Acetylated sialic acid glycoclusters and their use for treating infectious diseases - Patents.com

Abstract

The present invention relates to a glycocluster comprising at least two acetylated sialic acids covalently bound to a macrocycle, said macrocycle being selected from porphyrins, pillararenes, calixarenes, and fullerenes. According to one embodiment, each acetylated sialic acid is independently selected from 4-O-acetylated sialic acid, 7-O-acetylated sialic acid, 8-O-acetylated sialic acid, and 9-O-acetylated sialic acid. The present invention also relates to a glycocluster for use in the treatment and / or prevention of infectious diseases, such as coronavirus infections.

Description

[Technical field]

[0001] The present invention is in the field of infectious disease treatment and relates to novel glycoclusters containing multiple acetylated sialic acids for use in the treatment of infectious diseases, in particular SARS-CoV-2 infections. [Background technology]

[0002] Coronaviruses (CoVs) are viruses of the family Coronaviridae, subfamily Orthocoronavirinae. They are enveloped viruses with a positive-sense, single-stranded ribonucleic acid (RNA) genome and a helical nucleocapsid. Their name is due to their unique morphology, i.e., a series of club-shaped spikes protruding from the surface of their envelope, giving them a crown-like appearance. Coronaviruses are also characterized by unusually large RNA genomes and a specific replication strategy. RNA viruses, particularly coronaviruses, are responsible for a wide range of respiratory, systemic, gastrointestinal, and neurological diseases in mammals and birds.

[0003] Coronaviruses were first identified in humans about 50 years ago in the UK and the USA. Since then, they have generally only been considered to cause mild infectious respiratory illnesses, e.g., the common cold. At the beginning of the 20th century, two highly pathogenic coronaviruses were first identified: Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) and Middle East Respiratory Syndrome Coronavirus (MERS-CoV). In contrast to previous coronavirus infections, SARS and MERS were severe respiratory illnesses that caused hundreds of deaths. At the end of 2019, a novel infectious respiratory illness emerged in Wuhan (China), the cause of which was eventually identified as the novel human coronavirus SARS-CoV-2. The genome of SARS-CoV-2 is about 80% identical to that of SARS-CoV and about 96% identical to that of the bat coronavirus BatCoV RaTG13. The disease caused by SARS-CoV-2 infection has been named "coronavirus disease 2019" ("COVID-19"). COVID-19 rapidly emerged as a severe global pandemic in 2020.

[0004] SARS-CoV-2 infection is thought to be asymptomatic in a significant number of infected individuals. In symptomatic patients, COVID-19 generally (approximately 80%) manifests as a respiratory illness of mild severity, including fever, cough or other respiratory symptoms (such as mild shortness of breath or chest tightness), headache, fatigue or muscle pain, and loss of smell and taste. However, some patients (approximately 15%) develop severe symptoms, including dyspnea, hypoxia, or pneumonia ("COVID-19 pneumonia"). In a minority of cases (approximately 5%), severe symptoms such as respiratory failure, shock, or multiple organ failure are observed. It was estimated that approximately 20% of COVID-19 patients require hospitalization, and some of them (approximately 5%) must be admitted to an intensive care unit (ICU). Permanent damage to organs has been observed in some cases and some patients continue to suffer from various effects even months after recovery (“long COVID”). COVID-19 therefore causes considerable suffering and death and also puts at risk many healthcare systems around the world.

[0005] Several vaccines against SARS-CoV-2 have been approved and distributed to various countries. In contrast, although research is ongoing to develop drugs that inhibit the virus for preventive and / or therapeutic purposes, the first-line treatment of COVID-19 remains symptomatic treatment, as well as supportive measures such as supportive care and isolation. Moreover, some mass vaccination campaigns have faced significant shortcomings in many countries, leaving many people vulnerable to COVID-19 in the near future.

[0006] therefore, There remains a need for effective treatment and / or prevention of coronavirus infections, particularly respiratory coronavirus infections that cause diseases such as SARS, MERS, or COVID-19. In particular, there is a need for therapeutic agents that are highly efficient against coronavirus replication, with few or no significant adverse effects, good chemical stability, and / or low cost.

[0007] Entry of SARS-CoV into host cells is mediated by its transmembrane spike (S) glycoprotein, which forms a homotrimer that protrudes from the viral surface. The S glycoprotein contains two functional subunits, either responsible for binding to the host cell receptor (the S1 subunit, which contains the receptor-binding domain (RBD)) or for fusion of the virus with the cellular membrane (the S2 subunit). Angiotensin-converting enzyme 2 (ACE2), previously identified as the cellular receptor for SARS-CoV, also acts as a receptor for the novel coronavirus (SARS-CoV-2). In the case of SARS-CoV-2, the S glycoprotein on the virion surface mediates receptor recognition and membrane fusion. Recently, a high-resolution cryo-electron microscopy structure obtained for full-length human ACE2 in the presence of the RBD of the S glycoprotein of SARS-CoV-2 suggested simultaneous binding of two S glycoprotein trimers to the ACE2 dimer. The S2 subunit is further cleaved by host proteases located immediately upstream of the fusion peptide, resulting in activation of the glycoprotein, which undergoes extensive irreversible conformational changes that facilitate the membrane fusion process.

[0008] The binding of S protein to ACE2 has already been extensively studied, and there is a consensus on its central role in infection, with several studies suggesting an important role for other cell surface molecules co-receptors / adhesins. In particular, it was recently suggested that 9-O-acetylsialoglycan, in addition to ACE2, may be involved in the early steps of SARS-CoV-2 binding to cells (Yang, J. et al., Nature Communications, 2020, Vol. 11, Paper No.: 4541). Binding of acetylated sialic acid to coronavirus S glycoprotein and coronavirus hemagglutinin esterase has also been reported (Tortorici, MA et al., Nature Structural Molecular Biology, 2019, Vol. 26, pp. 481-489). Therefore, based on this preliminary information, it is believed that sialic acid derivatives may act as competitive inhibitors that block the interaction between the SARS-CoV-2 spike protein S and host cells, which normally mediates the first step of infection (Tortorici, MA et al., Nature Structural Molecular Biology, 2019, Vol. 26, pp. 481-489).

[0009] however, It has been suggested that sialic acid and its acetylated derivatives may bind, at least weakly, to the spike protein of SARS-CoV-2, but there is no direct quantitative evidence to define which of these two carbohydrates is the optimal partner (Yang, J. et al., Nature Communications, 2020, Vol. 11, Paper No.: 4541; Nguyen, K. et al., Viruses, May 2021, Vol. 13, No. 5, p. 927).

[0010] Moreover, it is known in the art that individual glycans, such as sialic acid or its derivatives, generally have relatively low affinity, typically at best moderate, for their protein targets (Sauter, NK et al., Biochemistry 1989, Vol. 28, p. 8388; Dormitzer, PR et al., Journal of Virology, October 2002, Volume 76, No. 20, pp. 10512-10517). This is further demonstrated in the examples of the present application. The limited affinity of individual sialic acids and their derivatives makes such glycans unsuitable for direct use in the actual treatment and / or prevention of viral infections.

[0011] Surprisingly, the applicant has demonstrated that when at least two sialic acids or their derivatives are bound to certain macrocycles (such as porphyrins, pillararenes, calixarenes, and fullerenes), the resulting glycoclusters are strong competitors for the initial attachment of SARS-CoV-2 to the host cell, despite the low affinity of sialic acids. Thus, in sharp contrast to sialic acids not bound to macrocycles ("free" sialic acids), the glycoclusters of the invention are suitable for use in the treatment and / or prevention of infectious diseases such as COVID-19. The present invention therefore paves the way for novel drugs against viral infections such as COVID-19. Summary of the Invention

[0012] The present invention relates to a glycocluster comprising at least two acetylated sialic acids covalently bound to a macrocycle, said macrocycle being selected from porphyrins, pillararenes, calixarenes, and fullerenes, and each acetylated sialic acid being independently selected from 4-O-acetylated sialic acid, 7-O-acetylated sialic acid, 8-O-acetylated sialic acid, and 9-O-acetylated sialic acid.

[0013] According to one embodiment, the glycocluster comprises at least four acetylated sialic acids covalently attached to said macrocycle. According to one embodiment, each acetylated sialic acid is independently selected from 7-O-acetylated sialic acid and 9-O-acetylated sialic acid. In one embodiment, each acetylated sialic acid is 9-O-acetylated sialic acid.

[0014] According to one embodiment, the macrocycle is selected from porphyrins, preferably from [Zn(tetraphenylporphyrin)] and tetraphenylporphyrin. According to one embodiment, the glycocluster further comprises at least one angiotensin-converting enzyme 2 (ACE2) binding inhibitor, preferably the angiotensin-converting enzyme 2 (ACE2) binding inhibitor is selected from an ACE2 binding inhibitor peptide, an ACE2 binding inhibitor protein, and an anti-ACE2 antibody or an antigen-binding fragment thereof.

[0015] According to one embodiment, the glycocluster has the formula (I), (Ia), (II), (III) or (IV): [ka] or a pharma- ceutically acceptable salt and / or solvate thereof, In the formula, each L 1 is a linker independently selected from -a single bond, or -alkyl, heteroalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, heteroaryl, heteroarylalkyl, and alkylheteroaryl, wherein said alkyl, heteroalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, heteroaryl, heteroarylalkyl, or alkylheteroaryl optionally comprises at least one coupling product, and each R 1 are independently selected from acetylated sialic acid and ACE2 binding inhibitors, provided that at least two R 1 is an acetylated sialic acid and M is a metal cation.

[0016] According to one embodiment, each L 1 is a linker selected from alkyl, heteroalkyl, alkylaryl, arylalkyl, heteroarylalkyl, and alkylheteroaryl, wherein said alkyl, heteroalkyl, alkylaryl, arylalkyl, heteroarylalkyl, or alkylheteroaryl comprises one or two coupling products, preferably at least one of said coupling products is triazolyl.

[0017] According to one embodiment, each L 1 -R 1 is -(CH2) m -R C -(CH2) n -R 1 , -O-(CH2) m -R C -(CH2) n -R 1 , -C(O)-(CH2) m -R C -(CH2) n -R 1 , -C(O)O-(CH2) m -R C -(CH2) n -R 1 , -phenyl-(CH2) m -R C -(CH2) n -R 1 , and -phenyl-O-(CH2) m -R C -(CH2) n -R 1 wherein R C is a coupling product, preferably triazolyl, m and n are independently integers ranging from 1 to 8, preferably integers ranging from 1 to 4, and each R 1 is as defined above. According to one embodiment, each R 1 is a 9-O-acetylated sialic acid.

[0018] According to one embodiment, the glycocluster has the formula (011), (005), (008) or (014): [ka] JPEG2024533417000004.jpg131159, or a pharma- ceutically acceptable salt or solvate thereof; In the formula, each R 1 is represented by the formula (9-AcSA) [ka] where the wavy line represents the R 1 Represents the attachment point of

[0019] The present invention also relates to a pharmaceutical composition comprising a glycocluster according to the invention and at least one pharma- ceutically acceptable carrier.The present invention also relates to a glycocluster according to the invention or a pharmaceutical composition according to the invention for use as a medicament.The present invention also relates to a glycocluster according to the invention or a pharmaceutical composition according to the invention for use in the treatment and / or prevention of an infectious disease, preferably a coronavirus or picornavirus infection, more preferably a SARS-CoV-2 infection.

[0020] The present invention also relates to a process for producing the glycoclusters according to the invention, comprising a step of coupling of each of the acetylated sialic acids with a macrocycle, preferably a coupling step in which the reaction between a terminal alkyne and an azide leads to the formation of a triazolyl group.

[0021] definition In the present invention, the following terms have the following meanings.

[0022] chemical definition When describing the compounds of the present invention, the terms used should be construed according to the following definitions unless otherwise indicated. When a chemical substituent is a combination of chemical groups, the point of attachment of the substituent to the molecule is through the last chemical group listed. For example, an arylalkyl substituent is attached to the remainder of the molecule through the alkyl moiety and can be represented as: "aryl-alkyl-".

[0023] "Acetyl" or "ethanoyl", represented by the symbol "Ac", refers to a methyl acyl moiety of the formula CH3-C(O)-.

[0024] "Alkene" or "alkenyl" refers to a straight or branched hydrocarbon chain containing at least one double bond and typically from 2 to 12 carbon atoms, preferably from 3 to 6 carbon atoms. Non-limiting examples of alkenyl groups include ethenyl, 2-propenyl, 2-butenyl, 3-butenyl, 2-pentenyl and its isomers, 2-hexyl and its isomers, and 2,4-pentadienyl.

[0025] "Alkyl" refers to a saturated straight or branched hydrocarbon chain, typically containing 1 to 12 carbon atoms, preferably 1 to 6 carbon atoms, more preferably 1 to 3 carbon atoms. In the present invention, an alkyl group can be monovalent or divalent (i.e., "alkylene" groups are included in the definition of "alkyl"). Non-limiting examples of alkyl groups include methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl and t-butyl, pentyl and its isomers (e.g., n-pentyl, isopentyl), and hexyl and its isomers (e.g., n-hexyl, isohexyl). Preferred alkyl groups include methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl and t-butyl.

[0026] "Alkylaryl" refers to an aryl group substituted with an alkyl group: alkyl-aryl-.

[0027] "Alkylheteroaryl" refers to a heteroaryl group substituted with an alkyl group: alkyl-heteroaryl-.

[0028] "Alkyne" or "alkynyl" refers to a straight or branched hydrocarbon chain containing at least one triple bond and typically containing from 2 to 12 carbon atoms, preferably from 3 to 6 carbon atoms. Examples of alkynyl groups include ethynyl, 2-propynyl, 2-butynyl, 3-butynyl, 2-pentynyl and its isomers, and 2-hexynyl and its isomers.

[0029] "Amide" refers to a functional group having a -(C=O)-NH- connectivity.

[0030] "Amido" refers to the group -(C=O)-NH2.

[0031] "Amine" refers to the -NH2 group and to secondary amines -NHR, where R is different from hydrogen, and preferably R is an alkyl group.

[0032] "Amino" refers to the group -NH2.

[0033] "Aminooxy" refers to the group -O-NH2.

[0034] "Aryl" refers to a cyclic polyunsaturated aromatic hydrocarbyl group containing at least one aromatic ring. An aryl group can have a single ring (i.e., phenyl) or multiple aromatic rings fused together (e.g., naphthyl) or covalently bonded. Typically, an aryl group has 5 to 12 carbon atoms, preferably 6 to 10 carbon atoms. The aromatic ring may optionally contain 1 to 2 additional rings (either cycloalkyl, heterocycloalkyl, or heteroaryl) fused thereto. Aryl is also intended to include the partially hydrogenated derivatives of the carbocyclic ring systems enumerated herein, so long as at least one ring is aromatic. Non-limiting examples of aryl groups include phenyl, biphenyl, biphenylenyl, 5- or 6-tetralinyl, naphthalene-1- or -2-yl, 4-, 5-, 6- or 7-indenyl, 1-, 2-, 3-, 4- or 5-acenaphthylenyl, 3-, 4- or 5-acenaphthenyl, 1- or 2-pentalenyl, 4- or 5-indanyl, 5-, 6-, 7- or 8-tetrahydronaphthyl, 1,2,3,4-tetrahydronaphthyl, 1,4-dihydronaphthyl, 1-, 2-, 3-, 4- or 5-pyrenyl. A preferred aryl group is phenyl.

[0035] "Arylalkyl" refers to an alkyl group substituted by an aryl group: aryl-alkyl-.

[0036] "Azide" refers to a group of the formula -N3.

[0037] "(C x ~C y )" means, in accordance with common terminology in the chemical art, that the group contains x to y carbon atoms.

[0038] "Carboxylic acid" refers to a group of the formula --COOH.

[0039] "Coupling functional group" refers to a functional group that can react with another functional group to form a covalent bond, such as a bond or a linear group of atoms. A coupling functional group that is reactive under suitable reaction conditions can therefore chemically react with another coupling functional group on a different molecule to form a new covalent bond. A coupling functional group generally represents a point of attachment to another molecule. Coupling functional groups generally include nucleophiles, electrophiles, and / or photoactivatable groups. Non-limiting examples of coupling functional groups include alcohols; alkenes; alkynes (e.g., -C≡CH); amines; amides; aminooxy; anhydrides, such as glutaric anhydride, succinic anhydride, or maleic anhydride; azides; carboxylic acids; activated carboxylic acids, such as acid anhydrides or acid halides; chloroformic acid; activated esters, such as N-hydroxysuccinimide ester, N-hydroxyglutarimide ester, or maleimide ester; glutamic acid; halides (halogen atoms); haloacetamides, such as chloroacetamide, bromoacetamide, or iodoacetamide (i.e., -NH-C(O)CH2X moieties, where X is a halogen atom); hydrazides; isocyanates; isothiocyanates; ketones; maleimides; norbornene; phosphonic acids; siloxy; tetrazines, and thiols. The reaction between two coupling functional groups can result in a "coupling product" as defined herein.

[0040] "Coupling product" refers to the residue of a coupling functional group resulting from the reaction between two coupling functional groups in different molecules, such as a functionally related group of atoms (such as an amide-C(O)-NH- group or a double bond), or a heterocycle (such as a divalent triazolyl group). In other words, a coupling product is the remaining portion of one or two coupling functional groups after a coupling reaction between two coupling functional groups. For example, a coupling reaction between two coupling functional groups A and B can result in the following coupling product, as shown in Table 1 below, where X represents a halogen atom (e.g., Br or Cl). The coupling product may be included in the "linker" as defined herein.

[0041] [Table 1]

[0042] "Covalently linked" means that two moieties are covalently linked together either directly, i.e., by a single, double, or triple covalent bond (typically a single bond), or indirectly, i.e., by a "linker," as described herein, that includes multiple covalent bonds.

[0043] "Glycocluster" refers to a cluster of glycans, i.e., a molecule or ensemble of molecules that contains multiple glycan units. Thus, a glycocluster contains at least two polysaccharide, oligosaccharide, and / or monosaccharide moieties, typically at least two monosaccharides. In a glycocluster, the glycan units are clustered together by their attachment to a common scaffold (e.g., a macrocycle, polymer, or metal nanoparticle) and are in relatively close proximity to each other. Glycoclusters are often used for drug delivery, but in the present invention, they can be used as inhibitors of cell binding and / or infectivity for use in treating infectious diseases.

[0044] "Halide," "halo," or "halogen" refers to a fluorine, chlorine, bromine, or iodine atom, preferably a chlorine or bromine atom.

[0045] "Heteroalkyl" refers to an alkyl group as defined above, in which one or more carbon atoms are replaced by a heteroatom selected from oxygen, nitrogen, and sulfur. In a heteroalkyl group, the heteroatoms are only bonded to carbon atoms along the alkyl chain, i.e., each heteroatom is separated from other heteroatoms by at least one carbon atom. The nitrogen and sulfur heteroatoms can be optionally oxidized, and the nitrogen heteroatom can be optionally quaternized. A heteroalkyl is only bonded to another group or molecule through a carbon atom, i.e., the adjacent atom is not selected from among the heteroatoms contained in the heteroalkyl group. Non-limiting examples of heteroalkyl include alkoxy, ethers and polyethers, secondary amines, tertiary amines, and thioethers.

[0046] "Heteroaryl" refers to an aromatic ring or ring system containing 5-12 carbon atoms, preferably 6-10 carbon atoms, having one or two rings fused or covalently bonded together, where at least one ring is aromatic and one or more carbon atoms in one or more of the rings are replaced by oxygen, nitrogen, and / or sulfur atoms. "Heteroaryl" may also be considered an "aryl" group as defined herein, where at least one carbon atom in the aryl group is replaced with a heteroatom, and the resulting molecule is chemically stable. The nitrogen and sulfur heteroatoms may be optionally oxidized, and the nitrogen heteroatom may be optionally quaternized.Non-limiting examples of heteroaryl groups include furanyl, thiophenyl, pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, triazolyl, oxadiazolyl, thiadiazolyl, tetrazolyl, oxatriazolyl, thiatriazolyl, pyridinyl, pyrimidyl, pyrazinyl, pyridazinyl, oxazinyl, dioxinyl, thiazinyl, triazinyl, imidazo[2,1-b][1,3]thiazolyl, thieno[3,2-b]furanyl, thieno[3,2-b]thiophenyl, thieno[2,3-d][1,3]thiazolyl, thieno[2,3-d]imidazolyl, tetrazolo[1,5-a]pyridinyl, indolyl, indolizinyl, isoindolyl, benzofuranyl, isobenzofuranyl, benzothiophenyl, isobenzothiophenyl, indyl, Zolyl, benzimidazolyl, 1,3-benzoxazolyl, 1,2-benzisoxazolyl, 2,1-benzisoxazolyl, 1,3-benzothiazolyl, 1,2-benzisothiazolyl, 2,1-benzisothiazolyl, benzotriazolyl, 1,2,3-benzoxadiazolyl, 2,1,3-benzoxadiazolyl, 1,2,3-benzothiadiazolyl, 2,1,3-benzothia These include diazolyl, thienopyridinyl, purinyl, imidazo[1,2-a]pyridinyl, 6-oxo-pyridazin-1(6H)-yl, 2-oxopyridin-1(2H)-yl, 6-oxo-pyridazin-1(6H)-yl, 2-oxopyridin-1(2H)-yl, 1,3-benzodioxolyl, quinolinyl, isoquinolinyl, cinnolinyl, quinazolinyl, and quinoxalinyl.

[0047] "Heteroarylalkyl" refers to an alkyl group substituted by a heteroaryl group: heteroaryl-alkyl-.

[0048] "Hydroxyl" refers to the -OH group.

[0049] "Ketone" refers to a functional group having a C-(C=O)-C connectivity.

[0050] "Linker" refers to a moiety that covalently links two molecules to one another and includes a series of polyvalent atoms selected from C, N, O, S, and P bound together by a stable covalent bond. The moiety typically incorporates 1 to 30 atoms, such as, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, or 30. Linkers can be linear or nonlinear, and some linkers have pendant side chains, or pendant functional groups, or both. In one embodiment, the linker is composed of any combination of single, double, triple, or aromatic carbon-carbon, carbon-nitrogen, nitrogen-nitrogen, carbon-oxygen, and carbon-sulfur bonds. In one embodiment, the linker is composed of a combination of moieties selected from alkyl, -C(O)NH-, -C(O)O-, -NH-, -S-, -O-, -C(O)-, -S(O)-, -S(O)2, and 5- or 6-membered monocyclic aryl or heteroaryl. In one embodiment, the linker comprises at least one "coupling product" as defined herein, typically one or two coupling products. In this context, "comprise" means that the linker can be interrupted by at least one coupling product (i.e., the coupling product is incorporated into the atom chain of the linker) and / or that the linker ends with at least one coupling product (i.e., the coupling product terminates the linker). In this embodiment, the coupling product is considered to be part of the linker, which specifically means that the atoms of the coupling product are counted in the total atom count of the linker.

[0051] "Macrocycle" refers to a molecule or ion containing 12 or more membered rings, such as pillararenes, calixarenes, porphyrins, fullerenes, crown ethers, and cyclodextrins. For brevity and clarity, in this application, the term "macrocycle" and its specific types (e.g., pillararenes, calixarenes, porphyrins, and fullerenes) refer to both the macrocycle itself and any macrocycle-based moiety resulting from the coupling of the macrocycle to another molecule, either directly by a single bond or through a "linker" as defined herein. Where appropriate, the latter will be specifically referred to as a "residue of a macrocycle" or a "macrocycle residue."

[0052] "Peptide" refers to a linear polymer of less than 50 amino acids joined together by peptide bonds.

[0053] "Sialic acid" or "SA" refers to a monosaccharide belonging to the class of alpha-keto acid sugars having a nine-carbon backbone, as commonly understood in the art. Specifically, sialic acid refers to acetylneuraminic acid (Neu5Ac) of the following formula: [ka]

[0054] The configuration in which the carboxylic acid is in the axial position is the α-anomer, while the configuration in which the carboxylic acid is in the equatorial position is the β-anomer. When bound to another molecule, sialic acid is predominantly the α-anomer. The carbon numbering of sialic acid is as shown in the formula above (left: α-anomer, right: β-anomer).

[0055] Sialic acid may be "acetylated" as commonly understood in the art, i.e., a group (typically a hydrogen atom) in sialic acid may be replaced by an acetyl group. Although Neu5Ac includes an acetylated amine (AcHN), in this application, only sialic acids in which acetylation has occurred at both the hydroxyl (OH) group and the amine are considered to be "acetylated" in the sense of the present invention. Specifically, an "acetylated sialic acid" is a sialic acid in which at least one hydroxyl (-OH) at positions 4, 7, 8, and 9 is replaced by an acetyl group, thereby resulting in an acetic acid moiety (-OC(O)CH3). Synthetic methods for acetylation of one or more positions in sialic acid, including esterification reactions with acetic acid, are well known in the art. Specifically, acetylated sialic acid may refer to 9-O-acetyl-sialic acid (9-AcSA) of the following formula (left: α-anomer, right: β-anomer). [ka]

[0056] Sialic acid or acetylated sialic acid, whether natural or artificial, can be bound to another compound directly by a single covalent bond or through a linker. For example, in the present invention, sialic acid and / or acetylated sialic acid are bound to macrocycles to form glycoclusters. Non-acetylated hydroxyls can be used as coupling functional groups, in particular, to obtain covalent bonds. For the sake of brevity and clarity, in this application, the term "sialic acid" and its specific types (e.g., acetylated sialic acid) refer to both the sialic acid monosaccharide itself and any sialic acid-based moiety resulting from the coupling of sialic acid to another molecule, either directly by a single bond or through a "linker" as defined herein. Where appropriate, the latter will be specifically referred to as "residue of sialic acid" or "sialic acid residue". When sialic acid is bound to another molecule by a non-acetylated hydroxyl (e.g., hydroxyl at position 2), the remaining oxygen is considered to be part of the sialic acid residue for purposes of definition and expression.

[0057] "Siloxy" refers to the functional group -O-Si(R)3, where R represents, for example, alkyl or aryl.

[0058] "Triazolyl" has the general formula [ka] “H” refers to monovalent, divalent, or trivalent derivatives of heteroaryl (i.e., triazole) of the formula: [ka]

[0059] general definition "About" is used herein to mean approximately, roughly, around, or in the region of. The term "about" preceding a number means ±10% of the value of the number. When the term "about" is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the stated numerical values ​​by 10%.

[0060] "Administration" or variations thereof (e.g., "administering") means providing a therapeutic agent (e.g., a compound of the invention), either alone or as part of a pharma- ceutically acceptable composition, to a patient whose condition, symptom, or disease is to be treated and / or prevented.

[0061] "Human" refers to a male or female subject at any stage of development, including neonates, infants, juveniles, adolescents, and adults.

[0062] "Patient" refers to an animal, typically a warm-blooded animal, preferably a human, awaiting medical care, or receiving medical care, or being / will be the subject of a medical procedure. A patient may also be a subject of preventive care or treatment.

[0063] "Pharmaceutically acceptable" means that the components of the composition are compatible with each other and not harmful to the patient to whom it is administered.

[0064] "Pharmaceutically acceptable carrier" refers to an excipient that does not produce adverse, allergic, or other untoward reactions when administered to animals, preferably humans. It includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. For human administration, preparations should meet sterility, pyrogenicity, general safety, and purity standards as required by regulatory authorities, such as FDA or EMA.

[0065] "Prevent," "preventing," and "prevention" refer to delaying or hindering the onset of a condition and / or disease and / or any one of its associated symptoms, barring a patient from contracting a condition or disease, or reducing a patient's risk of contracting a condition and / or disease and / or any one of its associated symptoms.

[0066] "Prodrug" refers to a pharmacologically acceptable derivative of a therapeutic agent (e.g., a compound of the present invention) whose in vivo biotransformation product is the therapeutic agent (active drug). Prodrugs are typically characterized by increased bioavailability and are readily metabolized in vivo to the active compound. Non-limiting examples of prodrugs include amide prodrugs and carboxylic acid ester prodrugs, particularly alkyl esters, cycloalkyl esters, and aryl esters.

[0067] "Solvate" refers to a molecular complex that contains a compound together with one or more molecules of one or more solvents in stoichiometric or substoichiometric amounts, typically the solvent is a pharma- ceutically acceptable solvent such as ethanol. The term "hydrate" refers to when the solvent is water (HO).

[0068] "Therapeutic agent", "active pharmaceutical ingredient" and "active ingredient" refer to a health-related compound for use in therapy. In particular, a therapeutic agent (e.g., a compound of the present invention) may be indicated for treating and / or preventing a disease, preferably an infectious disease. An active ingredient may also be indicated for improving the therapeutic activity of another therapeutic agent.

[0069] A "therapeutically effective amount" (briefly, "effective amount") refers to an amount of a therapeutic agent (e.g., a compound of the invention) sufficient to achieve a desired therapeutic or prophylactic effect in a patient to which it is administered.

[0070] "Treat," "treating," and "treatment" refer to alleviating, attenuating, or suppressing a condition and / or disease and / or any one of its associated symptoms, e.g., an infectious disease. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0071] Glycocluster The present invention relates to glycoclusters comprising at least two acetylated sialic acids covalently attached to a macrocycle. In this application, "glycoclusters" and "compounds of the invention" and similar terms are synonymous.

[0072] According to one embodiment, the glycocluster comprises at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 14, 15, or 16 acetylated sialic acids covalently attached to the macrocycle. In one embodiment, the glycocluster comprises at least 4 acetylated sialic acids covalently attached to the macrocycle. According to one embodiment, the glycocluster comprises 4 acetylated sialic acids covalently attached to the macrocycle (tetramer). According to one embodiment, the glycocluster comprises 10 acetylated sialic acids covalently attached to the macrocycle (decamer). According to one embodiment, the glycocluster comprises 12 acetylated sialic acids covalently attached to the macrocycle (dodecamer).

[0073] According to one embodiment, each acetylated sialic acid is not fully acetylated, i.e., at least one OH group at positions 4, 7, 8, and 9 is not substituted by acetyl. In one embodiment, each acetylated sialic acid is acetylated at one, two, three, or four positions among positions 4, 7, 8, and 9. In one embodiment, each acetylated sialic acid is acetylated at one, two, or three positions. In one embodiment, each acetylated sialic acid is acetylated at one or two positions. In one embodiment, each acetylated sialic acid is acetylated at positions 7 and 9.

[0074] According to one embodiment, each acetylated sialic acid comprises at least one OH group. In one embodiment, each acetylated sialic acid comprises one OH group. In one embodiment, each acetylated sialic acid comprises two OH groups. In one embodiment, each acetylated sialic acid comprises three OH groups. In one embodiment, each acetylated sialic acid comprises four OH groups.

[0075] According to one embodiment, each acetylated sialic acid is acetylated at only one position, i.e., each acetylated sialic acid is independently selected from 4-O-acetylated sialic acid, 7-O-acetylated sialic acid, 8-O-acetylated sialic acid, and 9-O-acetylated sialic acid. According to one embodiment, each acetylated sialic acid is independently selected from 7-O-acetylated sialic acid, 8-O-acetylated sialic acid, and 9-O-acetylated sialic acid. According to one embodiment, each acetylated sialic acid is independently selected from 7-O-acetylated sialic acid and 9-O-acetylated sialic acid. In one embodiment, each acetylated sialic acid is 4-O-acetylated sialic acid. In one embodiment, each acetylated sialic acid is 7-O-acetylated sialic acid. In one embodiment, each acetylated sialic acid is 8-O-acetylated sialic acid. In one embodiment, each acetylated sialic acid is a 9-O-acetylated sialic acid.

[0076] Without being bound by any theory, Applicants believe that sialic acids that are not fully acetylated, and in particular sialic acids in which only one OH is acetylated, are advantageous, particularly in terms of viral affinity or inhibition against SARS-CoV-2.

[0077] Typically, the macrocycles are selected from porphyrins, pillararenes, calixarenes, and fullerenes.

[0078] According to one embodiment, the macrocycle is selected from porphyrin. In one embodiment, the porphyrin is selected from porphin or tetraphenylporphyrin. In one embodiment, the porphyrin has a metal cation coordinated to four nitrogen atoms and has no hydrogen bonded to the nitrogen atom ("porphyrin chelate"). Non-limiting examples include [Zn(porphin)] and [Zn(tetraphenylporphyrin)]. In another embodiment, the porphyrin has no cation coordinated to four nitrogen atoms and has two hydrogen bonded to the nitrogen atom ("porphyrin free base").

[0079] According to one embodiment, the macrocycle is selected from pillararenes. In one embodiment, the pillararenes are selected from pillar[5]arenes.

[0080] According to one embodiment, the macrocycle is selected from calixarenes. In one embodiment, the calixarenes are selected from calix[4]arenes.

[0081] According to one embodiment, the macrocycle is selected from fullerenes. In one embodiment, the macrocycle is C, also known as Buckminsterfullerene. 60 -fullerenes.

[0082] In the present invention, the macrocycle can be unsubstituted (non-substituted) except for the acetylated sialic acid substituent. In the present invention, the macrocycle can include additional substituents, such as water-solubilizing substituents, or substituents that favor interaction with viruses.

[0083] According to one embodiment, the glycocluster comprises at least one angiotensin-converting enzyme 2 (ACE2) binding inhibitor. In one embodiment, the ACE2 binding inhibitor is an ACE2 binding inhibitor peptide or protein. In one embodiment, the ACE2 binding inhibitor is an ACE2 binding inhibitor peptide. In one embodiment, the ACE2 binding inhibitor is an ACE2 binding inhibitor protein. In one embodiment, the ACE2 binding inhibitor is an anti-ACE2 antibody or an antigen-binding fragment thereof, such as an anti-ACE2 monoclonal antibody. In one embodiment, the ACE2 binding inhibitor peptide is designed according to the sequence of the ACE2 receptor that forms a complex with the RBD domain of the S1 glycoprotein. In one embodiment, the ACE2 binding inhibitor peptide is selected from peptides [22-44] consisting of the amino acid sequence EEQAKTFLDKFNHEAEDLFYQSS (SEQ ID NO: 1), peptides [351-357] consisting of the amino acid sequence LGKGDFR (SEQ ID NO: 2), peptides [22-57] consisting of the amino acid sequence EEQAKTFLDKFNHEAEDLFYQSSLASWNYNTNITEE (SEQ ID NO: 3), and peptides [22-44-g-351-357] consisting of the amino acid sequence EEQAKTFLDKFNHEAEDLFYQSSGLGKGDFR (SEQ ID NO: 4), as described in reference: Yang, J. et al., Nature Communications, 2020, Vol. 11, paper number: 4541.

[0084] According to one embodiment, the glycocluster comprises at least one other substituent that is susceptible to binding to an S protein, such as an antibody, an antibody fragment, a nanobody or a lectin.

[0085] According to one embodiment, the glycocluster is a compound of formula (I). [ka]

[0086] In formula (I), M is a metal cation such as zinc, iron, copper, manganese, silver, gold, cobalt, nickel, tin, cadmium, lead, vanadium cation. In one embodiment, the metal cation has 2 or 3 positive charges, preferably 2 positive charges. In one embodiment, M is a metal cation selected from zinc (II), iron (II), iron (III), copper (II), copper (III), manganese (II), manganese (III), silver (II), silver (III), gold (III), cobalt (II), cobalt (III), nickel (II), nickel (III), tin (II), cadmium (II), lead (II), vanadium (II), and vanadium (III).

[0087] According to one embodiment, the glycocluster is a compound of formula (Ia). [ka]

[0088] In formulas (I) and (Ia), each L 1 is independently a single bond or a linker selected from alkyl, heteroalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, heteroaryl, heteroarylalkyl, and alkylheteroaryl, where alkyl, heteroalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, heteroaryl, heteroarylalkyl, or alkylheteroaryl optionally comprises at least one coupling product. 1is a linker selected from alkyl, heteroalkyl, alkylaryl, arylalkyl, heteroarylalkyl, and alkylheteroaryl, where the alkyl, heteroalkyl, alkylaryl, arylalkyl, heteroarylalkyl, or alkylheteroaryl comprises at least one coupling product. In one embodiment, the linker comprises one or two coupling products. In one embodiment, the linker comprises one coupling product (i.e., only one coupling product). In one embodiment, at least one of the coupling products is triazolyl.

[0089] In formulas (I) and (Ia), each R 1 are independently selected from acetylated sialic acid and ACE2 binding inhibitors, provided that at least two R 1 is acetylated sialic acid.

[0090] In one embodiment, each L 1 -R 1 is -(CH2) m -R C -(CH2) n -R 1 , -O-(CH2) m -R C -(CH2) n -R 1 , -C(O)-(CH2) m -R C -(CH2) n -R 1 , -C(O)O-(CH2) m -R C -(CH2) n -R 1 , -phenyl-(CH2) m -R C -(CH2) n -R 1 , and -phenyl-O-(CH2) m -R C -(CH2) n -R 1 wherein R Cis a coupling product, and m and n are independently integers ranging from 1 to 8. In one embodiment, R C is triazolyl. In one embodiment, m is an integer ranging from 1 to 6, preferably ranging from 1 to 4, more preferably ranging from 2 to 4, more preferably 2 or 3. In one embodiment, n is an integer ranging from 1 to 4, preferably ranging from 1 to 3, more preferably 1 or 2, more preferably 1.

[0091] In one embodiment, each L 1 -R 1 is -phenyl-O-(CH2) m -R C -(CH2) n -R 1 where R C is a coupling product, and m and n are independently integers ranging from 1 to 8.

[0092] In one embodiment, each R 1 is selected from 7-O-acetylated sialic acid and 9-O-acetylated sialic acid. 1 is a 7-O-acetylated sialic acid. 1 is a 9-O-acetylated sialic acid.

[0093] According to one embodiment, the glycocluster has the formula (II): [ka] wherein L 1 and R 1 is as defined above in formulas (I) and (Ia).

[0094] In one embodiment, each L 1 -R 1 is -O-(CH2) m -R C -(CH2) n -R 1 where R Cis a coupling product, and m and n are independently integers ranging from 1 to 8. In one embodiment, R C is triazolyl. In one embodiment, m is an integer ranging from 1 to 6, preferably ranging from 1 to 4, more preferably ranging from 2 to 4, more preferably 2 or 3. In one embodiment, n is an integer ranging from 1 to 4, preferably ranging from 1 to 3, more preferably 1 or 2, more preferably 1.

[0095] According to one embodiment, the glycocluster has the formula (III): [ka] wherein L 1 and R 1 is as defined above in formulas (I) and (Ia).

[0096] In one embodiment, each L 1 -R 1 is -O-(CH2) m -R C -(CH2) n -R 1 where R C is a coupling product, and m and n are independently integers ranging from 1 to 8. In one embodiment, R C is triazolyl. In one embodiment, m is an integer ranging from 1 to 6, preferably ranging from 1 to 4, more preferably ranging from 2 to 4, more preferably 2 or 3. In one embodiment, n is an integer ranging from 1 to 4, preferably ranging from 1 to 3, more preferably 1 or 2, more preferably 1.

[0097] According to one embodiment, the glycocluster has the formula (IV): [ka] wherein L 1 and R 1 is as defined above in formulas (I) and (Ia).

[0098] In one embodiment, each L 1 -R 1 is -C(O)O-(CH2) m -R C -(CH2) n -R 1 where R C is a coupling product, and m and n are independently integers ranging from 1 to 8. In one embodiment, R C is triazolyl. In one embodiment, m is an integer ranging from 1 to 6, preferably ranging from 1 to 4, more preferably ranging from 2 to 4, more preferably 2 or 3. In one embodiment, n is an integer ranging from 1 to 4, preferably ranging from 1 to 3, more preferably 1 or 2, more preferably 1.

[0099] According to one embodiment, the glycocluster has the formula (011): [ka] or a pharma- ceutically acceptable salt or solvate thereof.

[0100] According to one embodiment, the glycocluster has the formula (005): [ka] or a pharma- ceutically acceptable salt or solvate thereof.

[0101] According to one embodiment, the glycocluster has the formula (008): [ka] or a pharma- ceutically acceptable salt or solvate thereof.

[0102] According to one embodiment, the glycocluster has the formula (014): [ka] or a pharma- ceutically acceptable salt or solvate thereof.

[0103] In the above equations (011), (005), (008), and (014), each R 1 is as described in formulas (I) and (Ia).

[0104] In one embodiment, in the above formulas (011), (005), (008), and (014), each R 1 is represented by the formula (9AcSA) [ka] where the wavy line represents the R 1 Represents the attachment point of

[0105] In one embodiment, the glycocluster is selected from compounds of formula (011), (005), (008), and (014) above, wherein each R 1 is of formula (9AcSA), or a pharma- ceutically acceptable salt or solvate thereof, i.e., compounds 011, 005, 008, and 014, as depicted in Example 1 herein, or a pharma- ceutically acceptable salt or solvate thereof.

[0106] All references herein to the compounds of the invention (e.g., "glycoclusters" or "Formula (I)") include references to salts, preferably pharma- ceutically acceptable salts thereof, solvates, multicomponent complexes, and / or liquid crystals. All references herein to the compounds of the invention include references to polymorphs and / or crystal habits thereof. All references to the compounds of the invention include references to pharma- cetically acceptable prodrugs thereof. All references to the compounds of the invention include references to isotopically labeled compounds, including deuterated compounds.

[0107] The compounds of the present invention (e.g., "glycocluster" or "Formula (I)") and subformulas thereof contain at least one asymmetric center(s) and therefore can exist in different stereoisomeric forms. Thus, all references to the compounds of the present invention include references to all possible stereoisomers, including not only racemates but also individual enantiomers and non-racemic mixtures thereof. When a compound is desired as a single enantiomer, such a single enantiomer may be obtained by stereospecific synthesis, resolution of the final product or any convenient intermediate, or chiral chromatographic methods, each of which are known in the art. Resolution of the final product, intermediate, or starting material may be carried out by any suitable method known in the art.

[0108] The compounds of the present invention may be in the form of pharma- ceutically acceptable salts. Pharmaceutically acceptable salts include the acid addition and base salts thereof. Suitable acid addition salts are formed from acids which form non-toxic salts. Examples include acetate, adipate, aspartate, benzoate, besylate, bicarbonate / carbonate, bisulfate / sulfate, borate, camsylate, citrate, cyclamate, edisylate, esylate, formate, fumarate, gluceptate, gluconate, glucuronate, hexafluorophosphate, hybenzate, hydrochloride / chloride, hydrobromide / bromide, hydroiodide / iodide, isethionate, lactate, malate, maleate, malonate, mesylate, methylsulfate, naphthylate, 2-napsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate / hydrogenphosphate / dihydrogenphosphate, pyroglutamate, saccharate, stearate, succinate, tannate, tartrate, tosylate, trifluoroacetate, and xinafoate. Suitable base salts are formed from bases that form non-toxic salts. Examples include aluminum, arginine, benzathine, calcium, choline, diethylamine, 2-(diethylamino)ethanol, diolamine, ethanolamine, glycine, 4-(2-hydroxyethyl)-morpholine, lysine, magnesium, meglumine, morpholine, olamine, potassium, sodium, tromethamine, and zinc salts. Hemisalts of acids and bases can also be formed, such as hemisulphate and hemicalcium salts. When a compound contains an acidic group and a basic group, the compound can also form an intramolecular salt, and such compounds are within the scope of the present invention. When a compound contains a hydrogen-donating heteroatom (e.g., NH), the present invention also covers salts and / or isomers formed by the transfer of said hydrogen atom to a basic group or atom within the molecule.

[0109] Pharmaceutically acceptable salts of the compounds of the present invention can be prepared by one or more of these methods: (i) reacting the compound with a desired acid, (ii) reacting the compound with a desired base, (iii) removing an acid- or base-labile protecting group from a suitable precursor of the compound, or ring-opening a suitable cyclic precursor, such as a lactone or lactam, using a desired acid, and / or (iv) converting one salt of the compound to another by reaction with a suitable acid or a suitable ion exchange column. All these reactions are typically carried out in solution. The salt precipitates from the solution and can be collected by filtration or can be recovered by evaporation of the solvent. The degree of ionization of the salt can vary from completely ionized to nearly non-ionized.

[0110] Pharmaceutical Compositions The present invention also relates to pharmaceutical compositions comprising a compound of the present invention, as described herein, and at least one pharma- ceutically acceptable carrier.

[0111] According to one embodiment, the pharmaceutical composition further comprises at least one more therapeutic agent. In one embodiment, the therapeutic agent is an antiviral agent. In one embodiment, the at least one more therapeutic agent is an angiotensin-converting enzyme 2 (ACE2) binding inhibitor as described herein. In one particular embodiment, the enzyme 2 (ACE2) binding inhibitor is vectorized by glycocluster.

[0112] The compounds of the present invention may be formulated alone or together in suitable dosage unit formulations containing conventional non-toxic pharma- ceutically acceptable carriers, adjuvants, and vehicles appropriate for each route of administration.

[0113] Medical Use and Methods of Treatment The present invention also relates to a compound of the invention as described herein, or a pharmaceutical composition of the invention as described herein, for use as a medicament.

[0114] The present invention also relates to a compound of the invention as described herein, or a pharmaceutical composition of the invention as described herein, for use in the treatment and / or prevention of an infectious disease.

[0115] According to one embodiment, the infectious disease is a coronavirus infection or a picornavirus infection. In one embodiment, the infectious disease is a coronavirus infection.

[0116] In one embodiment, the coronavirus is an alphacoronavirus or a betacoronavirus, preferably a betacoronavirus. Non-limiting examples of alphacoronavirus include human coronavirus 229E (HCoV-229E) and human coronavirus NL63 (HcoV-NL63), also known as HcoV-NH or New Haven coronavirus. Non-limiting examples of betacoronavirus include human coronavirus OC43 (HcoV-OC43), human coronavirus HKU1 (HcoV-HKU1), Middle East Respiratory Syndrome-related coronavirus (MERS-CoV), formerly known as novel coronavirus 2012 or HcoV-EMC, severe acute respiratory syndrome coronavirus (SARS-CoV), also known as SARS-CoV-1 or SARS-classic, and severe acute respiratory syndrome coronavirus (SARS-CoV-2), also known as 2019-nCoV or novel coronavirus 2019. In one embodiment, the coronavirus is selected from HcoV-229E, HcoV-NL63, HcoV-OC43, HcoV-HKU1, MERS-CoV, SARS-CoV-1, and SARS-CoV-2. In one embodiment, the coronavirus is selected from MERS-CoV, SARS-CoV-1, and SARS-CoV-2.

[0117] According to one embodiment, the coronavirus is a SARS coronavirus. In one embodiment, the coronavirus is SARS-CoV-1 or SARS-CoV-2. In one embodiment, the coronavirus is SARS-CoV (also called SARS CoV-1), which causes Severe Acute Respiratory Syndrome (SARS). In one embodiment, the coronavirus is SARS-CoV-2, which causes COVID-19.

[0118] In this application, reference to "coronavirus" or "SARS-CoV-2" encompasses currently identified variants thereof. According to one embodiment, the coronavirus is the original haplotype (lineage A or B) of the SARS-CoV-2 pandemic or a variant thereof. In one embodiment, the coronavirus is SARS-CoV-2 alpha (lineage B.1.1.7, and sublineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 beta (lineage B.1.351, and sublineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 gamma (lineage P.1, and sublineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 delta (lineages B.1.617.2, XD, XF, XS, and sublineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 epsilon (lineages B.1.427, B1.429, and sublineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Zeta (lineage P.2, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Eta (lineage B.1.525, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Theta (lineage P.3, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Iota (lineage B.1.526, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Kappa (lineage B.1.617.1, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Lambda (lineage C.37, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Mu (lineage B.1.621, and sub-lineages thereof). In one embodiment, the coronavirus is SARS-CoV-2 Omicron (strains B.1.1.529, BA.1, BA.2, BA.3, BA.4, BA.5, XE, and substrains thereof). A list of all SARS-CoV-2 variants can be found on the cov-lineages website: https: / / cov lineages.org / lineage_list.html.

[0119] In one embodiment, the COVID-19 is moderate COVID-19. In one embodiment, the COVID-19 is mild to moderate COVID-19. In one embodiment, the COVID-19 is mild COVID-19. In one embodiment, the COVID-19 is mild to severe COVID-19. In one embodiment, the COVID-19 is severe COVID-19. A subject with COVID-19 may or may not be hospitalized.

[0120] In one embodiment, COVID-19 severity is assessed according to the World Health Organization (WHO) severity criteria, as follows: -Mild: Mild clinical symptoms with no signs of pneumonia on imaging. - Moderate: Fever and radiological evidence of pneumonia with respiratory symptoms requiring oxygen (O2): 3L / min < 5L / min. - Severe: meets any of the following criteria: tachypnea (respiratory rate (RR) ≥ 30 breaths / min); oxygen saturation (SpO2) ≤ 93% at rest in ambient air; or SpO2 ≤ 97% with O2 > 5 L / min; arterial blood oxygen pressure / inspired oxygen fraction ratio (PaO2 / FiO2) ≤ 300 mmHg (lmmHg = 0.133 kPa), where PaO2 / FiO2 in high altitude areas (altitudes above 1,000 meters above sea level) should be corrected by the following formula: PaO2 / FiO2 [multiply by] [atmospheric pressure (mmHg) / 760]; and / or chest imaging showing evident disease progression of > 50% within 24-48 hours. - Severe: if any of the following criteria are met: respiratory failure and requiring mechanical ventilation; shock; and / or other organ failure requiring ICU care. In one embodiment, the severity and / or progression of COVID-19 is assessed by the WHO 10-point progression scale as shown in Table 2 below. [Table 2]

[0121] In one embodiment, the subject treated according to the present invention has COVID-19 and has a score on the WHO 10-point progression scale for COVID-19 (as set forth in Table 2) in the range of 1 to 5, preferably in the range of 2 to 4, in the range of 3 to 6, preferably in the range of 4 to 5, or in the range of 5 to 9, preferably in the range of 6 to 8.

[0122] The compounds or pharmaceutical compositions of the present invention may be administered orally, parenterally (e.g., intramuscularly, intraperitoneally, intravenously, ICV, intracisternal injection or infusion, subcutaneous injection, or implant), by inhalation spray, nasal, vaginal, rectal, sublingual, or topical routes of administration. For the treatment and / or prevention of infectious diseases, suitable dosage levels may be about 0.01 to 500 mg / kg of patient body weight per day (mg / kg / day), and may be administered in single or multiple doses. Typically, dosage levels will be about 0.1 to about 250 mg / kg / day, preferably about 0.5 to about 100 mg / kg / day, and more preferably about 2.5 to about 20 mg / kg / day. The compounds may be administered on a regimen of 1 to 4 times per day, preferably once or twice per day. It will be understood, however, that the specific dose level and frequency of administration for any particular patient may vary and will depend upon a variety of factors, including the activity of the particular compound employed, the metabolic stability and length of action of that compound, age, body weight, general health, sex, diet, mode and time of administration, excretion rate, drug combination, the severity of the particular disease, and the host being treated.

[0123] The present invention also relates to the use of a compound of the present invention as described herein or a pharmaceutical composition of the present invention as described herein in the manufacture of a medicament for the treatment and / or prevention of an infectious disease.The present invention also relates to a method for the treatment and / or prevention of an infectious disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of the present invention as described herein or a pharmaceutical composition of the present invention as described herein.The present invention also relates to the use of a compound of the present invention as described herein or a pharmaceutical composition of the present invention as described herein in the treatment and / or prevention of an infectious disease.

[0124] Glycocluster synthesis In the present invention, the glycoclusters can be prepared by any synthetic method known in the art. In particular, suitable synthetic methods for creating a covalent bond between an acetylated sialic acid and a macrocycle are part of the general knowledge of a person skilled in the art of organic chemistry.

[0125] According to one embodiment, the acetylated sialic acid may be attached to the macrocycle by at least one non-acetylated hydroxyl group at positions 2, 4, 7, 8, or 9. In one embodiment, the acetylated sialic acid is attached to the macrocycle by a hydroxyl at position 2 or 4. In one embodiment, the acetylated sialic acid is attached to the macrocycle by a hydroxyl at position 2. The hydroxyl at position 2 is particularly advantageous for attaching the acetylated sialic acid to the macrocycle without compromising the biological activity associated with the opposite side of the acetylated sialic acid.

[0126] According to one embodiment, the non-acetylated hydroxyl group is used as a coupling functional group and reacts with a corresponding coupling functional group (e.g., halocarbon, carboxylic acid, or amine) of the macrocycle to form a bond between the acetylated sialic acid and the macrocycle (e.g., an ether, ester, or amide bond). According to another embodiment, the hydroxyl group is first replaced by a group containing a coupling functional group other than hydroxyl and / or converted to a coupling functional group other than hydroxyl, and then the other coupling functional group reacts with the corresponding coupling functional group of the macrocycle to form a bond between the acetylated sialic acid and the macrocycle.

[0127] According to one embodiment, the glycoclusters are prepared by copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC). In one embodiment, the glycoclusters are prepared in the presence of a copper(I) catalyst, such as a copper(I) salt (e.g., copper bromide or copper iodide), or a mixture of a copper(II) salt (e.g., copper sulfate) and a reducing agent (e.g., sodium ascorbate).

[0128] According to one embodiment, the glycocluster is prepared by reaction of at least two equivalents of an acetylated sialic acid containing an azide (-N3) with a macrocycle containing at least two terminal alkynes (-C≡CH). In one embodiment, the acetylated sialic acid is -O-(CH2) n -N3 groups, where n is an integer ranging from 1 to 8. In one embodiment, the macrocycle comprises at least two -O-(CH2) m It contains a -C≡CH group, where m is an integer ranging from 1 to 8.

[0129] According to another embodiment, the glycocluster is prepared by reaction of at least two equivalents of an acetylated sialic acid containing a terminal alkyne (-C≡CH) with a macrocycle containing at least two azides (-N3). In one embodiment, the acetylated sialic acid is -O-(CH2) nIn one embodiment, the macrocycle comprises at least two -C≡CH groups, where n is an integer ranging from 1 to 8. In one embodiment, the macrocycle comprises at least two -O-(CH2) m It contains a -N3 group, where m is an integer ranging from 1 to 8. [Brief description of the drawings]

[0130] [Figure 1] FIG. 1 is a scheme showing a method for the preparation of SA- or 9-AcSA-derived glycoclusters 004-014 from α-propargyl sialic acid (α-p-SA) 001 or α-propargyl acetylated sialic acid (α-p-9-AcSA) 002, and azide compounds 003, 006, 009 and 012 (shown in Scheme 3 of Example 1-d). [Diagram 2] Box plots showing the specific binding probability (BP) measured between S1-functionalized chips and surfaces coated with 9-AcSA, SA, or streptavidin before and after blocking with free 1 mM 9-AcSA as described in Example 2.1. One data point represents the binding frequency (BF) obtained with 1024 FD curves. Squares within the box indicate the mean, colored boxes indicate the 25th and 75th percentiles, and whiskers indicate the highest and lowest values. Lines within the box indicate the median. N=9 maps were examined for three independent experiments. P values ​​were determined by two-sample t-test in Origin. [Diagram 3] 2 is a graph showing the binding frequency (BF) plotted between S1 and 9-AcSA as a function of retention time, as described in Example 2.2. Least-squares fitting of the data to a single exponential decay curve (straight line) provides the average kinetic on-rate (kon) of the investigated interaction. Further calculation (koff / kon) results in the KD. One data point represents the BF obtained with 1024 FD curves. [Figure 4-9] 2 shows the results of screening the anti-binding properties of SA or 9-AcSA derived glycoclusters, as described in Example 2.5. [Figure 4-8]Histograms showing the inhibition efficiency of the tested molecules, which is assessed by measuring the binding probability (BP) of the interaction between 9-AcSA and SARS-CoV-2 before and after incubation with increasing concentrations (1-100 μM) of the tested molecules. The normalized histograms show the relative BP of the interaction between 9-AcSA and SARS-CoV-2 before and after incubation with 1, 10, or 100 μM of free acetylated sialic acid (9-AcSA) (Figure 4), SA- and 9-AcSA-pillar[5]arenes 004 and 005 (Figure 5), SA- and 9-AcSA-fullerenes 013 and 014 (Figure 6), SA- and 9-AcSA-porphyrins 010 and 011 (Figure 7), or SA- and 9-AcSA-calix[4]arenes 007 and 008 (Figure 8). [Figure 9] FIG. 2 is a graph showing the decrease in binding probability (BP) after incubation with increasing concentrations (1-100 μM) of the acetylated test molecules 9-AcSA-α-p, 005, 008, 011 and 014 as described in Example 2.5. Data are representative of at least N=3 independent experiments (chips and samples) per SA dendrimer concentration. P values ​​were determined by a two-sample t-test at origin. Error bars indicate standard deviation (sd) of the mean. [Figure 10-11] As described in Example 2.6, we present the results of an investigation of the efficiency of 9-AcSA-porphyrin glycocluster 011 in inhibiting the binding of SARS-CoV-2 to acetylated SA on a model surface (9-O-acetylated SA) and live cells (CHO cells) at low concentrations. [Figure 10] Box plot showing relative binding values ​​of the interaction between SARS-CoV-2 and 9-O-acetylated SA model surfaces before and after incubation with increasing concentrations (0.001-100 μM) of 9-AcSA-porphyrin glycocluster 011. [Figure 11]Box plot showing relative binding values ​​of the interaction between SARS-CoV-2 and CHO cells before and after incubation with increasing concentrations (0.1-10 μM) of 9-AcSA-porphyrin glycocluster 011. [Figure 12] FIG. 2 is a histogram showing the results of the infectivity assay, i.e., infectivity measured in the presence of free SA, free 9-AcSA, SA-porphyrin 010, and 9-AcSA-porphyrin 011, as described in Example 2.7. Each dot represents the infectivity from a well. The colored boxes represent the mean and the whiskers represent the sd of the mean. The line within the box represents the median. P values ​​were determined by two-sample t-test in Origin. [Figure 13A-B] Two box plots of binding probability (BP, %) between S1 and Lec2 or CHO cells (Figure 13A), or between SARS-CoV-2 and Lec2 or CHO cells (Figure 13B). n=10 (Figure 13A) or 12 (Figure 13B) maps were examined for three independent experiments. P values ​​were determined by two-sample t-test in Origin. EXAMPLES

[0131] The present invention is further illustrated by the following examples.

[0132] Example 1: Materials, Methods, and Synthesis and Characterization Results Example 1-a: General Materials and Methods Solvents used for chromatography were purchased in technical grade and further distilled before their use. Reagents and chemicals were purchased in ACS grade from Sigma Aldrich or Acros and used without purification.

[0133] All reactions were run on Merck aluminum roll silica gel 60-F using KMnO4 and phosphomolybdic acid solutions as developing solvents. 254The compounds were monitored by thin layer chromatography (TLC) performed at 37°C. Merck silica gel (60, particle size 40-63 μm) was employed for flash column chromatography. NMR spectra were recorded on a JEOL ECX 400 or 500 using the solvent peak as the reference. 1 H and 13 C NMR, and 1 H- 1 H and 1 H- 13 The nucleophiles were characterized by C correlation experiments. The abbreviations used to define the multiplicities are as follows: s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet, and br = broad. Chemical shifts (δ) are reported in ppm and indirectly referenced to residual solvent signals. High-resolution mass spectra (HRMS) were performed on a Bruker maXis Impact QTOF mass spectrometer and a Bruker MicroTOF-Q II XL spectrometer.

[0134] Example 1-b: Synthesis and characterization of biotinylated sialic acid Compounds B4 and B5 were prepared as shown in Scheme 1. [ka]

[0135] Intermediate compound B1 was prepared according to a known procedure (Jung, D. et al., Chemical Communications, 2014, Vol. 50, pp. 3044-3047). Intermediate compound B2 was prepared according to a known procedure (Gan, Z. & Roy, R., Canadian Journal of Chemistry, 2002, Vol. 80, pp. 908-916). Compound B5 was acetylated according to a method previously described (Ogura, H. et al., Carbohydrate Research, 1987, Vol. 167, pp. 77-86).

[0136] Intermediate compound (B3): To a solution of compound B1 (401 mg, 1.0 mmol, 1 equiv.), compound B2 (636 mg, 1.2 mmol, 1.2 equiv.), and Et3N (0.278 mL, 2.0 mmol, 2 equiv.) in dry dimethylformamide (DMF) (20 mL) was added CuI (38 mg, 0.2 mmol, 0.2 equiv.) under argon atmosphere. The solution was stirred for 12 h at room temperature. After the solvent was removed under vacuum, EtOAc (30 mL) and saturated ammonium chloride (30 mL) were added and the phases were separated. The organic layer was washed with brine (30 mL), dried over MgSO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography on silica gel using dichloromethane (DCM) / MeOH (20:1) as eluent to give a white solid (975 mg, 1.15 mmol, 96% yield). 1H NMR (500 MHz, CDCl3) 7.79 (s, 1H, H-12), 6.68 (br, 1H, NH), 6.43 (br, 1H, NH), 5.47 (d, J = 46.4 Hz, 1H, 10a), 5.45-5.42 (m, H-8), 5.35-5.32 (m, 1H, H-7), 4.92-4.87 (m, 2H, H-4, H-10b), 4.60-4.53 (m, 4H, H-13, H-17), 4.38-4.33 (m, 2H, H-9a, H-25), 4.16-4.05 (m, 3H, H-5, H-6, H-9b), 3.91 (t, J = 5.1 Hz, 2H, H-14), 3.80 (s, 3H, COCH3), 3.61 (dd, J = 6.0, 2.7 Hz, 2H, H-15), 3.56 (dd, J = 5.9, 2.7 Hz, 2H, H-15), 3.50 (t, J = 5.2 Hz, 2H, H-17), 3.41-3.40 (m, 2H, H-18), 3.17 (dd, J = 11.8, 7.3 Hz, 1H, H-24), 2.95-2.88 (m, 1H, H-28a), 2.77 (d, J = 12.9 Hz, 1H, H-3a), 2.62 (dd, J = 12.8, 4.6 Hz, 1H, H-28b), 2.24 (t, J = 7.5 Hz, 2H, H-20), 2.16 (s, 3H, OCOCH3), 2.15 (s, 3H, OCOCH3), 2.03 (s, 3H, OCOCH3), 2.02 (s, 3H, OCOCH3), 1.97 (t, J = 12.5 Hz, 1H, H-3b), 1.88 (s, 3H, NCOCH3), 1.77-1.63 (m, 4H, H-21, H-23), 1.48-1.42 (m, 2H, H-22). 1313C NMR (126 MHz, CDCl3) δ 173.5 (C-19), 171.0, 170.80, 170.5, 170.3, 170.2 (C=O, NCOCH3, OAc), 168.2 (C-1), 164.0 (Cq, C-26), 143.8 (C-11), 124.4 (C-12), 98.6 (C-2), 72.8 (C-6), 70.4 (C-16), 70.1 (C-17), 69.9 (C-15), 69.4 (C-14), 69.2 (C-4), 68.5 (C-8), 67.5 (C-7), 62.6 (C-9), 61.9 (C-25), 60.3 (C-27), 58.4 (C-10), 55.7 (C-24), 53.0 (OCH3), 50.2 (C-13), 49.2 (C-5), 40.5 (C-28), 39.1 (C-18), 37.9 (C-3), 35.9 (C-20), 28.3 (C-22), 28.1 (C-23), 25.6 (C-21), 23.2 (NCOCH3), 21.2, 21.0, 2 x 20.9 (OCOCH3). HRMS (TOF-MS-ESI + , m / z): C 39 H 60 N7O 17 S [M+H] + Calculated for 930.3761; found 930.3750.

[0137] Compound (B4) ("biot-SA"): A solution of compound B3 (345 mg, 0.371 mmol, 1 equiv.) and NaOMe (40 mg, 0.742 mmol, 2 equiv.) in dry MeOH (20 mL) was stirred at 0° C. for 30 min, then warmed to room temperature and stirred for an additional 1.5 h. Amberlyst® 15 ion exchange resin was then added to neutralize the base. The resin was filtered and washed with water (2×5 mL). The filtrate was evaporated under reduced pressure to give a white solid. Without further purification, the white solid was dissolved in water (15 mL) and LiOH·H2O (35 mg, 0.831 mmol, 3 equiv.) was added. The solution was stirred at room temperature for 1 h, after which Amberlyst® 15 ion exchange was added. The reaction mixture was then filtered, the resin washed with water (2×5 mL), and the filtrate was lyophilized to give a white solid (186 mg, 0.249 mmol, 90% yield). 1 H NMR (500 MHz, D2O) δ 8.06 (s, 1H, H-12), 4.92-4.89 (m, 1H, H-10a), 4.68-4.66 (m, 1H, H-10b), 4.62-4.60 (m, 2H, H-13), 4.57-4.53 (m, 1H, H-27), 4.38-4.34 (m, 1H, H-25), 3.95-3.93 (m, 2H, H-14), 3.88-3.79 (m, 4H, H-5, H-8, H-6, H-9a), 3.76-3.71 (m, 1H, H-4), 3.65-3.55 (m, 6H, H-15, H-16, H-9b, H-7), 3.52-3.49 (m, 2H, H-17), 3.32-3.30 (m, 2H, H-18), 3.27-3.24 (m, 1H, H-24), 2.95-2.90 (m, 1H, H-28a), 2.74-2.68 (m, 2H, H-3a, H-28b), 2.21 (dd, J = 13.2, 6.5 Hz, 2H, H-20), 2.01-2.00 (m, 3H, NAc), 1.74 (t, J = 12.2 Hz, 1H, H-3b), 1.61-1.48 (m, 4H, H-21, H-23), 1.36-1.33 (m, 2H, H-22). 13C NMR (126 MHz, D2O) δ 176.8 (C=O, NCOCH3), 175.0 (C-1), 171.8 (C=O, C-19), 165.3 (C=O, C-26), 143.6 (C-11), 125.7 (C-12), 99.5 (C-2), 72.9 (C-6), 71.1 (C-8), 69.7 (C-16), 69.3 (C-17), 68.9 (C-15), 68.7 (C-14), 68.3 (C-7), 67.7 (C-4), 62.9 (C-9), 62.1 (C-25), 60.3 (C-27), 57.2 (C-10), 55.4 (C-24), 51.8 (C-5), 50.1 (C-13), 2 x 39.7 (C-28, C-3), 38.9 (C-18), 35.5 (C-20), 27.9 (C-22), 27.7 (C-23), 25.2 (C-21), 22.1 (NCOCH3). HRMS (TOF-MS-ESI + , m / z): C 30 H 50 N7O 13 S [M+H] + Calculated value: 748.3182; measured value: 748.3173.

[0138] Compound (B5) ("biot-9-AcSA"): To a solution of compound B4 (200 mg, 0.267 mmol, 1 equiv.) and trimethyl orthoacetate (0.34 mL, 2.67 mmol, 10 equiv.) in dry dimethyl sulfoxide (DMSO) (1.2 mL) was added p-toluenesulfonic acid monohydrate (5.0 mg, 0.027 mmol, 0.1 equiv.). The solution was stirred at room temperature for 12 h. DCM (50 mL) was then added to precipitate the crude product. The crude product was purified by elution with C2H2O / MeOH (0-1 / 3, gradient) as eluent. 18 Purification was performed by silica gel flash chromatography. The fractions containing compound B5 were collected and concentrated under reduced pressure. The concentrated solution was lyophilized to give the desired compound as a white solid (25 mg, 0.0316 mmol, 12%). 1H NMR (500 MHz, CD3OD) δ 8.02 (s, 1H, H-12), 4.94 (d, J = 12.1 Hz, 1H, H-10a), 4.67 (d, J = 12.1 Hz, 1H, H-10b), 4.55-4.55 (m, 2H, H-13), 4.49 (dd, J = 7.8, 4.7 Hz, 1H, H-27), 4.38 (dd, J = 11.2, 1.8 Hz, 1H, H-25), 4.31 (dd, J = 7.9, 4.5 Hz, 1H, H-9a), 4.14-4.06 (m, 2H, H-8, H-9b), 3.90 (t, J = 5.1 Hz, 2H, H-14), 3.74-3.71 (m, 2H, H-5, H-4), 3.64-3.60 (m, 3H, H-6, H-16), 3.58-3.56 (m, 2H, H-17), 3.51-3.48 (m, 3H, H-7, H-15), 3.35-3.33 (m, 2H, H-18), 3.20-3.17 (m, 1H, H-24), 2.94-2.84 (m, 2H, H-3a, H-28a), 2.69 (d, J = 12.7 Hz, 1H, H-28b), 2.21 (t, J = 7.4 Hz, 2H, H-20), 2.05 (s, OCOCH3), 2.02 (s, NCOCH3), 1.74-1.71 (m, 1H, H-3b), 1.66-1.5 (m, 4H, H-21, H-23), 1.45-1.41 (m, 2H, H-22). 13C NMR (126 MHz, CD3OD) δ 176.2 (C=O, NCOCH3), 175.5 (C-1), 174.0 (C=O, 9-O-Ac), 173.1 (C=O, C-19), 166.1 (C=O, C-26), 146.5 (C-11), 125.9 (C-12), 101.9 (C-2), 74.2 (C-6), 71.4 (C-16), 71.2 (C-17), 70.7 (C-8), 2 x 70.6 (C-7, C-15), 70.3 (C-14), 69.6 (C-4), 67.3 (C-9), 63.3 (C-25), 61.6 (C-27), 58.9 (C-10), 57.0 (C-24), 54.1 (C-5), 51.3 (C-13), 42.6 (C-3), 41.1 (C-28), 40.3 (C-18), 36.7 (C-20), 29.7 (C-22), 29.5 (C-23), 26.8 (C-21), 22.6 (NCOCH3), 20.8 (OCOCH3). HRMS (TOF-MS-ESI + , m / z): C 32 H 52 N7O 14 S [M+H] + Calculated value: 790.3287; observed value: 790.3287.

[0139] Example 1-c: Synthesis and characterization of sialic acid derivatives Intermediate compound 001 was prepared according to known procedures (Daskhan, G. et al., ChemistryOpen, 2016, Vol. 5, pp. 477-484). Intermediate compound 002 was prepared according to known procedures (Ogura, H. et al., Carbohydrate Research, 1987, Vol. 167, pp. 77-86), as shown in Scheme 2. [ka]

[0140] Prop-2-ynyl 5-acetamido-9-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-onuropyranose (002): To a solution of compound 001 (380 mg, 1.1 mmol, 1 equiv.) and trimethyl orthoacetate (1.4 mL, 11 mmol, 10 equiv.) in dry DMSO (4 mL), p-toluenesulfonic acid monohydrate (10 mg, 0.05 mmol, 0.05 equiv.) was added. The solution was stirred at room temperature for 12 h. DCM (20 mL) was then added to precipitate the crude product. After slow cotton filtration, the solid on the filter was redissolved in methanol, transferred, and the solvent was evaporated. The residue was purified by silica gel chromatography using DCM / MeOH (20:3) to give the desired compound (2) as a white solid (167 mg, 0.429 mmol, 39% yield). [ka] 1 H NMR (500 MHz, CD3OD) δ: 4.42-4.35 (m, 2H, H-9a, H-10a), 4.17-4.03 (m, 3H, H-10b, H-9b, H-8), 3.74-3.65 (m, 2H, H-4, H-5), 3.57-3.54 (m, 1H, H-6), 3.49 (dd, J = 9.2, 1.9 Hz, 1H, H-7), 2.83 (dd, J = 12.3, 4.4 Hz, 1H, H-3a), 2.75 (t, J = 2.4 Hz, 1H, H-13), 2.06 (s, 3H, OCOCH3), 2.03 (s, 3H, NHCOCH3), 1.60 (t, J = 11.7 Hz, 1H, H-3b). 13C NMR (126 MHz, CD3OD) δ 175.6 (C=O, NAc), 173.5 (C-1), 173.1 (C=O, 9-O-Ac), 101.6 (C-2), 80.6 (C-11), 75.0 (C-12), 74.1 (C-6), 70.6 (C-8), 70.5 (C-7), 69.2 (C-4), 67.2 (C-9), 54.0 (C-5), 53.1 (C-10), 42.2 (C-3), 22.7 (NCOCH3), 20.8 (OCOCH3). HRMS (TOF-MS-ESI + , m / z): C 16 H 24 NO 10 [M+H] + Calculated value 390.1395; measured value 390.1395.

[0141] Example 1-d: Synthesis and characterization of glycoclusters [ka]

[0142] Intermediate compound 003 was prepared according to known procedures (Nierengarten, I. et al., Chemical Communications, 2012, Vol. 48, pp. 8072-8074). Intermediate compound 006 was prepared according to known procedures (Tikad, A. et al., Chemistry: A European Journal, 2016, Vol. 22, pp. 13147-13155). Intermediate compound 009 was prepared according to known procedures (Tikad, A. et al., Chemistry: A European Journal, 2016, Vol. 22, pp. 13147-13155; and Liu, Y. et al., Angewandte Chemie (International Edition English), 2016, Vol. 55, pp. 7952-7957). Intermediate compound 012 was prepared according to known procedures (Nierengarten, JF et al., Chemical Communications, 2010, Vol. 46, pp. 3860-3862).

[0143] Compounds 004, 007, 010, and 013 ("SA-glycoclusters") and 005, 008, 011, and 014 ("9-As-SA-glycoclusters") were prepared by grafting clickable α-propargyl sialic acids 001 and 002 onto polymeric azides 003, 006, 009, and 0012 using either a combination of copper(II) sulfate and sodium L-ascorbate or copper(I) bromide dimethylsulfide, as shown in Figure 1.

[0144] More specifically, an excess of 001 or 002 was coupled to tetraazido pillar[5]arene 003 with catalytic amounts of copper(II) sulfate and sodium L-ascorbate in a mixed solvent system (1,4-dioxane / H2O, 2:1) at room temperature overnight. 004 and 005 were then precipitated with acetone and purified by copper scavenger (Quadrasil MP) and size-exclusion Sephadex® G-25 chromatography. Purified decavalent pillar[5]arenes 004 and 005 were obtained in 74% and 62% yields, respectively. A specific method was optimized for porphyrin tetramers 010 and 011 to avoid ion exchange between copper and zinc and to address the solubility properties of 009. Therefore, a lower catalyst amount and a ternary solvent system (THF / DMSO / H2O, 3:3:1) were employed. The tetravalent porphyrin conjugates 010 and 011 were obtained in 61% and 87% yields, respectively. Calix[4]arenes 007 and 008 and fullerenes 013 and 014 were also obtained in high yields using a similar coupling and purification protocol. All multimeric species were 1 H NMR, 13 All reactions were confirmed to be complete as characterized by C NMR and mass spectrometry.

[0145] Compound (004): To a solution of compound 003 (21 mg, 0.01 mmol, 1 eq.) and compound 001 (70 mg, 0.20 mmol, 12 eq.) in 1,4-dioxane (2 mL), a freshly prepared solution of CuSO4 (3.2 mg, 0.02 mmol, 2 eq.) and NaAsc (13 mg, 0.066 mmol, 6.6 eq.) in HO (1 mL) was added under argon atmosphere. The reaction mixture was stirred vigorously overnight at room temperature. Acetone (40 mL) was then added to precipitate the crude product. After centrifugation, the crude product was dissolved in water (2 mL) and treated with Quadrasil MP (40 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex® G-25 column and eluted with water. The fractions that did not migrate by TLC elution with DCM / MeOH (1:0.3, KMnO4 staining) were collected and the collected fractions were lyophilized to give a white solid (59 mg, 0.0124 mmol, 74% yield). [ka] 1 H NMR (500 MHz, DMSO-d6) δ 8.17-8.06 (m, 10H, 10 x H-12), 6.57 (s, 10H, 10 x H-16), 4.77-4.13 (m, 60H, 10 x H-10, 10 x H-13, 10 x H-14), 3.61-3.31 (m, 80H, 10 x H-4, 10 x H-5, 10 x H-6, 10 x H-7, 10 x H-8, 10 x H-9, 10 x H-18), 2.63 (s, 10H, 10 x H-3a), 1.87 (s, 30H, 10 x NCOCH3), 1.43 (s, 10H, 10 x H-3b). 13C NMR (126 MHz DMSO-d6) δ 173.9 (C=O, NCOCH3), 172.0 (C-1), 149.7 (Cq, C-15), 145.2 (Cq, C-11), 129.6 (Cq, C-17), 125.2 (C-12), 116.1 (C-16), 100.5 (C-2), 73.5 (C-6), 72.1 (C-8), 69.3 (C-7), 68.1 (C-4, C-14), 63.6 (C-9), 57.8 (C-10), 53.2 (C-5), 50.8 (C-13), 41.6 (C-3), 29.0 (C-18), 23.2 (NCOCH3). HRMS (TOF-MS-ESI - , m / z): C 195 H 267 N 40 O 100 [M-3H] 3- Calculated value 1590.2365; measured value 1590.2262.

[0146] Compound (005): To a solution of compound 003 (13 mg, 0.01 mmol, 1 equiv.) and compound 002 (47 mg, 0.12 mmol, 12 equiv.) in 1,4-dioxane (1.2 mL), a freshly prepared solution of CuSO4 (3.2 mg, 0.02 mmol, 2 equiv.) and NaAsc (13 mg, 0.066 mmol, 6.6 equiv.) in HO (0.6 mL) was added under argon atmosphere. The reaction mixture was stirred vigorously overnight at room temperature. Acetone (20 mL) was then added to precipitate the crude product. After centrifugation, the crude product was dissolved in water (2 mL) and treated with Quadrasil MP (40 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex® G-25 column and eluted with water. The unmigrated fractions were collected by TLC elution with DCM / MeOH (1:0.3) and KMnO4 staining. The collected fractions were lyophilized to give a white solid (32 mg, 0.0062 mmol, 62% yield). [ka] 1 1H NMR (500 MHz, CD3OD) δ 8.16 (br, 10H, 10 x H-12), 6.62 (br, 10H, 10 x H-16), 4.69 (br, 30H, 10 x H-10b, 10 x H-13), 4.37 (br, 20H, 10 x H-9), 4.09 (br, 30H, 10 x H-8, 10 x H-14), 3.76 (br, 40H, 10 x H-4, 10 x H-5, 10 x H-6, 10 x H-18), 3.53 (br, 10H, 10 x H-7), 2.83 (br, 10H, 10 x H-3a), 2.03 (br, 60H, 10 x OCOCH3, 10 x NCOCH3), 1.65 (br, 10H, 10 x H-13b). 13 13C NMR (126 MHz, CD3OD) δ 175.3 (C=O, NCOCH3), 173.9 (C-1), 173.3 (C=O, 9-O-Ac), 151.0 (Cq, C-15), 146.3 (Cq, C-11), 130.1 (Cq, C-17), 126.0 (C-12), 117.1 (C-16), 101.7 (C-2), 74.3 (C-6), 70.6 (C-8), 69.3 (C-7), 68.6 (C-4, C-14), 67.3 (C-9), 58.9 (C-10), 54.0 (C-5), 51.4 (C-13), 42.1 (C-3), 31.0 (C-18), 22.9 (NCOCH3), 21.0 (OCOCH3). HRMS (TOF-MS-ESI + , m / z): C 215 H 293 N 40 O 110 [M + 3H] 3+ Calculated value for 1732.2869; measured value 1732.2863.

[0147] Compound (007): To a solution of compound 006 (36 mg, 0.03 mmol, 1 equiv.) and compound 001 (46 mg, 0.132 mmol, 4.4 equiv.) in dry DMSO (1 mL), CuBr·CH3SCH3 (3.7 mg, 0.018 mmol, 0.6 equiv.) was added under argon atmosphere. After 4 h at 80 °C under microwave irradiation, the crude product was cooled and precipitated by adding DCM (30 mL). The precipitate was then dissolved in water (2 mL) and treated with Quadrasil® MP (70 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex® G-15 column and eluted with water. The fractions that could not be transferred by TLC elution with DCM / MeOH (1:0.3) and KMnO4 staining were collected. The collected fractions were lyophilized to give a white solid (70 mg, 0.0276 mmol, 92% yield). [ka] 1 H NMR (500 MHz, CD3OD) δ 8.03 (s, 4H, 4 x H-12), 6.82 (s, 8H, 8 x H-18), 4.62 (br, 8H, 8 x H-13), 4.35 (br, 4H, 4 x H-22a), 3.89-3.58 (m, 52H, 4 x H-4, 4 x H-5, 4 x H-6, 4 x H-7, 4 x H-8, 4 x H-9, 4 x H-15, 16 x OH), 3.13 (br, 4H, 4 x H-22b), 2.82 (br, 4H, 4 x H-3a), 2.52 (br, 8H, 4 x H-14), 2.02 (s, 12H, 4 x NCOCH3), 1.66 (br, 4H, 4 x H-3b), 1.08 (s, 36H, 12 x H-21). 13C NMR (126 MHz, CD3OD) δ 175.3 (C=O, NCOCH3), 173.9 (C-1), 154.2 (Cq, C-16), 146.1 (Cq, C-11, C-19), 134.9 (Cq, C-17), 126.4 (C-18), 125.6 (C-12), 101.3 (C-2), 74.50 (C-6), 73.0 (C-15), 70.1 (C-7, C-8), 69.2 (C-4), 64.3 (C-9), 58.4 (C-10), 53.9 (C-5), 48.5 (C-13), 41.9 (C-3), 34.7 (Cq, C-20), 32.0 (C-14, C-21, C-22), 22.9 (NCOCH3). HRMS (TOF-MS-ESI + , m / z): C after deconvolution 120 H 170 N 16 O 44 Calculated value for [M] 2369.0978; observed value 2369.0877.

[0148] Compound (008): To a solution of compound 006 (37 mg, 0.03 mmol, 1 equiv.) and compound 002 (51 mg, 0.132 mmol, 4.4 equiv.) in dry DMSO (1 mL), CuBr·CH3SCH3 (3.7 mg, 0.018 mmol, 0.6 equiv.) was added under argon atmosphere. After 4 h at 80 °C under microwave irradiation, the crude product was precipitated by adding DCM (30 mL). The precipitate was then dissolved in water (2 mL) and treated with Quadrasil® MP (70 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex® G-15 column and eluted with water. The fraction that could not be transferred by TLC elution with DCM / MeOH (1:0.3, KMnO4 staining) was collected. The collected fractions were lyophilized to give a white solid (62 mg, 0.0244 mmol, 81% yield). [ka] 11H NMR (500 MHz, CD3OD) δ 8.21 - 8.06 (m, 4H, 4 x H-12), 6.82 (s, 8H, 8 x H-18), 4.63 (br, 8H, 4 x H-13), 4.37 (s, 8H, 4 x H-9a, 4 x H-22a), 4.13 (br, 8H, 4 x H-8, 4 x H-9b), 3.88 - 3.52 (m, 24H, 4 x H-4, 4 x H-5, 4 x H-6, 4 x H-7, 4 x H-15), 3.13 (s, 4H, 4 x H-22b), 2.79 (br, 4H, 4 x H-3a), 2.53 (br, 8H, 4 x H-14), 2.02 (br, 24H, 4 x NCOCH3, 4 x OCOCH3), 1.70 (br, 4H, 4 x H-3b), 1.33 (s, 6H, 2 x H-21), 1.08 (s, 24H, 8 x H-21), 0.85 (s, 6H, 2 x H-21). 13 13C NMR (126 MHz, CD3OD) δ 175.3 (C=O, NCOCH3), 173.1 (C=O, C-1, 9-O-Ac), 154.2 (Cq, C-16), 146.4 (Cq, C-11), 146.0 (Cq, C-19), 134.9 (Cq, C-17), 126.4 (Cq, C-12, C-18), 100.8 (C-2), 74.4 (C-6), 72.9 (C-15), 70.5 (C-8), 70.4 (C-7), 69.1 (C-4), 67.4 (C-9), 58.7 (C-10), 53.8 (C-5), 42.0 (C-3), 34.7 (Cq, C-20), 32.0 (C-14, C-21, C-22), 22.75 (OCOCH3), 20.9 (OCOCH3). HRMS (TOF-MS-ESI + , m / z): Calculated value for C 120 H 170 N 16 O 44 [M] is 2537.1400; measured value is 2537.1298.

[0149] Compound (010): To a solution of compound 009 (40 mg, 0.037 mmol, 1 eq.) and compound 001 (57 mg, 0.164 mmol, 4.4 eq.) in THF / DMSO (1.5 mL, 2:3) was added a freshly prepared solution of CuSO4 (2.36 mg, 0.0148 mmol, 0.4 eq.) and NaAsc (8.7 mg, 0.0439 mmol, 1.2 eq.) in HO (0.3 mL) under argon atmosphere. The reaction mixture was stirred vigorously overnight at room temperature. Acetone (30 mL) was then added to precipitate the crude product. The crude product was then dissolved in water (2 mL) and treated with Quadrasil® MP (80 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex® G-15 column and eluted with water. The fractions that did not migrate by TLC elution with DCM / MeOH (1:0.3, KMnO4 stain) were collected and the collected fractions were lyophilized to give a dark green solid (56 mg, 0.0227 mmol, 61% yield). [ka] 1 H NMR (400 MHz, D2O / DMSO-d6) δ 9.07 (br, 8H, 8 x H- Porph ), 8.32 (br, 12H, 4 x H-12, 8 x H-18), 7.51 (br, 8H, 8 x H-17), 3.74 (br, 28H, 4 x H-4, 4 x H-5, 4 x H-6, 4 x H-7, 4 x H-8, 4 x H-9), 2.78 (br, 12H, 4 x H-14, 4 x H-3a), 2.05 (br, 16H, 4 x H-3b, 4 x NCOCH3). 1313C NMR (101 MHz, D2O / DMSO-d6) δ 176.3 (C=O, NCOCH3), 173.9 (C-1), 159.5 (C-16), 151.3 (Cq, C-19), 146.6 (Cq, C-11), 137.2 (C-18), 135.5 (Cq, C- Porph ), 133.3 (C- Porph ), 127.0 (C-12), 121.5 (Cq, C- Porph ), 114.5 (C-17), 101.9 (C-2), 74.3 (C-6), 73.1 (C-8), 69.7 (C-4, C-7), 67.0 (C-15), 64.1 (C-9), 59.1 (C-10), 53.4 (C-5), 49.4 (C-13), 41.8 (C-3), 30.9 (C-14), 23.6 (NCOCH3). HRMS (TOF-MS-ESI + , m / z): C 112 H 134 N 20 O 40 Zn [M+2H] 2+ Calculated for 1232.4186; found 1232.4390.

[0150] Compound (011): To a solution of compound 009 (40 mg, 0.037 mmol, 1 eq.) and compound 002 (63 mg, 0.162 mmol, 4.4 eq.) in THF / DMSO (1.8 mL, 1:1) was added a freshly prepared solution of CuSO4 (2.36 mg, 0.0148 mmol, 0.4 eq.) and NaAsc (8.7 mg, 0.0439 mmol, 1.2 eq.) in HO (0.3 mL) under argon atmosphere. The reaction mixture was stirred vigorously overnight at room temperature. Acetone (30 mL) was then added to precipitate the crude product. The crude product was then dissolved in water (2 mL) and treated with Quadrasil® MP (80 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex® G-15 column and eluted with water. The fractions that did not migrate by TLC elution with DCM / MeOH (1:0.3) were collected and lyophilized to give a dark green solid (82 mg, 0.0323 mmol, 87% yield). [ka] 1 H NMR (500 MHz, DMSO-d6) δ 8.60 (br, 8H, 8 x H- Porph ), 8.11 (br, 4 x H-12), 7.72 (br, 8H, 8 x H-18), 6.89 (br, 8H, 8 x H-17), 4.18-4.04 (m, 56H, 4 x H-4, 4 x H-5, 4 x H-8, 4 x H-9, 4 x H-10, 4 x H-13, 4 x H-15, 12 x OH, 4 x NH), 3.46-3.33 (m, 8H, 4 x H-6, 4 x H-7), 2.26 (8H, 4 x H-14), 1.88 (br, 28H, 4 x NCOCH3, 4 x OCOCH3, 4 x H-3b). 13C NMR (126 MHz, DMSO-d6) δ 174.0 (C=O, NCOCH3), 172.3 (C=O, C-1, 9-O-Ac), 158.3 (Cq, C-16), 150.3 (Cq, C-19), 135.7 (C-18), 132.2 (C- Porph ), 120.7 (Cq, C- Porph ), 113.2 (C-17), 73.1 (C-6), 69.6 (C-8), 68.0 (C-4, C-7), 67.0 (C-9), 65.2 (C-15), 58.1 (C-10), 53.1 (C-5), 47.8 (C-13), 41.8 (C-3), 30.1 (C-14), 23.1 (NCOCH3), 21.3 (OCOCH3). HRMS (TOF-MS-ESI + , m / z): C 120 H 140 N 20 O 44 Zn [M+2H] 2+ Calculated value 1313.4228; observed value 1313.3966.

[0151] Compound (013): To a solution of compound 012 (40 mg, 0.0172 mmol, 1 equiv.) and compound 001 (95 mg, 0.274 mmol, 16 equiv.) in DMSO (1.4 mL), CuBr·CH3SCH3 (3.5 mg, 0.017 mmol, 1 equiv.) was added under argon atmosphere. After 4 h at 80 °C under microwave irradiation, the solution was cooled, precipitated with DCM (30 mL), and centrifuged. The crude product was redissolved in water (2 mL) and treated with Quadrasil® MP (80 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex™ G-25 column and eluted with water. The fractions that could not be transferred by TLC elution with DCM / MeOH were collected. The collected fractions were lyophilized to give an orange solid (89 mg, 0.0137 mmol, 80% yield). [ka] 11H NMR (400 MHz, DMSO-d6) δ 8.07 - 7.97 (m, 12H, 12 x H-12), 4.67 (s, 12H, 12 x H-10a), 4.40 (s, 36H, 12 x H-10b, 12 x H-15), 4.15 (br, 84H, 12 x H-12, 48 x OH, 12 x NH), 3.69 - 3.29 (m, 84H, 12 x H-4, 12 x H-5, 12 x H-6, 12 x H-7, 12 x H-8, 12 x H-9), 2.67 (s, 12H, 12 x H-3a), 2.15 (s, 24H, 12 x H-14), 1.86 (s, 36H, 12 x NAc), 1.36 (m, 12H, 12 x H-3b). 13 13C NMR (101 MHz, DMSO-d6) δ 174.0 (C=O, NCOCH3), 171.9 (C=O, C-1), 164.0 (Cq, C-16), 145.6, 141.5 (C-11, C sp2 ), 125.2 (C-12), 100.7 (C-2), 73.4 (C-6), 72.1, 69.4, 68.2 (C-18, C-8, C-7, C-4), 65.1 (C-15), 63.6 (C-9), 57.8 (C-10), 53.3 (C-5), 47.2 (C-17, C-13), 41.9 (C-3), 29.4 (C-14), 23.2 (NCOCH3). HRMS (ESI+-MS, m / z): C 282 H 328 N 48 O 132 [M + 4H] 4+ Calculated value for [M + 4H] is 1623.2468; found 1623.2138.

[0152] Compound (014): To a solution of compound 012 (22 mg, 0.0096 mmol, 1 equiv.) and compound 002 (60 mg, 0.154 mmol, 16 equiv.) in DMSO (0.8 mL), CuBr·CH3SCH3 (2 mg, 0.0096 mmol, 1 equiv.) was added under argon atmosphere. After 4 h at 80 °C under microwave irradiation, the solution was cooled, precipitated with DCM (30 mL), and centrifuged. The crude product was redissolved in water (2 mL) and treated with Quadrasil® MP (60 mg) to remove residual copper ions. After filtration through a 45 μm sterile filter, the filtrate was passed through a Sephadex™ G-25 column and eluted with water. The fraction that could not be transferred by TLC elution with DCM / MeOH (1:0.3, KMnO4 staining) was collected. The collected fractions were lyophilized to give an orange solid (55 mg, 0.00785 mmol, 82% yield). [ka] 1 H NMR (500 MHz, DMSO-d6) δ 8.24-8.01 (m, 12H, 12 x H-12), 4.94-3.25 (m, 192H, 12 x H-4, 12 x H-5, 12 x H-6, 12 x H-7, 12 x H-8, 12 x H-9, 12 x H-10, 12 x H-13, 12 x H-15), 2.68 (br, 12H, H-3a), 2.16-1.15 (m, 80H, 12 x H-3b, 12 x H-14, 12 x NCOCH3, 12 x OCOCH3). 13 C NMR (101 MHz, DMSO-d6) δ 173.5 (C=O, NCOCH3), 171.5 (C=O, C-1, 9-O-Ac), 163.57 (Cq, C-16), 145.7, 145.2 (C-11, C sp2), 124.5 (C-12), 100.4 (C-2), 72.9 (C-6), 69.4 (C-18, C-8, C-7), 67.8 (C-4), 66.5 (C-9), 64.5 (C-15), 57.64 (C-10), 53.1 (C-5), 46.7 (C-17, C-13), 41.6 (C-3), 29.2 (C-14), 22.9 (NCOCH3), 21.2 (OCOCH3). HRMS (ESI+-MS, m / z): C 306 H 348 N 48 O 144 [M+5H] 5+ Calculated value 1400.6350; observed value 1400.6368.

[0153] Example 2: AcSA-derived glycoclusters are potent inhibitors of SARS-CoV-2 cell binding and infectivity material and method Emergence of UV-inactivated SARS-CoV-2 SARS-CoV-2 (strain BavPat1, European Virology Archives) was propagated in Vero E6 and used at passage 3. 500 μL of passage 3 stock (supernatant of infected cells) was added to a 6-well dish. The 6-well dish was placed without the lid in a UV Stratalinker 1800 (Stratagene) and the virus-containing supernatant was exposed to 5000 J of UV irradiation. Viral inactivation was confirmed by adding 10 μL of UV-treated supernatant to a 48-well plate containing 50000 naive Vero E6 cells and monitoring infection after 48 h by indirect immunofluorescence.

[0154] Sialic acid coated surface The gold-coated silicon substrate was first washed with ethanol and cleaned by UV-O treatment (Jetlight) for 15 min. The surface was then stained with a biotinylated bovine serum albumin (BBSA) solution (25 μg mL in PBS). -1The plates were incubated in 100 µg / mL PBS (Sigma) overnight at 4 °C. After rinsing with PBS, the plates were rinsed with 1 drop of streptavidin (10 µg mL PBS). -1 , Sigma) was pipetted onto the BBSA surface for 1 h at 4 °C, followed by rinsing with PBS. Finally, the BBSA-streptavidin surface was washed with biotinylated biot-SA (B4) or biot-9-AcSA (B5) solution (10 μg mL in PBS). -1 ) for 1 h, followed by a final rinse with PBS. The surface exhibited a uniform and stable morphology under repeated scanning and a thickness of 1.1 ± 0.1 nm. The thickness of the deposited layer was estimated by scanning a small area of ​​the surface (1 μm × 1 μm) at high force to remove bound biomolecules, followed by imaging a wider square of the same area (5 μm × 5 μm) at a lower force.

[0155] Virus particle imaging Droplets of 80 μL of virus solution (~10 7 particles·mL -1 ) was deposited on a freshly cleaved mica substrate and incubated for 1 h at +4 °C. After rinsing 10 times with MilliQ water, the sample was dried for 1 h at 37 °C. AFM imaging was performed in air using a PeakForce-Hirs-FA tip (nominal spring constant 0.4 N m -1 The imaging parameters used were: tip oscillation frequency 1 kHz, maximum peak force 250 pN, scanning speed 0.25 kHz, and display 256 pixels / line.

[0156] AFM tip functionalization For AFM tip functionalization, NHS-PEG 24-Ph-aldehyde linker (Broadpharm) was used. AFM tips (MSCT-D probes, Bruker) were immersed in chloroform for 10 min, rinsed with ethanol, dried in a gentle stream of filtered nitrogen, cleaned with UV light and ozone cleaner (JetLight) for 15 min, and immersed overnight in ethanolamine solution [3.3 g ethanolamine hydrochloride in 6.6 mL dimethylsulfoxide (DMSO)]. The cantilevers were then washed three times with DMSO and three times with ethanol and dried with nitrogen. Meanwhile, 3.3 mg of NHS-PEG was added to the cantilevers. 24 The -Ph-aldehyde linker was dissolved in 0.5 mL of chloroform. Ethanolamine-coated cantilevers were immersed in this solution along with 30 μL of triethylamine. After an incubation time of 2 h, they were washed three times with chloroform, dried with nitrogen and placed in a star configuration (tips facing each other) on parafilm (Bemis NA). 50 μL of S1 subunit protein solution (0.1 mg mL -1 , Genscript Z03501) or UV-inactivated SARS-CoV-2 virions (10 8 particles·mL -1 ), and 2 μL of freshly prepared NaCNBH3 solution (6 wt% vol -1 , in 0.1 M NaOH(aq) was pipetted onto them and incubated for 1 h at 4 °C. Finally, 5 μL of 1 M ethanolamine (pH = 8) was added to the droplets for 10 min to quench the reaction. After washing with PBS, the chips were stored in PBS until the experiment.

[0157] Binding probability (BP) assay Force spectroscopy experiments were performed on the model surfaces using a Nanoscope Multimode 8 (Bruker) operated in force volume (contact) mode (Nanoscope software v9.1) with an MSCT-D probe (nominal spring constant 0.03 N m -1) was used to record a 5 μm × 5 μm map with a lamp size of 200 nm, a maximum force of 500 pN, and no surface retardation. The sample was scanned using a line frequency of 1 Hz and 32 pixels / line (32 lines). Both the approach and retraction speeds were set at 1 μm s -1 was kept constant.

[0158] Inhibitory screening To study the role of sialic acids during the first step of SARS-CoV-2 binding to the host cell surface, FD curve-based AFM was used to compare SARS-CoV-2 binding to SA(Neu5Ac) and 9-AcSA and to characterize the binding free energy landscape of the interaction to 9-AcSA. To mimic the exposure of cell surface glycans in vitro, biotinylated SA, either biot-9-AcSA (B5) or biot-SA (B4), was immobilized on a streptavidin-coated surface and verified by AFM imaging and scratching experiments, revealing a deposited layer with a thickness of 1.1 ± 0.1 nm. The interaction between the spike S1 subunit and the SA-coated surface was monitored by FD-based AFM.

[0159] To study the inhibitory potential of the synthesized (9-Ac)-SA derived glycoclusters (see Example 1-d) on the binding affinity between SARS-CoV-2 and 9-AcSA, the binding probability (BP) (percentage of the curve showing binding events) was measured before and after incubation with different concentrations (0, 1, 10 and 100 μM, respectively) of SA-glycoclusters (004, 007, 010 and / or 013) and four 9-AsSA-glycoclusters (005, 008, 011 and / or 014). Three force volume maps were recorded for three different regions, as previously described, in the absence of glycoclusters. The tested glycoclusters were then added to the fluid cell and three maps were recorded for each concentration.

[0160] dynamic force spectroscopy Dynamic force spectroscopy experiments were performed using a ForceRobot 300 (JPK). The same parameters were used as for the binding probability (BP) assay, with 0.1, 0.2, 1, 5, 10 and 20 μm s -1 Various retraction rates of 1000 Hz to 1000 Hz were used. Origin software (OriginLab) was used to display the results in DFS plots and generate rupture force histograms for distinct LR ranges, and various force spectroscopy models were applied as previously described (Alsteens, D. et al., Nature Nanotechnology, 2017, Vol. 12, pp. 177 183). These models were used to quantify the energy landscape of this interaction and to estimate the kinetic off-rate k off and the distance to the transition state x u In kinetic on-rate analysis, BP is determined at a specific contact time (t), where the contact time is the time that the tip is in contact with the surface.

[0161] The data were fitted and the K D was calculated (Rankl, C. et al., Proceedings of the National Academy of Sciences of the United States of America, 2008, Vol. 105, pp. 17778-17783). Briefly, the relationship between the interaction time (τ) and BP is given by the following equation:

number

number

[0162] Cell line culture CHO cells were cultured in 10% FBS (fetal bovine serum), penicillin (100 U mL -1 ), streptomycin (100 μg mL -1 ) (Invitrogen), and 2 mM L-glutamine (Sigma).

[0163] Transduction of Lec2 cells Lec2 cells were transduced to express nuclear eGFP and cytoplasmic mCherry using H2BeGFP and actin-mCherry expressing lentiviruses.

[0164] Labeling UV-inactivated SARS-CoV-2 virions with Atto488 NHS ester dye 10 of UV-inactivated SARS-CoV-2 virions 8 particles·mL ―1 200 μL of the solution was mixed with 10 μL of Atto488 NHS ester dye (Atto-Tec) (10 mM in dry DMSO) in 500 μL of acetone buffer (pH 4.5) for 2 h under gentle agitation. Free dye was then eliminated and the fluorescently labeled virions were concentrated to the initial concentration by filtration using Amicon Ultra-0.5 centrifugal filter units, MWCO 10 kDa (Sigma) (centrifugation at 10000 g for 10 min for purification and 1000 g for 2 min for product recovery).

[0165] Virus binding assay Co-cultures of CHO and Lec2 (fluorescently labeled with actin-mCherry) were incubated with 10 μl of UV-inactivated SARS-CoV-2 virions conjugated with Atto488 NHS ester dye.8 particles·mL ―1 The cells were incubated with the solution for 1 h on ice (to prevent internalization). The cells were then rinsed three times with PBS and fixed with formaldehyde (4% in PBS, 15 min) (Invitrogen, Thermo Fisher Scientific). After a final wash with PBS, the cells were imaged with a laser scanning confocal microscope (Zeiss LSM 980) using a 40x water immersion objective. Images were analyzed with Zen blue 2.3 software (Zeiss). Maximum intensity projections were performed to obtain single images from z-stacks.

[0166] FD-based AFM and fluorescence microscopy of live cells AFM correlative images of CHO cells were acquired using a Bioscope Resolve AFM (Bruker) connected to an inverted epifluorescence microscope (Zeiss Observer Z.1) or a confocal laser scanning microscope (Zeiss LSM 900) in PeakForce QNM mode (Nanoscope software v9.2). All experiments were performed using a 40x oil immersion objective (NA = 0.95). Cell images (30–50 μm 2) were recorded at a force of 500 pN using a PFQNM-LC probe (Bruker) with a tip length of 17 μm, a tip radius of 65 nm, and an opening angle of 15°. All fluorescence and AFM experiments were realized under cell culture conditions, using an integrated AFM and fluorescence microscope chamber, at 37 °C, in either Mem α (nucleoside-containing) medium or Ham's F12 medium depending on the cell type (Alsteens, D. et al., Nature Nanotechnology, 2017, Vol. 12, pp. 177-183). The cantilever was calibrated using the thermal noise method (Hutter, JL & Bechhoefer, J., Review of Scientific Instruments, 1993, Vol. 64, pp. 1868-1873), obtaining values ​​in the range of 0.08-0.14 N / m. The AFM tip was oscillated sinusoidally at 0.25 kHz, with an amplitude of 750 nm. Samples were scanned using a frequency of 0.125 Hz and 128 or 256 pixels / line. AFM images and FD curves were analyzed using Nanoscope analysis software (v1.9, Bruker), Origin, and ImageJ (v1.52e). Individual FD curves detecting dissociation events between the cell surface and S1 or SARS-CoV-2 were analyzed using Nanoscope analysis and Origin software. The baseline of the retraction curves was corrected using linear fitting to the last 30% of the retraction curve. The force-time curves were used to determine the loading rate (slope) of each rupture event. Optical images were analyzed using Zen Blue software (Zeiss).

[0167] Monitoring the effect of 9-AcSA porphyrin addition Live cell experiments were performed in the same manner as above by scanning a suitable area of ​​a confluent layer of cells followed by addition of 10 nM, 100 nM, 1 μM, or 10 μM of 9-AcSA porphyrin 011 to the medium. The same area was then scanned again to monitor potential changes following addition of 9-AcSA oligomers.

[0168] Production of SARS-CoV-2 spike-pseudotyped VSV viruses The pCG1 SARS-CoV-2 spike protein with C-terminal truncation of 18 amino acid residues plasmid was transfected into HEK-293 cells. The next day, VSV-deltaG virions were transduced into the cells at an MOI of 5 FFU / cell. After 1 h of incubation at 37 °C in a humidity-saturated environment containing 5% CO2, the medium was removed and the cells were washed with PBS. The transduced cells were cultured in DMEM supplemented with 5% FBS, 1% penicillin, 1% streptomycin, 2 mM L-glutamine, 1 mM Na-pyruvate, and NEAA, and anti-VSV-G antibody (1:1000). The produced cells were harvested from the medium the day after transduction. Cell debris was removed by centrifugation (1250 × g, 10 min) and 0.22 μm filter.

[0169] Infectivity assay A549 or A549 ACE2 stable cells (1 × 10 4 ) were seeded in 96-well plates. A mixture of pseudotyped virus at MOI 5 and increasing concentrations (0.001 μM, 0.01 μM, 0.1 μM, 1 μM, 10 μM) of SA(Neu5Ac), 9-AcSA, SA-porphyrin glycocluster 010, or 9-AcSA-porphyrin glycocluster 011 (as shown in Example 1-d) was incubated at room temperature for 15 min. The mixture was added to the medium and cells were incubated for 1 h. Cells were washed with PBS and incubated with fresh cell medium for 24 h. Infectivity was monitored by fluorescence and images were acquired with a bioimager device (Amersham Typhoon). The number of infected cells was counted by Fiji.

[0170] result, 1. Binding of the S1 subunit to 9-O-acetylated sialic acid (9-O-AcSA) Strikingly, a significant difference of approximately three-fold was observed between acetylated (B5) and non-acetylated SA (B4), demonstrating a significantly higher binding activity of the S1 subunit to 9-AcSA compared to SA.

[0171] Furthermore, the specificity of the interaction was confirmed by performing additional independent control experiments: both competition assays with free 9-AcSA (Figure 2) or surfaces coated with streptavidin only (no SA) resulted in significantly lower binding probabilities (BP), confirming the specificity of the interaction of 9-AcSA with the SARS-CoV-2 S1 domain.

[0172] 2. Kinetics of the interaction between the S1 subunit and 9-O-AcSA The binding free energy landscape of the interaction between S1 and 9-AcSA was characterized using single-molecule dynamic force spectroscopy (DFS). By monitoring the effect of contact time on BP (Figure 3), the binding rate (k on ) is estimated by assuming that the receptor-binding complex can be approximated by a pseudo-first-order kinetic equation. By fitting the data with monoexponential growth, it is found to be (4.2 ± 0.2) × 10 4 M -1 s -1 k on is extracted, and the affinity constant (K D ) is k off and k on This was calculated as the ratio of 0.01 to 0.1, resulting in a value of 5.7 ± 5 μM.

[0173] Values ​​in the μM range correspond to moderate affinity and support the hypothesis that SARS-CoV-2 uses glycans such as 9-AcSA on the host cell surface as an initial moderate affinity foothold to facilitate subsequent strong binding to the ACE2 receptor.

[0174] 3. SARS-CoV-2 virion-level binding Non-replicating SARS-CoV-2 particles (i.e., native SARS-CoV-2 virions inactivated by UV radiation) were used to assess the physiological relevance of the investigated interactions. Binding of these non-replicating SARS-CoV-2 particles to 9-AcSA was assessed by grafting whole virions onto an AFM tip. The interactions were monitored at medium speed (1 μm s-1 ) and high speed (20 μm s -1 ), the DFS plots were reconstructed and overlaid with data obtained with the purified S1 domain.

[0175] A good agreement was observed between data collected on the purified S1 domain alone or on full virions: both distinct rupture forces at the single molecule level were very close, as were the kinetic parameters. At the virion level, higher forces were also recorded, consistent with multiple contacts.

[0176] 4. Verification of interactions in living cells The interaction was then verified directly on live cells by investigating the interaction between either purified S1 or non-replicating fulvirions and co-cultures of unlabeled CHO cells, which naturally express sialic acid (SA), and Lec2 cells (fluorescently labeled with nuclear protein H2B-GFP and actin-mCherry). To prove the role of SA as a surface receptor, the inventors performed laser scanning confocal microscopy on co-cultures of CHO and Lec2 cells and observed that UV-inactivated SARS-CoV-2 virions (fluorescently labeled with Atto488-NHS dye) preferentially bound to SA-expressing CHO cells. Confluent monolayers of co-cultured CHO and Lec2 cells were then scanned by AFM using AFM tips functionalized with either S1 glycoprotein or fulvirions, using conditions in which both cell types grow.

[0177] While CHO cells showed a significantly higher density of attachment events (~9% for S1 and ~13% for SARS-CoV-2, Fig. 13a, n=10 cells for S1 and n=12 cells for SARS-CoV-2), Lec2 cells showed only a sparse distribution of these events (~3% for S1 and ~5% for SARS-CoV-2, Fig. 13b, n=10 cells for S1 and n=12 cells for SARS-CoV-2), confirming the establishment of specific SARS-CoV-2 binding to SA on live cells.

[0178] 5. Inhibition of SARS-CoV-2 binding using SA- and AcSA-glycoclusters Eight glycoclusters decorated with either SA or 9-AcSA were tested for their blocking properties using our SMFS approach and non-replicating SARS-CoV-2 full particles. First, the inhibitory potential of monovalent commercially available 9-acetylated sialic acid (9-AcSA) (Carbosynth) was evaluated at 0, 1, 10, and 100 μM (Figure 4). Only a modest decrease in binding frequency (BF) of 20-30% was observed over the range of concentrations investigated. This result was expected, as it was previously determined that active AcSA (compared to SA) exhibits only moderate affinity for the SARS-CoV-2 spike, as also evident in Example 2.2.

[0179] Four glycoclusters functionalized with either SA or 9-AcSA were compared (Figures 5-8). Relative binding frequency (BF) plots show no or only slight inhibition for SA-glycoclusters, while a gradual and effective BF drop was observed for 9-AcSA-glycoclusters. Furthermore, a 50% drop per 10 μM was observed for all 9-AcSA-clusters. The 9-AcSA-derived porphyrin 011 appears to be the most efficient inhibitor, reaching a drop of about 50% already at 1 μM (Figures 7 and 9).

[0180] Thus, the applicants have surprisingly demonstrated that multiple AcSAs covalently bound to porphyrins, pillararenes, calixarenes, and fullerenes result in potent inhibition of SARS-CoV-2 despite the low affinity of free AcSA. Thus, the glycoclusters of the present invention may be used for the treatment and / or prevention of infectious diseases such as COVID-19.

[0181] 6. Characterization of AcSA-induced porphyrin inhibition Since the first screen (Example 2.5 herein) pointed to a porphyrin-based glycocluster (compound 011) as the most efficient 9-AcSA-derived glycocluster for SARS-CoV-2 anti-binding, its binding ability was evaluated in more detail.

[0182] Surprisingly, it was found that already at 10 nM, 9-AcSA-porphyrin 011 caused a greater than 50% reduction in the investigated interaction between SARS-CoV-2 and 9-AcSA, following an exponential decay as a function of concentration (Figure 10).

[0183] The inhibitory potency on live cells was examined and this trend was confirmed even under physiologically relevant conditions (Figure 11).

[0184] Thus, the applicants have unexpectedly demonstrated that multiple AcSAs covalently bound to porphyrins provide particularly significant inhibition of SARS-CoV-2 even at low concentrations, despite the low affinity of free AcSA.Accordingly, the AcSA-porphyrin-based glycoclusters of the present invention may be used for the treatment and / or prevention of infectious diseases such as COVID-19.

[0185] 7. Characterization of the neutralizing properties of AcSA-derived porphyrins After assessing the anti-adhesion potential (examples 2.5 and 2.6), we focused on evaluating the neutralizing properties of AcSA-porphyrin glycocluster 011, i.e., its ability to prevent viral entry and therefore infection. A robust virus infectivity assay was used that can be performed in low biosafety conditions. Briefly, a growth-incompetent G-defective vesicular stomatitis virus (VSV) trans-complemented with the SARS-CoV-2 spike protein and encoding a GFP reporter protein (VSV-SARS-CoV-2) was used with or without various concentrations of interfering molecules on A549 cells and A549 cells transduced with the ACE2 receptor. A549-ACE2 was infected with VSV-SARS-CoV-2 at an MOI of 5. Infectivity was monitored by measuring GFP fluorescence in cells 24 hours post-infection. Although A549 cells were not infected by VSV-SARS-CoV-2, overexpression of ACE2 strongly promoted infection. Infectivity assays were then performed incubating VSV-SARS-CoV-2 with SA, 9-AcSA, SA-porphyrin 10, and AcSA-porphyrin 11.

[0186] As expected, both SA and 9-SA did not significantly reduce VSV-SARS-CoV-2 infectivity (Figure 12). Furthermore, while SA-porphyrin 010 did not provide a significant reduction in VSV-SARS-CoV-2 infectivity, we found that 9-AcSA-porphyrin 011 significantly reduced VSV-SARS-CoV-2 infectivity, with an estimated IC in the range of 0.1-1 µM. 50 It was observed that the .alpha.-amino acid was .alpha.-amino acid (.alpha.) (FIG. 12

[0187] This cell-based assay confirms previous results regarding the effect of sialic acid on SARS-CoV-2 binding and further demonstrates that effective inhibition of binding of the virus to its receptor by the glycoclusters of the invention results in a significant reduction in infectivity.

[0188] 8. Comparison of 9-AcSA porphyrin binding to different SARS-CoV-2 strains To assess whether SARS-CoV-2 mutations observed in recent circulating variants affect the efficacy of inhibition of viral binding to its receptor, affinity measurements of 9-AcSA porphyrin 011 to the S1 domain from either the original Wuhan strain or the most recent Omicron strain were performed. Briefly, the S1 domain of the spike protein from either strain was covalently attached to a BioLayer Interferometric Sensor and binding of 9-AcSA porphyrin 011 was assessed.

[0189] No decrease in affinity was observed, and analysis gave affinity constants of the order of 141±4 nM for the Wuhan strain and 100±3 nM for the Omicron strain.

[0190] These results confirm the efficacy of the acetylated sialic acid glycoclusters not only against the original strain of SARS-CoV-2 but also against the Omicron strain, despite it emerging nearly 2 years after the original haplotype and containing many mutations.

Claims

1. A glycocluster comprising at least two acetylated sialic acids covalently attached to a macrocycle, the macrocycle is selected from porphyrins, pillararenes, calixarenes, and fullerenes; each acetylated sialic acid is independently selected from 4-O-acetylated sialic acid, 7-O-acetylated sialic acid, 8-O-acetylated sialic acid, and 9-O-acetylated sialic acid; Glycocluster.

2. The glycocluster of claim 1 , wherein the glycocluster comprises at least four acetylated sialic acids covalently attached to the macrocycle.

3. 2. The glycocluster of claim 1, wherein each acetylated sialic acid is independently selected from 7-O-acetylated sialic acid and 9-O-acetylated sialic acid.

4. The glycocluster of claim 3, wherein each acetylated sialic acid is a 9-O-acetylated sialic acid.

5. 2. The glycocluster of claim 1, wherein the macrocycle is selected from porphyrins, preferably from [Zn(tetraphenylporphyrin)] and tetraphenylporphyrin.

6. The glycocluster of claim 1, further comprising at least one angiotensin-converting enzyme 2 (ACE2) binding inhibitor, preferably selected from an ACE2 binding inhibitor peptide, an ACE2 binding inhibitor protein, and an anti-ACE2 antibody or its antigen-binding fragment.

7. The glycocluster has the formula (I), (Ia), (II), (III) or (IV): 【Chemical 1】 or a pharmaceutically acceptable salt and / or solvate thereof, During the ceremony, Each L １ is, independently, - a single bond, or a linker selected from alkyl, heteroalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, heteroaryl, heteroarylalkyl, and alkylheteroaryl; and wherein the alkyl, heteroalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, heteroaryl, heteroarylalkyl, or alkylheteroaryl optionally comprises at least one coupling product; Each R １ are independently selected from acetylated sialic acid and ACE2 binding inhibitors, provided that at least two R １ is acetylated sialic acid, M is a metal cation; The glycocluster of claim 1.

8. Each L １ is a linker selected from alkyl, heteroalkyl, alkylaryl, arylalkyl, heteroarylalkyl, and alkylheteroaryl, wherein the alkyl, heteroalkyl, alkylaryl, arylalkyl, heteroarylalkyl, or alkylheteroaryl comprises one or two coupling products, preferably at least one of the coupling products is triazolyl.

9. Each L １ -R １ is -(CH ２ ) ｍ -R Ｃ - (CH ２ ) ｎ -R １ , —O—(CH ２ ) ｍ -R Ｃ - (CH ２ ) ｎ -R １ , -C(O)-(CH ２ ) ｍ -R Ｃ - (CH ２ ) ｎ -R １ , -C(O)O-(CH ２ ) ｍ -R Ｃ - (CH ２ ) ｎ -R １ , -phenyl-(CH ２ ) ｍ -R Ｃ - (CH ２ ) ｎ -R １ , and -phenyl-O-(CH ２ ) ｍ -R Ｃ - (CH ２ ) ｎ -R １ is selected from In the formula, R Ｃ is a coupling product, preferably triazolyl, m and n are independently integers ranging from 1 to 8, preferably from 1 to 4, and each R １ is as defined in claim 7, The glycocluster of claim 7.

10. Each R １ The glycocluster of claim 7, wherein is 9-O-acetylated sialic acid.

11. The glycocluster may have the formula (011), (005), (008) or (014): 【Chemistry 2】 【change】 or a pharmaceutically acceptable salt and / or solvate thereof, In the formula, each R １ is the formula (9-AcSA) 【Chemistry 3】 It is of where the wavy line represents the R １ represents the connection point of The glycocluster of claim 10.

12. A pharmaceutical composition comprising a glycocluster according to any one of claims 1 to 11 and at least one pharmaceutically acceptable carrier.

13. A pharmaceutical composition comprising a glycocluster according to any one of claims 1 to 11 and at least one pharmaceutically acceptable carrier for use in the treatment and / or prevention of an infectious disease.

14. The pharmaceutical composition of claim 13, wherein the infectious disease is a coronavirus or picornavirus infection, preferably a SARS-CoV-2 infection.

15. 12. A process for producing a glycocluster according to any one of claims 1 to 11, comprising a step of coupling each of the acetylated sialic acids with the macrocycle, preferably a coupling in which the reaction between a terminal alkyne and an azide results in the formation of a triazolyl group.

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