Treatment of viral infections

Abstract

The present invention relates to dextran sulfate or a pharma- ceutical acceptable salt thereof for use in the prophylaxis, inhibition and / or treatment of flavivirus infection or infectious disease. The dextran sulfate or a pharma-ceutical acceptable salt thereof has an average molecular weight of 10,000 Da or less and an average sulfur content of 15% or more.

Description

[Technical field]

[0001] The present invention relates generally to the treatment of viral and infectious diseases, and in particular to the use of dextran sulfate in the treatment of flavivirus infections and infectious diseases caused by flavivirus infections. [Background technology]

[0002] Flavivirus is a genus of positive-stranded ribonucleic acid (RNA) viruses in the family Flaviviridae, which includes West Nile virus, Dengue virus, tick-borne encephalitis virus, yellow fever virus, Zika virus, and several other viruses that can cause encephalitis.

[0003] Flaviviruses generally share several common characteristics, including size (40-65 nm), symmetry (an enveloped icosahedral nucleocapsid), nucleic acid (single-stranded positive-stranded RNA viruses of approximately 10,000-11,000 bases), and appearance under an electron microscope.

[0004] Most of these viruses are classified as arboviruses because they are primarily transmitted by the bite of infected arthropods (mosquitoes or ticks). For most of these arboviruses, human infection is accidental, and humans are dead-end hosts because they cannot replicate the virus to high enough titers to reinfect the arthropods needed to continue the viral life cycle. Exceptions to this are yellow fever virus, dengue virus, and Zika virus. Although these three viruses still require a mosquito vector, they are well adapted to humans and do not necessarily depend on an animal host.

[0005] Flaviviruses can cause a variety of infections depending on the specific virus species that infects the human host. The dengue virus causes dengue fever, which in severe cases can develop into dengue hemorrhagic fever or dengue shock syndrome (commonly referred to as severe dengue, which is potentially fatal). The Zika virus causes Zika fever. Zika fever, also known as Zika virus disease, has symptoms similar to dengue fever. However, maternal-fetal transmission during pregnancy can cause microcephaly and other brain abnormalities in the child. Additionally, Zika virus infection has been associated with Guillain-Barré syndrome (GBS) in adults. Yellow fever is caused by the yellow fever virus and symptoms include fever, chills, loss of appetite, nausea, and muscle pain, but often last for less than five days. However, about 15% of infected individuals develop liver damage, yellowing of the skin, and kidney problems.

[0006] US2004 / 0009953 and WO2005 / 004882 reviewed the antiviral effects of high molecular weight (40kDa, 500kDa) and low sulfated (6-13%) dextran sulfates against the HIV-1 virus. Antiviral Research 2017,143:186-194 reviewed the effects of high sulfated heparin, high molecular weight and low sulfated dextran sulfate (2.3 sulfurs per disaccharide, 40kDa) and suramin against Zika virus infection.

[0007] Once a subject has been infected with a flavivirus, treatment to reduce the risk of infection and spread of the virus is generally required. Summary of the Invention

[0008] The general object is to provide a treatment for a flavivirus infection or infection.

[0009] This and other objectives are met by the embodiments disclosed herein.

[0010] An embodiment of the present invention relates to dextran sulfate or a pharma- ceutically acceptable salt thereof for use in the prophylaxis, inhibition and / or treatment of a flavivirus infection or infectious disease. The dextran sulfate or a pharma- ceutically acceptable salt thereof has an average molecular weight of 10,000 Da or less and an average sulfur content of 15% or more.

[0011] The experimental data presented herein demonstrate that dextran sulfate, or a pharma- ceutically acceptable salt thereof, can be used to treat flavivirus infections and flavivirus infections. More specifically, dextran sulfate produced significant dose-dependent inhibition of Zika, Dengue, and Yellow Fever virus infection of human cells at clinically relevant concentrations.

[0012] The embodiments, together with further objects and advantages thereof, may best be understood by reference to the following description taken together with the accompanying drawings, in which: [Brief description of the drawings]

[0013] [Figure 1A] FIG. 1 illustrates that dextran sulfate inhibits Zika virus infection in human HeLa cells in a concentration-dependent manner at an MOI of 0.5. Data represent the mean + SEM. [Figure 1B] FIG. 1 illustrates that dextran sulfate inhibits Zika virus infection in human HeLa cells in a concentration-dependent manner at an MOI of 3. Data represent the mean + SEM. [Figure 1C] FIG. 1 illustrates that dextran sulfate inhibits Zika virus infection in human HeLa cells in a concentration-dependent manner at an MOI of 5. Data represent the mean + SEM. [Diagram 2] 2A-2B illustrate that dextran sulfate inhibits Zika virus infection in Vero cells in a concentration-dependent manner at an MOI of 0.05 (2A) and 0.01 (2B). Data represent the mean + SEM. [Diagram 3] FIG. 1 illustrates control monensin inhibition of Zika virus infection in human HeLa cells. [Figure 4A]FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4B] FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4C] FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4D] FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4E]FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4F] FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4G] FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Figure 4H] FIG. 1 illustrates the ability of dextran sulfate and the positive control viral infectivity inhibitor monensin to prevent infection of mammalian cells by the indicated flaviviruses in vitro. Concentration-effect data were analyzed by iterative curve fitting with a four-parameter logistic equation. The x-axis is in μM for dextran sulfate and monensin, respectively. Data represent individual data points from a single experiment. [Diagram 5] FIG. 1 illustrates that dextran sulfate inhibits Dengue, Zika, and Yellow Fever virus infection in human cells by >90% in a concentration-dependent manner. Data represent the mean + SD. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] The present invention relates generally to the treatment of viral and infectious diseases, and in particular to the use of dextran sulfate in the treatment of flavivirus infections and infectious diseases caused by flavivirus infections.

[0015] Dextran sulfate was able to prevent flavivirus infections, represented by Zika virus (ZIKV), Dengue virus (DENV), and Yellow fever virus (YFV) infections in human cells at clinically relevant concentrations. More specifically, dextran sulfate treatment significantly reduced the number of infected cells compared to controls, providing evidence that dextran sulfate can significantly prevent or at least inhibit flavivirus infections in human cells.

[0016] Flavivirus particles, or virions, are enveloped and spherical with a diameter of about 50 nm. The surface proteins are arranged in an icosahedral-like symmetry. Mature virions contain primarily two virally encoded membrane proteins designated protein M and protein E, whereas immature virions contain membrane protein precursors.

[0017] Flavivirus infections in general, and Zika, Dengue and Yellow Fever virus infections in particular, can be considered as a two-step process: initial virus-cell attachment followed by internalization into the host cell. Initial virus-cell attachment is mediated through interactions between the viral envelope protein E and negatively charged glycosaminoglycans (GAGs) on the cell surface (Infection, Genetics and Evolution 2019 69:22-29), enriching the virus particles on the cell surface. The actual internalization into the host cell is mediated by cell surface receptors on the host cell. Neural cell adhesion molecule (NCAM1), as well as heparan sulfate (Virology 2002,292:162-168), have recently been identified as potential flavivirus receptors for cell internalization, and Zika virus receptors in particular (Nature Communications 2020 11:3896). However, no direct Zika virus-heparin interaction was observed in heparin-binding assays, and downregulation or removal of cellular heparan sulfate did not inhibit Zika virus infection (Antiviral Research 2017,143:186-194), indicating that cell surface heparan sulfate is not utilized by Zika virus as an attachment receptor.

[0018] A possible mechanism for the inhibitory effect of dextran sulfate on flavivirus infection may be to interfere with initial virus cell attachment and / or binding to cell surface receptors. The dextran sulfate of the present embodiment exhibits a negative charge and may mimic heparan sulfate and other negatively charged GAGs on the cell surface. Thus, the dextran sulfate of the present embodiment may block or at least interfere with the binding between the viral envelope protein E and heparan sulfate and other negatively charged GAGs on the host cell surface. In addition, NCAM1 and other cell surface receptors contain heparin binding domains (HBDs) (Journal of Neuroscience Research 1992 33(4):538-548). The dextran sulfate of the present embodiment may bind to such HBDs or act as a competitive antagonist, and thus may interfere with the binding of flavivirus particles to NCAM1 and other cell surface receptors containing such HBDs or heparan sulfates.

[0019] Thus, the experimental results seen with dextran sulfate in this embodiment may be obtained by disrupting the initial virus-cell attachment and disrupting the interaction between viral proteins and cell surface receptors (such as NCAM1 and heparan sulfate).

[0020] An embodiment of the invention relates to dextran sulfate, or a pharma- ceutically acceptable salt thereof, for use in preventing, inhibiting, or treating a flavivirus infection or infectious disease.

[0021] In one embodiment, the flavivirus infection is selected from the group consisting of flavivirus infections caused by a flavivirus selected from the group consisting of Dengue virus (DENV), Japanese encephalitis virus (JEV), Kunjin virus (KUNV), Langat virus (LGTV), Louping ill virus (LIV), Murray Valley encephalitis virus (MVEV), St. Louis encephalitis virus (SLEV), Powassan virus (POWV), West Nile virus (WNV), Yellow fever virus (YFV), and Zika virus (ZIKAV). In another embodiment, the flavivirus infection is selected from the group consisting of flavivirus infections caused by a flavivirus selected from the group consisting of DENV, JEV, KUNV, LIV, MVEV, SLEV, POWV, WNV, YFV, and ZIKV.

[0022] In certain embodiments, the flavivirus infection is selected from the group consisting of flavivirus infections caused by a flavivirus selected from the group consisting of DENV, YFV, and ZIKV.

[0023] In a preferred embodiment, the flavivirus infection is a flavivirus infection caused by ZIKV.

[0024] In another preferred embodiment, the flavivirus infection is a flavivirus infection caused by DENV.

[0025] In a further embodiment, the flavivirus infection is a flavivirus infection caused by YFV.

[0026] In one embodiment, the flavivirus infection is caused by a flavivirus other than Japanese encephalitis virus, West Nile virus, dengue virus, and yellow fever virus.

[0027] Flavivirus infections, as used herein, include dengue fever, dengue hemorrhagic fever, and dengue shock syndrome caused by DENV, Japanese encephalitis caused by JEV, non-encephalitic Kunjin virus disease and encephalitic Kunjin virus disease caused by KUNV, louping disease caused by LIV, Murray Valley encephalitis caused by MVEV, St. Louis encephalitis caused by SLEV, encephalitis caused by POWV, West Nile fever caused by WNV, yellow fever caused by YFV, and Zika fever or Zika virus disease caused by ZIKAV.

[0028] In a preferred embodiment, the flavivirus infection is selected from the group consisting of dengue fever, dengue hemorrhagic fever, and dengue shock syndrome caused by DENV, yellow fever caused by YFV, and Zika fever or Zika virus disease caused by ZIKAV. In a specific embodiment, the flavivirus infection is Zika fever or Zika virus disease caused by ZIKAV. In another specific embodiment, the flavivirus infection is dengue fever, dengue hemorrhagic fever, or dengue shock syndrome caused by DENV. In a further specific embodiment, the flavivirus infection is yellow fever caused by YFV.

[0029] In one embodiment, the flavivirus infection is a flavivirus infection other than Japanese encephalitis caused by JEV, West Nile fever caused by WNV, dengue fever, dengue hemorrhagic fever, and dengue shock syndrome caused by DENV, and yellow fever caused by YFV.

[0030] The present invention also relates to the use of dextran sulfate, or a pharma- ceutically acceptable salt thereof, for the manufacture of a medicament for the prevention, inhibition, and / or treatment of a flavivirus infection or infectious disease.

[0031] The present invention further relates to a method for preventing, inhibiting and / or treating a flavivirus infection or infection, the method comprising administering to a subject suffering from or at risk of suffering from a flavivirus infection or infection an effective amount of dextran sulfate, or a pharma- ceutically acceptable salt thereof.

[0032] Treatment of a flavivirus infection or infection, as used herein, does not necessarily mean therapeutic treatment of a flavivirus infection or infection, but also includes inhibition or alleviation of short-term and long-term symptoms of a flavivirus infection or infection. Thus, treatment also includes delaying the onset of a flavivirus infection or infection, including delaying the onset of symptoms associated with a flavivirus infection or infection, preventing the onset, and resolving established pathology.

[0033] Other viruses also use similar mechanisms to flaviviruses to attach to and internalize cells. For example, rabies virus also uses cell surface receptor NCAM1 for internalization (Philosophical Transactions Royal Society B 2015,370:2014035). More specifically, rabies virus and other lyssaviruses contain glycoprotein G, which is involved in host cell interaction and internalization. Thus, the dextran sulfate of the present embodiment can also be used to prevent, inhibit, or treat lyssavirus infection or infection.

[0034] In one embodiment, the lyssavirus infection is selected from the group consisting of lyssavirus infections caused by a lyssavirus selected from the group consisting of Australian bat lyssavirus (ABLV), Duvenhagerissavirus (DUVV), European bat lyssavirus (EBLV), Lagos bat virus (LBV), Mokola virus (MOKV), and rabies virus. In a particular embodiment, the lyssavirus infection is caused by the rabies virus.

[0035] In one embodiment, the lyssavirus infection is rabies, encephalitis, or rabies-like encephalitis disease. In a particular embodiment, the lyssavirus infection is rabies.

[0036] In one embodiment, the lyssavirus infection is caused by a lyssavirus other than the rabies virus.

[0037] In the following, references to the (average) molecular weight and sulfur content of dextran sulfate also apply to any pharma- ceutically acceptable salts of dextran sulfate. Thus, pharma- ceutically acceptable salts of dextran sulfate preferably have the average molecular weight and sulfur content as described in the following embodiments.

[0038] Dextran sulfate outside the preferred range of this embodiment may be less effective and / or may cause negative side effects to the cell or subject.

[0039] For example, dextran sulfate with a molecular weight of more than 10,000 Da (10 kDa) generally has a lower effect-to-side effect profile than dextran sulfate with a lower average molecular weight. This means that the maximum dose of dextran sulfate that can be safely administered to a subject is lower for larger dextran sulfate molecules (>10,000 Da) than for dextran sulfate molecules with average molecular weights within the preferred range. As a result, such larger dextran sulfate molecules are less suitable for clinical use when dextran sulfate is administered to a subject in vivo. For example, US2004 / 0009953 discloses that dextran sulfate with an average molecular weight of 40 kDa or 500 kDa and an average sulfur content of 17% was highly toxic to rats.

[0040] The dextran sulfate or its pharma- ceutically acceptable salt according to the present invention is non-toxic as shown in the examples presented. Therefore, the dextran sulfate or its pharma- ceutically acceptable salt according to the present invention does not have toxicity problems compared with the high molecular weight dextran sulfate disclosed in US2004 / 0009953.

[0041] Dextran sulfate is a sulfated polysaccharide, specifically a sulfated glucan, i.e. a polysaccharide made up of many glucose molecules. The average molecular weight defined herein indicates that the molecular weight of an individual sulfated polysaccharide may differ from this average molecular weight, but the average molecular weight represents the average molecular weight of the sulfated polysaccharide. This further means that for dextran sulfate samples, there is a natural distribution of molecular weights around this average molecular weight.

[0042] The average molecular weight of dextran sulfate (more precisely, the weight-average molecular weight (M w ) is usually determined using indirect methods such as gel exclusion / permeation chromatography, light scattering, or viscosity. The determination of average molecular weight using such indirect methods depends on many factors, including the choice of column and eluent, flow rate, calibration procedure, etc.

[0043]

number

[0044] In one embodiment, the dextran sulfate or a pharma- ceutically acceptable salt thereof is w In certain embodiments, the dextran sulfate or a pharma- ceutically acceptable salt thereof has an M w is in the range of 2,000Da to 10,000Da.

[0045] In another embodiment, the dextran sulfate or a pharma- ceutically acceptable salt thereof is w In a particular embodiment, the dextran sulfate or a pharma- ceutically acceptable salt thereof has an M w is within the interval of 3,500 Da to 9,500 Da, for example within the interval of 3,500 Da to 8,000 Da.

[0046] In other particular embodiments, the dextran sulfate or a pharma- ceutically acceptable salt thereof is M w is within the interval of 4,500 Da to 7,500 Da, for example within the interval of 4,500 Da to 6,500 Da, or within the interval of 4,500 Da to 5,500 Da.

[0047] Thus, in some embodiments, dextran sulfate or a pharma- ceutically acceptable salt thereof is M w is 10,000 Da or less, 9,500 Da or less, 9,000 Da or less, 8,500 Da or less, 8,000 Da or less, 7,500 Da or less, 7,000 Da or less, 6,500 Da or less, 6,000 Da or less, or 5,500 Da or less.

[0048] In some embodiments, the dextran sulfate or a pharma- ceutically acceptable salt thereof is w is 1,000 Da or more, 1,500 Da or more, 2,000 Da or more, 2,500 Da or more, 3,000 Da or more, 3,500 Da or more, 4,000 Da or more, or 4,500 Da or more. Any of these embodiments may be referred to as M w This may be combined with any of the embodiments presented above that specify an upper limit of, for example, 10,000 Da or less.

[0049] In certain embodiments, the M of dextran sulfate or a pharma- ceutically acceptable salt thereof as set forth above is w is the average M wand is preferably determined by gel exclusion / permeation chromatography, size exclusion chromatography, light scattering or viscosity-based methods.

[0050]

number

[0051] In one embodiment, dextran sulfate or a pharma- ceutically acceptable salt thereof has an M n is in the range of 1,850 to 3,500 Da.

[0052] In certain embodiments, dextran sulfate or a pharma- ceutically acceptable salt thereof has a M n is within the interval of 1,850 Da to 2,500 Da, preferably within the interval of 1,850 Da to 2,300 Da, for example within the interval of 1,850 Da to 2,000 Da.

[0053] Thus, in some embodiments, dextran sulfate or a pharma- ceutically acceptable salt thereof is M n In addition, the dextran sulfate or a pharma- ceutically acceptable salt thereof has an M of 3,500 Da or less, 3,250 Da or less, 3,000 Da or less, 2,750 Da or less, 2,500 Da or less, 2,250 Da or less, or 2,000 Da or less. n is 1,850 Da or more.

[0054] In one embodiment, dextran sulfate or a pharma- ceutically acceptable salt thereof has an average sulfate number per glucose unit in the interval of 2.5 to 3.0.

[0055] In a particular embodiment, dextran sulfate or a pharma- ceutically acceptable salt thereof has an average number of sulfates per glucose unit in the interval 2.5 to 2.8, preferably in the interval 2.6 to 2.7.

[0056] In one embodiment, the dextran sulfate or a pharma- ceutically acceptable salt thereof has an average number of glucose units in the interval of 4.0 to 6.0.

[0057] In a particular embodiment, the dextran sulfate or a pharma- ceutically acceptable salt thereof has an average number of glucose units in the interval 4.5 to 5.5, preferably in the interval 5.0 to 5.2.

[0058] In one embodiment, dextran sulfate or a pharma- ceutically acceptable salt thereof has an M n is within the range of 1,850 to 3,500 Da, the average number of sulfates per glucose unit is within the range of 2.5 to 3.0, and the average sulfation at the C2 position of the glucose unit of dextran sulfate is at least 90%.

[0059] In one embodiment, the dextran sulfate has an average number of glucose units of about 5.1, an average number of sulfates per glucose unit in the range of 2.6 to 2.7, and an M n is in the range of 1,850Da to 2,000Da.

[0060] In one embodiment, the pharma- ceutically acceptable salt of dextran sulfate is the sodium salt of dextran sulfate. In a particular embodiment, the sodium salt of dextran sulfate has an average number of glucose units of about 5.1, an average number of sulfates per glucose unit in the range of 2.6 to 2.7, and an M with a Na counterion. n is in the range of 2,100Da to 2,300Da.

[0061] In one embodiment, dextran sulfate has an average number of glucose units of 5.1, an average number of sulfates per glucose unit of 2.7, and an average Na+-free Mn Approximately 1,900-1,950 Da, with an average M of 0.01, including Na+, as measured by NMR spectroscopy n is approximately 2,200 to 2,250 Da.

[0062] Dextran sulfate is a polyanionic derivative of dextran and contains sulfur. The average sulfur content of the dextran sulfate of this embodiment is 15% or more. In one embodiment, the average sulfur content of the dextran sulfate of this embodiment is preferably 15-20%, more preferably about 17%, usually corresponding to about or at least two sulfate groups per glucosyl residue. In a particular embodiment, the sulfur content of the dextran sulfate is preferably equal to or at least close to the maximum possible sulfur content of the corresponding dextran molecule.

[0063] Dextran sulfate according to this embodiment may be provided as a pharma- ceutically acceptable salt of dextran sulfate, such as the sodium or potassium salt.

[0064] A currently preferred dextran sulfate according to this embodiment is disclosed in WO2016 / 076780.

[0065] The subject is preferably a mammalian subject, more preferably a primate, particularly a human subject. However, dextran sulfate or its pharma- ceutically acceptable salts can also be used in veterinary applications. Non-limiting examples of animal subjects include primates, cats, dogs, pigs, horses, mice, and rats.

[0066] Dextran sulfate or a pharma- ceutically acceptable salt thereof is preferably administered to a subject by injection, in particular intravenous (iv), subcutaneous (sc) or intraperitoneal (ip) injection, preferably iv or sc injection. Other parenteral administration routes that can be used include intramuscular and intraarticular injection. Alternatively or additionally, injection of dextran sulfate or a pharma- ceutically acceptable derivative thereof can be performed, for example, directly into a tissue or organ or other site in the subject's body where the target effect is to occur.

[0067] The dextran sulfate or its pharma- ceutically acceptable salt of this embodiment is preferably formulated as an aqueous injection solution using a selected solvent or excipient. The solvent is advantageously an aqueous solvent, in particular a buffer solution. Non-limiting examples of such buffer solutions are citrate buffers, such as citrate monohydrate (CAM) buffer, or phosphate buffers. For example, the dextran sulfate of this embodiment can be dissolved in saline, such as 0.9% NaCl saline, and optionally buffered with 75 mM CAM, adjusting the pH to about 5.9 using sodium hydroxide. Non-buffered solutions are also possible, including aqueous injection solutions such as saline, i.e. NaCl (aqueous). Furthermore, if a buffer solution is required, other buffer systems other than CAM can also be used.

[0068] This embodiment is not limited to injection, and other routes of administration may alternatively be used, including oral, nasal, buccal, rectal, transdermal, tracheal, bronchial, or topical administration. The active compound, dextran sulfate, is then formulated with a suitable excipient or carrier selected based on the particular route of administration.

[0069] Suitable dosage ranges of dextran sulfate or a pharma- ceutically acceptable salt thereof may vary depending on the application, such as in vitro versus in vivo, the size and weight of the subject, the condition the subject is being treated, and other considerations. Specifically, for a human subject, possible dosage ranges may be 1 μg / kg to 100 mg / kg body weight, preferably 10 μg / kg to 50 mg / kg body weight.

[0070] In a preferred embodiment, dextran sulfate or a pharma- ceutically acceptable salt thereof is formulated to be administered at a dose in the range of 0.05-50 mg / kg of subject body weight, preferably 0.05 or 0.1-40 mg / kg of subject body weight, more preferably 0.05 or 0.1-30 mg / kg, or 0.1-25 mg / kg, or 0.1-15 mg / kg, or 0.1-10 mg / kg of subject body weight. A preferred dose is selected in the range of 0.25-5 mg / kg, preferably 0.5-2.5 mg / kg, more preferably 0.75-2 mg / kg of subject body weight.

[0071] Dextran sulfate or a pharma- ceutical acceptable derivative thereof can be administered in a single dose, such as in the form of a single bolus injection. This bolus dose can be injected very quickly into the subject, but it can also be advantageous to inject the dextran sulfate solution over a period of time, such that the dextran sulfate solution is infused into the patient over a period of several minutes, such as 5-10 minutes.

[0072] Alternatively, dextran sulfate or a pharma- ceutically acceptable salt thereof can be administered on multiple (ie, at least two) occasions during the treatment period.

[0073] Dextran sulfate or a pharma- ceutically acceptable salt thereof can be administered sequentially, simultaneously, or in the form of a composition comprising dextran sulfate or a pharma- ceutically acceptable salt thereof and at least one other active agent, The at least one active agent can be selected from among any agent useful in any of the aforementioned diseases, disorders, or conditions. EXAMPLES

[0074] Example 1 - Antiviral activity against ZIKV The objective of this study was to investigate the antiviral activity of dextran sulfate against Zika virus (ZIKV).

[0075] Materials and Methods In this example, the sodium salt of dextran sulfate was used (ILB®, Tikomed AB, Viken, Sweden, WO2016 / 076780). Dextran sulfate was provided in a stock solution of 100 mg / ml in 0.9% NaCl.

[0076] Viral infection was investigated in two cell lines using Zika virus PF13: human HeLa Kyoto cells and African green monkey kidney cells (Vero AD cells). The following conditions were tested: HeLa Kyoto with multiplicities of infection (MOI), i.e., the number of virions per cell at the start of infection, of 0.5, 3, and 5, and Vero AD with MOIs of 0.05 and 0.01. Cytotoxicity was determined using an MTT assay against uninfected cells treated with the same concentration of dextran sulfate.

[0077] reagent Complete medium: 5% M199 for Vero AD, 10% DMEM (Gibco) supplemented with 10% FBS (Gibco) and 1X p / s (Gibco) for HeLa Kyoto.

[0078] Supplementary medium: same as above for VeraAD, same as above for HeLa Kyoto but with 2% FBS.

[0079] Experimental procedure The antiviral activity of three dilutions of dextran sulfate was examined by pre-incubating the drug with the assay cells for 1 h prior to infection with Zika virus. Virus and formulations were left on the cells for the entire experiment, 48 h. Cytotoxicity of the same concentration range of dextran sulfate was determined by MTT assay in the same cell lines. For Zika virus, three MOIs were tested in parallel to ensure that the correct range of infectivity was reached.

[0080] cell seeding Cells were separated and counted. Two 96-well plates were seeded per cell type; one for antiviral assay and one for cytotoxicity assay. Cells were seeded at a density of 8,000 cells / well in 100 μl / well of complete medium. After seeding, plates were incubated at room temperature for 5 min to ensure even distribution, and then incubated at 37°C, 5% CO2 until the next day.

[0081] Plates contained 3 wells per dextran sulfate concentration (600μg / ml, 200μg / ml, 60μg / ml) and MOI, 9 infected control wells per MOI, 18 uninfected control wells per MOI and 8 monensin control wells. See table below. Bold represents 1st MOI, italics represents 2nd MOI and underline represents 3rd MOI. [Table 1] 600:600μg / ml dextran sulfate; 200:200μg / ml dextran sulfate; 60:60μg / ml dextran sulfate M: monensin (monensin sodium salt, Sigma); I: infected; U: uninfected

[0082] Compound pretreatment Dextran sulfate was prepared in three different concentrations from a stock solution (100 mg / ml).

[0083] 600 μg / ml: 6 μl of dextran sulfate stock in 994 μl of supplemented medium 200 μg / ml: 2 μl of dextran sulfate stock in 998 μl of supplemented medium 60 μg / ml: 100 μl of 600 μg / ml in 900 μl supplemented medium.

[0084] 2.4 μl of 10 mM monensin stock was added to 1200 μl of supplemented medium followed by vortex mixing. 240 μl of supplemented medium was added to three wells in each row of a round-bottom 96-well plate (designated M above). 360 μl of diluted monensin was added to the three top wells. 120 μl of the well volume in the three top wells was transferred to three wells in the next row, and so on, resulting in a 3-fold dilution of monensin per row.

[0085] Media was removed from the cells and replaced with 100 μl of prepared compound (dextran sulfate or monensin) or media (uninfected and untreated controls) for 1 hour.

[0086] Towards the end of the incubation, fresh dextran sulfate and monensin were prepared in 50 μl volumes at twice the final concentrations (as they were diluted to final concentrations with an equal volume of virus or medium).

[0087] 1,200 μg / ml: 7.2 μl of dextran sulfate stock in 592.8 μl of supplemented medium 400 μg / ml: 2.4 μl of dextran sulfate stock in 597.6 μl of supplemented medium 120μg / ml: 60μl of 1,200μg / ml dilution in 540μl supplemented medium

[0088] 2.4 μl of 10 mM monensin stock was added to 600 μl of supplemented medium followed by vortex mixing. 120 μl of supplemented medium was added to three wells in each row of a round-bottom 96-well plate (designated M above). 180 μl of diluted monensin was added to the three top wells. 60 μl of the well volume in the three top wells was transferred to three wells in the next row, and so on, resulting in a 3-fold dilution of monensin per row.

[0089] cytotoxicity Dextran sulfate and monensin were added simultaneously with the preincubation drugs and were left on for the duration of the experiment. Monensin was removed after 8 hours and replaced with supplemented medium to reduce cytotoxicity.

[0090] Virus preparation and transmission Virus was added according to the following MOI:

[0091] Zika MOI 0.5 (HeLa Kyoto): 27 μl of undiluted virus (1.5 × 10 7 PFU / ml) + 2,473 μl of supplemented medium Zika MOI3 (HeLa Kyoto): 160 μl undiluted virus + 2,340 μl supplemented medium Zika MOI5 (HeLa Kyoto): 270 μl undiluted virus + 2,230 μl supplemented medium

[0092] Zika MOI 0.5 (Vero AD): 27 μl of undiluted virus (1.5 × 10 7 PFU / ml) + 2,473 μl of supplemented medium Zika MOI 0.05 (Vero AD): 2.7 μl undiluted virus + 2,500 μl supplemented medium Zika MOI 0.01 (Vero AD): 0.6 μl undiluted virus + 2,500 μl supplemented medium

[0093] At the end of compound preincubation, media was removed from cells and replaced with 50 μl of diluted compound (dextran sulfate or monensin) or media (untreated and uninfected controls). 50 μl of diluted virus or media (uninfected controls) was added immediately. Plates were incubated at 37° C. and 5% CO2 for 48 hours, with formulations+virus remaining on the cells for the entire duration of the experiment.

[0094] Fixation and deployment After 48 hours, plates were washed with PBS, fixed with 4% formaldehyde for 30 minutes, washed again with PBS, and stored in PBS at 4° C. until staining.

[0095] Cytotoxicity plates were treated with MTT to determine cell viability.

[0096] Measures of infectivity Cells were immunostained by immunofluorescence staining. Briefly, any residual formaldehyde was quenched with 50 mM ammonium chloride, after which cells were permeabilized (0.1% Triton X100) and stained with an antibody recognizing flavivirus envelope protein (Millipore, MAB10216). Primary antibodies were detected with Alexa-488 conjugated secondary antibodies (Life Technologies, A11001), and cell nuclei were stained with Hoechst. Images were acquired using a 10X objective on a CellInsight CX5 high content platform (Thermo Scientific), and infection rates were calculated (infected cells / total cells x 100) using CellInsight CX5 software.

[0097] Cytotoxicity measurements Cytotoxicity was detected by MTT assay. Briefly, MTT reagent (Sigma) was added to cells for 2 h at 37°C and 5% CO2, after which the medium was removed and the precipitate was solubilized with a 1:1 mixture of isopropanol:DMSO for 20 min. The supernatant was transferred to a clean plate and the signal was read at 570 nm.

[0098] Determination of % Inhibition The normalized percentage of inhibition was calculated using the following formula:

number

[0099] For the control drug, EC 50 Values ​​were extrapolated from curves representing best fits (nonlinear regression analysis, variable slope) of the logarithm of compound concentration versus normalized percentage of inhibition using GraphPad Prism (version 9).

[0100] Determination of cytotoxicity The percentage of cytotoxicity was calculated using the following formula:

number

[0101] TC vs. control drug 50 Values ​​were extrapolated from curves representing best fits (nonlinear regression analysis, variable slope) of the logarithm of compound concentration versus percentage normalized cytotoxicity using GraphPad Prism (version 9).

[0102] result The bar graphs in Figures 1A-1C and 2A-2B show the % inhibition of infection (bars) and % cytotoxicity (dots) for dextran sulfate at the three concentrations tested for the indicated viruses. Dextran sulfate caused significant dose-dependent inhibition of Zika virus in both cell types.

[0103] No significant cytotoxicity was observed at any of the concentrations tested.

[0104] Given the unmet clinical need for effective antivirals against Zika and other flaviviruses, the results presented herein demonstrate that dextran sulfate of the present embodiments has therapeutic potential against Flaviviridae.

[0105] The efficacy of dextran sulfate against Zika virus was quite surprising considering that dextran sulfate was shown in Antiviral Research 2017, 143:186-194 to have a % infection inhibition of about 60% at 200 μg / ml in Vero cells, compared to a % infection inhibition of over 80% at 200 μg / ml with dextran sulfate according to the present invention (FIG. 2A). The dextran sulfate used in Antiviral Research 2017, 143:186-194 is low sulfate dextran sulfate (2.3 sulfates per disaccharide) from Sigma. In addition, dextran sulfate has a significantly higher average molecular weight (40 kDa). These differences in sulfation number and average molecular weight between the dextran sulfate of the present invention and the dextran sulfate in Antiviral Research 2017, 143:186-194 are believed to be the basis for the significant difference in antiviral activity against Zika virus.

[0106] Example 2 - Antiviral activity against DENV, ZIKV, and YFV The objective of this study was to investigate the antiviral activity of dextran sulfate against dengue virus (DENV), Zika virus (ZIKV), and yellow fever virus (YFV).

[0107] Materials and Methods In this example, the sodium salt of dextran sulfate was used (ILB®, Tikomed AB, Viken, Sweden, WO2016 / 076780). Dextran sulfate was provided in a stock solution of 100 mg / ml in 0.9% NaCl.

[0108] Viral infections were investigated in two cell lines: human Huh-7 cells (VRS stock P1) using a panel of four serotypes of Dengue virus (DENV) and the vaccine strain of Yellow Fever 17D (YF17D), and human HeLa Kyoto cells (VRS stock P1) using two strains of Zika virus (ZIKV).

[0109] Virus: DENV1 (Hawaii strain, GenBankTM code EU848545) DENV2 (New Guinea C strain, GenBankTM code AF038403) DENV3 (H87 strain, GenBank code M93130) DENV4 (strain H241; GenBank code AY947539) African ZIKV lineage (MP1751 strain) ZIKV Asian lineage (PF / 13) YF17D (VRS stock)

[0110] reagent Complete medium: DMEM (Gibco) supplemented with 10% FBS (Gibco) and 1X p / s (Gibco). Supplemented medium: DMEM (Gibco) supplemented with 2% FBS (Gibco) and 1X p / s (Gibco).

[0111] Experimental procedure The antiviral activity of eight dilutions of dextran sulfate was examined by pre-incubating the drug with the assay cells for 1 h before adding the virus. The virus and formulations were left on the cells for the entire experiment, 48 h. The cytotoxicity of the same concentration range of dextran sulfate was determined by MTT assay.

[0112] cell seeding Cells were dissociated and counted. Two 96-well plates were seeded per cell type; one for antiviral assay and one for cytotoxicity assay. Huh-7 and HeLa cells were seeded at a density of 8,000 cells / well in 100 μl / well of complete medium. After seeding, plates were incubated at room temperature for 5 min to ensure even distribution, and then incubated at 37°C, 5% CO2 until the next day.

[0113] Virus preparation and transmission Virus stocks were diluted in supplemented medium to reach the following MOIs: DENV1-3: 0.3 DENV4:1 ZENV:5 YF17D:0.1

[0114] Formulation Dilution Dextran sulfate formulations were prepared at twice the final concentration (because they were diluted to the final concentration with an equal volume of virus or medium).

[0115] For each virus, a dextran sulfate test solution was prepared by adding 150 μl of a 100 mg / ml stock to 600 μl of supplemented medium (20 mg / ml).

[0116] Monensin, as a control inhibitor, was prepared for each relevant virus by adding 3 μl of a 10 mM stock to 747 μl of supplemented medium (40 μM).

[0117] In row A of a round-bottom 96-well plate, the dilutions prepared above were added in triplicate (180 μl / well). In rows B-H, 120 μl of supplemented medium was added. For 3-fold serial dilutions, 60 μl was transferred from the previous row to the next row with 15 mixings in between.

[0118] Cell treatment The medium was removed from the cells and 50 μl of supplemented medium was added immediately followed by 50 μl of the diluted formulation. Medium alone was added to columns 11 and 12 (untreated control). After mixing, the plate was incubated at 37° C., 5% CO2 for 1 hour.

[0119] Cytotoxicity Testing The medium was removed from the cells and replaced with 50 μl of supplemented medium, followed by 50 μl of diluted formulation or medium. After mixing, the plates were incubated at 37° C. and 5% CO2 for 48 hours.

[0120] cell infection After 1 hour, the medium was removed from the cells and replaced with 50 μl of virus or medium (uninfected, untreated controls), immediately followed by 50 μl of previously prepared formulation dilutions. Plates were incubated at 37° C. and 5% CO2 for 48 hours, with formulation+virus remaining on the cells for the entire duration of the experiment.

[0121] Fixation and deployment After 48 hours, plates were washed with PBS, fixed with 4% formaldehyde for 30 minutes, washed again with PBS, and stored in PBS at 4° C. until staining.

[0122] Cytotoxicity plates were treated with MTT to determine cell viability.

[0123] Measures of infectivity Cells were immunostained by immunofluorescence staining. Briefly, any residual formaldehyde was quenched with 50 mM ammonium chloride, after which the cells were permeabilized (0.1% Triton X100) and stained with antibodies recognizing dengue virus envelope proteins (Invitrogen MA1-27093, for DENV1-3), pan-flavivirus envelope (Millipore MAB10216, for DENV4 and ZIKV), and yellow fever E antibody 3576 (Santa Cruz, sc-58083, for YF17D). Primary antibodies were detected with Alexa-488 conjugated secondary antibodies (Life Technologies, A11001), and cell nuclei were stained with Hoechst. Images were acquired using a 4X objective on a CellInsight CX5 high content platform (Thermo Scientific), and infection rates were calculated using CellInsight CX5 software (infected cells / total cells × 100).

[0124] Cytotoxicity measurements Cytotoxicity was detected by MTT assay. Briefly, MTT reagent (Sigma) was added to cells for 2 h at 37°C and 5% CO2, after which the medium was removed and the precipitate was solubilized with a 1:1 mixture of isopropanol:DMSO for 20 min. The supernatant was transferred to a clean plate and the signal was read at 570 nm.

[0125] EC 50 Concentration determination Concentration-effect data were analyzed using KaleidaGraph (v5; Synergy Software) to fit a four-parameter logistic equation (m1 = maximal inhibition of infectivity, m2 = EC 50 concentration, m3 = Hill coefficient, m4 = maximum infectivity; if m1>m4 after analysis, the analysis was repeated such that m1 was forced to be equal to the original m4, m1 was missing from the table associated with the relevant graph, and if m4 after analysis exceeds the highest replicate of the vehicle data by more than 10%, m4 was forced to be the average of the vehicle data, and m4 was missing from the table associated with the relevant graph). All replicates of % infection in the absence (vehicle) or presence of the indicated drug at the indicated concentration were included in the analysis.

[0126] T.C. 50 Concentration determination The percentage of cytotoxicity was calculated using the following formula:

number

[0127] Maximum cytotoxicity is TC 50 If the value is not reached, TC 50 The value > the maximum concentration tested is reported.

[0128] result Table 1 shows the EC 50 , T.C. 50 , and the selection index (SI(=TC 50 / EC 50 ), as well as the EC for the positive control inhibitor monensin.50 Indicates the value.

[0129] Inhibition of all flaviviruses (i.e., four dengue virus strains, two Zika virus strains, and a yellow fever virus strain) was evident with both dextran sulfate and the positive control drug, with the EC 50 is in the range of approximately 0.03 to 0.6 mg / ml (30 to 600 μg / ml).

[0130] Low sub-TC at the highest concentration tested (10 mg / ml) 50 No cytotoxicity was observed at any of the dextran sulfate concentrations tested, except at the 100% level (maximum cytotoxicity of 16%). [Table 2]

[0131] Dengue virus Dengue fever is a viral infection transmitted by mosquitoes (usually species Aedes aegypti and Aedes albopictus) and is widespread in many parts of the world (Asia, Africa, America, Australia, and the Caribbean). Dengue infections arise from four antigenically distinct dengue virus types, designated DENV1, DENV2, DENV3, and DENV4. They belong to the Flavivirus genus, which comprises the most numerous arboviral diseases in humans in terms of geographic distribution, morbidity, and mortality. At least 2.5 billion people are at risk of the disease, and it is estimated that there are 100 to 400 million cases of dengue infection each year. Symptoms are usually mild (a mild febrile illness known as "dengue fever"), but an estimated 500,000 cases each year become very severe and life-threatening ("severe dengue fever" also known as dengue hemorrhagic fever) requiring hospitalization, and approximately 2.5% of affected individuals die. According to the WHO, severe dengue is a leading cause of hospitalization and death among children and adults in Asian and Latin American countries.

[0132] There is no drug treatment or any widely available safe vaccine against dengue, so prevention of dengue relies on effective vector control.

[0133] This assay assessed the ability of each of the four dengue virus types (1–4) to infect Huh-7 cells (a human epithelial tumor cell line) and the ability of dextran sulfate to prevent infection. Dextran sulfate demonstrated potent inhibition of infection by all four dengue virus types, with IC 50 The values ​​were below 0.6 mg / ml (Figures 4A-4D). The assay was validated by inhibition of infectivity from each of the four dengue virus variants by monensin, an ionophore with antiviral activity.

[0134] Data on dextran sulfate supported the potential of this agent to reduce human infection with dengue virus.

[0135] Yellow fever virus Yellow fever is another serious arbovirus disease caused by a flavivirus infection spread by mosquitoes (mainly Aedes aegypti and other Aedes species) that occurs in Africa, South America, Central America, and the Caribbean. Yellow fever is an acute hemorrhagic disease. The "yellow" describes the jaundice that affects some patients. Other symptoms include fever, headache, muscle pain, nausea, vomiting, and fatigue, and in severe cases can be fatal.

[0136] This assay assessed the ability of yellow fever virus to infect Huh-7 cells (a human epithelial tumor cell line) and the ability of dextran sulfate to prevent infection. Dextran sulfate demonstrated potent inhibition of infection, with IC 50 values ​​were less than 0.05 mg / ml (Fig. 4E). The assay was validated by inhibition of infectivity by monensin, an ionophore with antiviral activity.

[0137] Data on dextran sulfate supported the potential of this agent to reduce human infection with yellow fever virus.

[0138] Zika virus Like the Dengue virus, the flavivirus Zika is transmitted by mosquitoes (again, usually species Aedes aegypti and Aedes albopictus) and occurs in association with tropical and subtropical malaria regions of the world (e.g., Africa, Central and South America, South and Southeast Asia). There are two main Zika virus lineages, known as the African lineage and the Asian lineage.

[0139] While Zika virus disease is mild and temporary for most people, the disease can have devastating consequences for pregnant women, who may transmit the virus to their fetus. This may precipitate miscarriage or congenital abnormalities, such as a potentially fatal abnormally small head (microcephaly) as well as congenital Zika syndrome, a developmental disorder in which the brain does not function properly. In 2015–2016, there was a large outbreak in Brazil alone with over 200,000 cases, and 8,000 children were born with congenital abnormalities. It is currently estimated that 5–10% of pregnancies with confirmed Zika virus infection result in congenital abnormalities, increasing to 8–15% if confirmed infection is limited to the first trimester. In addition, there is an increasing understanding that a low percentage of patients develop neurological problems, such as Guillain-Barré syndrome, neuropathy, or myelitis.

[0140] There is no current drug therapy to mitigate Zika virus infection, and prevention relies on precautions to reduce exposure to mosquitoes. Infection is suspected based on symptoms and confirmed by laboratory testing.

[0141] This assay assessed the ability of both Zika virus strains (Asian and African) to infect HeLa cells (a human epithelial tumor cell line) and the ability of dextran sulfate to prevent infection. Dextran sulfate demonstrated potent inhibition of infection by the two Zika virus strains, with IC 50 The values ​​were less than 0.140 mg / ml (Figures 4G-4H). The assay was validated by inhibition of infectivity from both mutants by monensin, an ionophore with antiviral activity.

[0142] Data on dextran sulfate supported the potential for this agent to reduce human infection with Zika virus.

[0143] conclusion The ability of dextran sulfate to inhibit the infectivity of the flaviviruses Dengue virus (four major types), Zika virus (two major types), and Yellow Fever virus was evaluated. See summary of results in Figure 5 for Zika Asia virus (ZIKV), Dengue Hawaii virus (DENV), and YF17D.

[0144] Dextran sulfate demonstrated a strong ability to inhibit the infectivity of all flaviviruses tested. In addition, the general cytotoxicity of dextran sulfate was also evaluated in the same assays, and no worrisome levels were evident in any of the assays. The data support the potential of dextran sulfate as a treatment to prevent human infection with potentially devastating flaviviruses for which there is currently no drug therapy.

[0145] The above-described embodiments should be understood as some illustrative examples of the present invention. As will be understood by those skilled in the art, various modifications, combinations and variations may be made to the embodiments without departing from the scope of the present invention. In particular, different part solutions in different embodiments may be combined in other configurations, if technically possible.

Claims

1. Use of dextran sulfate or a pharma- ceutical acceptable salt thereof in the manufacture of a medicament for the prevention, inhibition, or treatment of a flavivirus infection or infectious disease, wherein the dextran sulfate or the pharma-ceutical acceptable salt thereof has an average molecular weight of 10,000 Da or less and an average sulfur content of 15% or more.

2. 2. The use according to claim 1, wherein the flavivirus infection is selected from the group consisting of flavivirus infections caused by flaviviruses selected from the group consisting of Dengue virus, Japanese encephalitis virus, Kunjin virus, Langat virus, Louping ill virus, Murray Valley encephalitis virus, St. Louis encephalitis virus, Powassan virus, West Nile virus, Yellow fever virus, and Zika virus, preferably selected from the group consisting of Dengue virus, Japanese encephalitis virus, Kunjin virus, Louping ill virus, Murray Valley encephalitis virus, St. Louis encephalitis virus, Powassan virus, West Nile virus, Yellow fever virus, and Zika virus.

3. 3. The use according to claim 2, wherein the flavivirus infection is selected from the group consisting of flavivirus infections caused by a flavivirus selected from the group consisting of dengue virus, yellow fever virus, and Zika virus.

4. The use according to claim 3 , wherein the flavivirus infection is a flavivirus infection caused by Zika virus.

5. The use according to claim 3, wherein the flavivirus infection is a flavivirus infection caused by a dengue virus.

6. The use according to claim 3, wherein the flavivirus infection is a flavivirus infection caused by yellow fever virus.

7. The use according to claim 1, wherein the flavivirus infection is caused by a flavivirus other than Japanese encephalitis virus, West Nile virus, dengue virus, and yellow fever virus.

8. 2. The use of claim 1, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof is formulated for systemic administration to a subject.

9. 9. The use according to claim 8, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof is formulated for intravenous or subcutaneous administration to the subject, preferably formulated for subcutaneous administration to the subject.

10. The use according to claim 1, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof has an average sulfur content in the range of 15-20%.

11. The use according to claim 1, wherein the average molecular weight is in the range of 2000 to 10000 Da, preferably in the range of 3000 to 10000 Da, more preferably in the range of 3500 to 9500 Da.

12. The use according to claim 11, wherein said average molecular weight is in the range of 4500 to 7500 Da, preferably in the range of 4500 to 5500 Da.

13. 2. The use according to claim 1, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof has a number average molecular weight (Mn) measured by nuclear magnetic resonance (NMR) spectroscopy in the range of 1850 to 3500 Da, preferably in the range of 1850 to 2500 Da, more preferably in the range of 1850 to 2300 Da.

14. The use according to claim 13, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof has a Mn in the range of 1850-2000 Da as measured by NMR spectroscopy.

15. The use according to claim 1, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof has an average number of sulfates per glucose unit in the range of 2.5 to 3.0, preferably in the range of 2.5 to 2.8, more preferably in the range of 2.6 to 2.

7.

16. 2. The use according to claim 1, wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof has an average of 5.1 glucose units and an average number of sulfates per glucose unit of 2.6 to 2.

7.

17. The use according to claim 1 , wherein the dextran sulfate or the pharma- ceutically acceptable salt thereof is formulated as an aqueous injection solution.

18. The use according to claim 1, wherein the pharma- ceutically acceptable salt is the sodium salt of dextran sulfate.

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