to claim 1, wherein the weight ratio of the compound (b) to the indocyanine green (a) is in the range of 0.15 to 15.

3. 3. The indocyanine green composition of claim 1 or claim 2, wherein the compound (b) is ascorbic acid.

4. 3. The indocyanine green composition of claim 1 or claim 2, wherein compound (b) is a salt of ascorbic acid.

5. 5. The indocyanine green composition of claim 4, wherein compound (b) is selected from sodium ascorbate, potassium ascorbate, calcium ascorbate, magnesium ascorbate, and mixtures thereof, preferably sodium ascorbate.

6. 6. The indocyanine green composition of claim 1, further comprising at least one compound (c) selected from reduced glutathione, thioglycerol, sorbitol, methionine, sodium chloride, histidine, dithiothreitol (DTT) and mixtures thereof.

7. 7. The indocyanine green composition according to claim 6, wherein the weight ratio of compound (c) to indocyanine green (a) is in the range of 1:500 to 100:1, preferably 1:100 to 10:1, more preferably 1:100 to 4:

1.

8. 8. The indocyanine green composition of claim 1, comprising up to 5% by weight of sodium iodide, based on the weight of indocyanine green.

9. The indocyanine green composition according to any one of claims 1 to 8, wherein the concentration of indocyanine green in the aqueous composition is 1 to 25 mg / ml.

10. 10. The aqueous composition according to claim 1, wherein the content of ICG in the composition before storage is 90% or more, and the content of indocyanine green is measured as an area percentage (%) at 240 nm by HPLC.

11. The indocyanine green composition according to any one of claims 1 to 10 for use as a pharmaceutical or diagnostic agent.

12. A kit comprising at least 1) a lyophilized powder of the composition of any one of claims 1 to 10, and 2) an aqueous diluent in separate compartments.

13. A kit that can be used for the preparation of the composition according to any one of claims 1 to 10, comprising in separate compartments an aqueous solution comprising at least 1) indocyanine green (a), and at least 2) a compound (b) selected from ascorbic acid, its salts and mixtures thereof.