FIELD OF THE INVENTION

The invention relates to dendritic polyglycerol (dPG) compounds having alkyl-sulfonate/sulfate groups, which irreversibly inhibit viral infection (virucidal effect) in the nanomolar concentration range.

BACKGROUND OF THE INVENTION

The recent emergence of SARS-CoV-2 resulted in a global pandemic (COVID-19), threatening the health of the world’s population and causing dramatic socio-economic damage. It is known that new viruses can emerge or re-emerge every 3-4 years, as previously shown by H1N1, Ebola, H5N1, Zika, etc., all episodes that revealed how our society is unprepared to respond to novel viruses. Indeed, even the percentages of people infected by known viruses such as HSV, HIV, and influenza evidence the urgency of developing novel strategies in fighting viral diseases.

At the moment, there are two weapons against viruses: vaccines and antivirals. Vaccines are preventive drugs composed of modified or attenuated pathogens that are meant to stimulate an immunological bio-response prior to exposure to a live virus. At the moment, vaccines represent the most effective approach to preventing viral infections. However, the durability of protection following vaccination is not 100%. Vaccines are not always available, particularly in underdeveloped countries, and existing vaccines are highly unlikely to be effective against a virus that has not yet emerged. Thus, there remains a large unmet medical need for therapeutic interventions that can help at-risk and infected individuals. Antivirals are drugs designed to fight against viruses and viral infections directly.

The life cycle of a virus is composed of multiple steps: 1) attachment, 2) entry, 3) uncoating, 4) biosynthesis, and 5) assembly and release. The typical mechanism of action of existing antivirals involves inhibiting a step of the viral life cycle, thereby stopping replication. Most antivirals target one or more of steps 2-5, requiring each antiviral to be specific for the manner in which such step is carried out by a particular virus. Given the error-prone nature of viral replication, viruses are often known to mutate and develop resistance to antivirals. The first step of the viral life cycle is attachment. In this step the virus recognizes a host cell using receptors on viral attachment ligands (VALs) that recognize and bind to specific proteins present on host cell membranes. It is known that the VALs of a significant percentage of all viruses target either heparan sulfate proteoglycans (HSPG) or sialic acid (SA) terminal moieties of proteins present on cell membranes. This facilitates a different approach to designing antivirals by mimicking HSPG or SA with a molecule (ranging from polymers to dendrimers, oligomers, nanoparticles, liposomes, monoclonal antibodies, and small molecules) that will bind to a virus and block viral entry. Many of these compounds have shown broad-spectrum activity and limited toxicity, yet none has been translated into a successful drug. The main limitation of such binding inhibitors lies in their mechanism itself. Binding is a reversible event, particularly when the environment (e.g., the bloodstream) surrounding a compound that is bound to a virus causes dissociation of the virus-compound complex, separating the virus from the compound that prevented binding and leaving the virus free to bind again. Unfortunately, dilution is a common event, especially in vivo. Such temporary blocking of viral attachment and/or replication is referred to as virustatic.

The irreversible inhibition of the infectivity of a virus following interaction with an antiviral compound or composition is referred to as virucidal. Many known compounds, ranging from strong surfactants to alcohols, can irreversibly inhibit the infectivity of viruses. Most of these compounds, however, have not translated into acceptable drugs due to issues such as toxicity. Viruses are made of components similar to those of the host, so a drug that damages or interferes with such common components in a virus or an infected cell can also damage (i.e., be toxic to) the host. Only a few compounds have demonstrated virucidal properties together with low toxicity, such as certain reported peptides, but these have been virus-specific, not broad spectrum.

Therefore, there remains a need for virucidal agents that have low toxicity, excellent virucidal activity and broad-spectrum action. SUMMARY OF THE INVENTION

An aspect of the present invention provides a compound of Formula I or a pharmaceutically acceptable salt or ester thereof, wherein dPGC is a dendritic polyglycerol core having an average molecular weight from about 5 to about 100 kDa as measured by GPC;

R can be the same or different and is selected from the group comprising: -H, - SO ₃ ^" , -(CH ₂ ) _z -S-(CH ₂ ) _y -CH ₃ , -optionally substituted C ₅ to C ₃₀ alkyl, -C ₃ to C ₃₀ alkenyl, C ₅ to C ₃₀ ω -hydroxyalkyl, C ₃ to C ₃₀ co-hydroxyalkenyl, C ₅ to C ₃₀ co- hydroxyalkylthioalkyl, C ₅ to C ₃₀ w-hydroxyalkoxyalkyl, C ₅ to C ₃₀ w-haloalkyl, and C ₅ to C ₃₀ w-haloalkoxyalkyl, -(optionally substituted C ₅ to C ₃₀ alkyl)-SO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-SO ₃ ^" , -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-OSO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -O-SO ₃ -, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ -, and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ -; y is an integer from about 4 to about 30; z is an integer from about 2 to about 20; y + z is an integer from about 6 to about 30; and having a DF of at least about 30% as measured by 1HNMR.

In Formula I, "OR" represents the free OH groups within and at the periphery of the core.

Another aspect of the present invention provides a pharmaceutical composition comprising an effective amount of the one or more compounds of the invention and at least one pharmaceutically acceptable excipient, carrier and/or diluent.

Another aspect of the present invention provides the compounds of the invention for use in treating and/or preventing viral infections and/or diseases associated with viruses. Another aspect of the present invention provides a virucidal composition comprising an effective amount of one or more compounds of the invention and at least one suitable carrier or aerosol carrier.

Another aspect of the present invention provides a device comprising the virucidal composition of the invention or one or more compounds of the invention and means for applying and/or dispensing thereof.

Another aspect of the present invention provides a method of disinfection and/or sterilization of non-living surfaces using one or more compounds of the invention or the virucidal composition of the invention.

Another aspect of the present invention provides a use of one or more compounds of the invention or the virucidal composition of the invention for sterilization and/or for disinfection of human or animal skin and/or hair.

Another aspect of the present invention provides a use of one or more compounds of the invention or the virucidal composition of the invention for manufacturing virucidal surfaces.

Another aspect of the present invention provides a device comprising a surface coated with one or more compounds of the invention or with the virucidal composition of the invention.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 shows a dendritic polyglycerol core and a route for its synthesis by anionic ring opening of glycidol.

Figure 1A illustrates another dendritic polyglycerol core.

Figure 2 shows functionalization of dendritic polyglycerols by allyl bromide. The dendritic polyglycerol core is shown as a solid sphere.

Figure 3 shows the ¹ HNMR spectra of dPG _10kDa-allyl.

Figure 4 shows a synthetic route for the synthesis of 11-mercapto-l-undecanesulfonate (MUS). Figure 5 shows ¹ HNMR of 11-mercapto-l-undecanesulfonate (MUS).

Figure 6 shows synthetic route for the synthesis dPG-MUS.

Figure 7 shows ¹ HNMR spectra of MUS (top), dPG-allyl (middle) and dPG-MUS (bottom). Figure 8 shows two-step approach for the synthesis of the dPG-Cl l-sulfate _50/» (RX, R21). Figure 9 shows ¹ HNMR spectrum of dPG-Cl 1 -sulfate (RX).

Figure 10 shows two-step approach for the synthesis of the dPG-Cl l-sulfateso _/» (R19B). Figure 11 shows ¹ HNMR spectrum of dPG-Cl 1 -sulfate (R19B).

Figure 12 shows one-pot approach for the synthesis of the a) dPG-C4-sulfonate (RP3B) and b) dPG-C ₃ -sulfate (RN4) along with ¹ HNMR characterizing data.

Figure 13 shows ¹ HNMR of dPG-C4-sulfonate (top) and dPG-C ₃ -sulfate (bottom).

Figure 14 shows on the top, dose-response curve of dendritic polymer 2 (solid circles), R17 (solid triangles), R18 (solid squares) and Cl (empty squares). At the bottom is a table that reports the characterizations andIC ₅₀ of each compound.

Figure 15 shows the results of a virucidal assay of dendritic polyglycerols 2 and R17. Both show strong virucidal activity.

Figure 16 shows on the top, dose-response curve of dendritic polyglycerols RP3, RP3B, RN4, RN4B. (The compounds RP32 and RN42 are mentioned in the dose-response curve, having been tested at the same time, but do not form part of the invention.) At the bottom are shown the virucidal assay results for RP3B and RNB, showing that these compounds have virustatic (reversible) but not virucidal activity. The table at the bottom summarizes the characterizations and IC ₅₀ for RP3, RP3B, RN4 and RN4B. Figure 17 shows on the top, a dose-response curve for dendritic polyglycerols R21 and RX. At the bottom are shown the virucidal assay results for both compounds, showing virucidal activity. The table at the bottom summarizes their characterizations and IC ₅₀ .

Figure 18 shows on the top, a dose-response curve and the cell viability of dendritic polyglycerol RX. At the bottom, results of the virucidal assay that shows virucidal activity. A table on top recaps the data.

Figure 19 shows on the top, the dose-response curve of dendritic polyglycerol R19B. At the bottom, a table reports the characterizations andIC ₅₀ for RX and R19B.

Figure 20 shows a comparison of the antiviral activity of MUS:OT gold nanoparticles (solid circles) MUS-CD (solid squares) and RX (solid triangles), on the top in concentration (log of ug/ml) and at the bottom in molarity (log of nM). In both cases RX outperforms the other two compounds.

Figure 21 shows the hydrodynamic diameter measured by DLS (dynamic light scattering) of the non-functionalized core dPG (Cl) in phosphate buffer at concentration lmg/mL. The results are for three measurements.

Figure 22 shows gel permeation chromatography (GPC) diagram of the non-functionalized core dPG (Cl). (Mn:7.2 kDa, Mw: 10 kDa, PDI: 1.4)

Figure 23 shows the ¹ HNMR spectra of compounds (2), (R17) and (R18).

Figure 24 shows cytotoxicity data for compounds (RX), (R17) and (R21).

DETAILED DESCRIPTION OF THE INVENTION

All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. The publications and applications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. In addition, the materials, methods, and examples are illustrative only and are not intended to be limiting. Definitions

In the case of conflict, the present specification, including definitions, will control. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in art to which the subject matter herein belongs. As used herein, the following abbreviations and definitions are supplied in order to facilitate the understanding of the present invention.

As used in the specification and claims, the singular form "a", "an" and "the" includes plural references unless the context clearly dictates otherwise.

The term “about” as used in conjunction with a number or a range of numbers, indicates that such number/range will be understood to be approximate. Thus, “about 2” encompasses the integers 1, 2, 3 and 4. The term “about 5 to 30” should be read as "about 5 to about 30" and encompasses, e.g., ranges from 4 to 32, 4 to 28, 6 to 33 and 3 to 27.

As used herein the term “alkenyl” refers to a monoradical branched or unbranched, unsaturated or polyunsaturated hydrocarbon chain, having from about 2 to 30 carbon atoms, more preferably about 5 to 30 carbon atoms and still more preferably about 7 to about 15 carbon atoms. This term is exemplified by groups such as ethenyl, but-2-enyl, hex-2, 5-dienyl, (2E,6E)-

5-methyl-^-nona^, 6-diene and the like. The term “alkenyl” when recited to specify a group linking to another moiety [such as alkenyl sulfonate or -(C ₅ to C ₃₀ alkenylj-SCUH] refers to a diradical branched or unbranched, unsaturated or polyunsaturated hydrocarbon chain consisting of an alkenyl monoradical, a terminal hydrogen of which is substituted by such other moiety.

As used herein, the term “alkyl” refers to a monoradical branched or unbranched saturated hydrocarbon chain containing from 1 to 50 carbon atoms, preferably 5 to 30 carbon atoms. Representative examples of alkyl include, but are not limited to methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl and

6-isopropyl-3-methyl-l0λ ³ -decane. The term "alkyl" when recited to specify a group linking to another moiety [such as alkylsulfate or -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ ^' ] refers to a diradical branched or unbranched saturated hydrocarbon chain derived from an alkyl monoradical, a terminal hydrogen of which is substituted by such other moiety; exemplified by groups such as methylene (-CH ₂ -), ethylene (-CH ₂ CH ₂ -), propylene isomers [e.g., -CH ₂ CH ₂ CH ₂ - and -CH(CH ₃ )-CH ₂ -] and the like. The term “substituted alkyl” refers to an alkyl group in which 1 or more (up to about 5, preferably up to about 3) hydrogen atoms is/are independently replaced by a substituent selected from the group comprising: alkenyl, alkenylthio, alkenyloxy, alkoxy, alkoxyalkoxy, alkoxyalkoxyalkoxy, alkoxyalkoxyalkyl, alkoxyalkyl, alkoxycarbonyl, alkoxycarbonylalkoxy, alkoxycarbonylalkyl, alkoxysulfonyl, alkylamidoalkyl, alkylcarbonyl, alkylcarbonylalkoxy, alkylcarbonylalkyl, alkylcarbonylalkylthio, alkylcarbonyloxy, alkylcarbonylthio, alkylsulfmyl, alkylsulfmylalkyl, alkyl sulfonyl, alkylsulfonylalkyl, alkylthio, alkylthio alkyl, alkylthioalkoxy, alkynyl, alkynyloxy, alkynylthio, aryl, arylcarbonyl, aryloxy, arylsulfonyl, cyano, cyanoalkoxy, cyanoalkyl, cyanoalkylthio, 1,3-dioxolanyl, dioxanyl, dithianyl, ethylenedioxy, formyl, formylalkoxy, formylalkyl, haloalkenyl, haloalkenyloxy, haloalkoxy, haloalkyl, haloalkynyl, haloalkynyloxy, halogen, heterocycle, heterocyclocarbonyl, heterocycloxy, heterocyclosulfonyl, hydroxy, hydroxyalkoxy, hydroxyalkyl, mercapto, mercapto alkoxy, mercapto alkyl, methylenedioxy, and nitro. Preferred substiituents for "substituted alkyl" are selected from the group comprising: alkenyl, alkenylthio, alkoxysulfonyl, alkylcarbonylthio, alkylsulfmyl, alkylsulfmylalkyl, alkyl sulfonyl, alkylsulfonylalkyl, alkylthio, alkylthioalkoxy, alkynylthio, aryl, arylcarbonyl, aryloxy, arylsulfonyl, cyanoalkylthio, dithianyl, heterocyclosulfonyl, hydroxy, hydroxyalkoxy, hydroxyalkyl, mercapto, mercapto alkoxy, and mercapto alkyl.

As used herein, the term “and/or” used in a phrase such as “A and/or B” herein is intended to include “A and B”, “A or B”, “A”, and “B”.

As used in the specification and claims, the term “at least one” used in a phrase such as “at least one C atom” can mean “one C atom” or “two C atoms” or more than two C atoms.

The term “comprise” is generally used in the sense of include, that is to say permitting the presence of one or more features or components. Also as used in the specification and claims, the language “comprising” can include analogous embodiments, as contrasted with the terms “consisting of’ which includes only the embodiment recited and “consisting essentially of’ which includes analogous embodiments to the extent that they do not materially affect the basic and novel characteristics of the claimed invention. As used herein, the term “degree of functionalization” can be abbreviated as “DF The term refers to the number of R functional groups bearing a moiety selected from the group: -(optionally substituted C ₅ to C ₃₀ alkyl)-SO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-SO ₃ ^" , -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-OSO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -O-SO ₃ -, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ - and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ -, determined as a percentage of all of the free hydroxyl groups in a given dendritic polyglycerol. The DF for the dendritic polyglycerols of the invention is at least about 30% (i.e., between about 30% and 100%) or preferably between about 30% and 90%, more preferably between about 40% and 75%, or about 50%. Degree of functionalization can be measured by ¹ HNMR, as described in greater detail below. It will be appreciated by those skilled in the art that some, but not all of the free hydroxyl groups within the dPGC will be functionalized in the syntheses described below and bear the functional moiety represented by R and that it is not feasible to assign the specific location of a particular functional group as existing at one or the other of the two groups shown as R in Formula I. Such functionalization within the dPGC will diminish proceeding inward from the more accessible periphery to the less accessible center of the core, but will be inherrently counted in the degree of functionlization measurement by ¹ HNMR.

As used herein, the term “dendrimer” refers to nano-sized, synthetic, highly branched polymers and oligomers having a well-defined chemical structure that that radially, symmetrically, identically branches from an initial monomeric unit, typically forming spherical (e.g., ovoid, ellipsoid, etc...) macromolecules.

As used herein, the term “dendritic” refers to dendrimer-like highly branched polymers, copolymers or oligomers having a chemical structure resembling that of a dendrimer. Dendritic compounds have a core including a given number of generations of branches or arms, and a plurality of end groups. The branches start from an initial monomeric unit (e.g., trimethylolpropane) but are not identical, typically as the result of incomplete bonding in the early steps of polymerization. The generations of arms consist of structural monomeric units; these can be identical or incomplete for a given generation of arms (or non-identical in the case of dendritic copolymers), and can be the same as the first generation or can branch differently for subsequent generations of arms. (These monomeric units are glycerol in a dendritic polyglycerol.) The generations of arms extend radially in a geometrical progression from the initial monomeric unit until the end (or N ^th ) generation (also described as the periphery). Dendritic (in the sense of dendrimer-like) includes molecules containing non-symmetrical branching. Dense star polymers, starburst polymers and rod-shaped dendrimers can be considered dendritic.

As used herein, the term "dendritic polyglycerol" or "dPG" refers to a glycerol polymer having a plurality of branch points and multifunctional branches that lead to further branching with polymer growth. Dendritic polymers can be obtained by a one- step polymerization process and form a polydisperse system with varying degrees of branching. Figure 1 illustrates the structure and a method for synthesizing a dPG. Methods of making a variety of such polymers are known in the art and further described herein.

As used herein, the term “dendritic polyglycerol core” or “dPGC” refers to an entire dendritic polyglycerol serving as a substrate for functionalization.

As used herein, the term "functionalized" means having chemically bound substituent groups, also referred to as functional groups, functional moieties or functional units, such as bioactive ligands. The dendritic polyglycerols useful for the present invention can contain only a single functional unit per branch or can contain two of the same or different functional units per branch.

As used herein, the terms “glycerol” and “glycerine” and “glycerin” all refer to the monomeric unit propane- 1,2, 3 -triol.

As used herein, the term "hyperbranched" as in "hyperbranched polymer" or "HBP" or “hBP” is used synonymously with "dendritic) when it refers to a polymer or oligomer that branches radially from a central core incorporating plural copies of at least one branching monomer unit. This term is not synonymous with "dendritic" in the case of linear polymers that branch following a cylindrical symmetry or any other branching macromolecule that does not follow a radial branching symmetry. In contrast, hyperbranched polymers (HBPs) incorporate monomers that have three or more reacting groups and thus result in branched polymers. HBPs can be homopolymers composed of a hyperbranched single monomer, or can be copolymers of branching monomers (those able to react at three or more positions) with other branching monomers or with linear monomers (those able to react at only two positions). The HBP compounds employed herein typically are considered to be biocompatible or pharmaceutically acceptable polymers, such that they are suitable for administration to human and/or veterinary subjects. Certain disclosed embodiments of the HBP, e.g., the hyperbranched polyglycerol (hPG) polymer are homopolymers that contain only repeating glycerol subunits. In another example, the HBP can be a heteropolymer that includes one, two or more other polymer subunits. HBPs are well known in the art (see, e.g., Gao and Yan, Prog. Polym. Sci. 29 (2004) 183-275). Examples of HBP compounds, methods of synthesizing them using, for example, a single monomer methodology and double-monomer methodology, modifying, and functionalizing the compounds are disclosed herein and in Macromolecules 1999, 32, 4240- 4246 (polyglycerol) and in Biomaterials 2006, 27:5471-5479, and Gao and Yan, Prog. Polym. Sci. 29 (2004) 183-275.

As used herein, the term “mammal” (for purposes of treatment) refers to any animal classified as a mammal, including humans, domestic and farm animals or pet animals, such as dogs, horses, cats, cows, monkeys etc. Preferably, the mammal is human.

The term “optional” or “optionally” means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances in which it does not. For example, “optionally substituted alkyl” means either “alkyl” or “substituted alkyl,” as defined. It will be understood by those skilled in the art with respect to any group containing one or more substituents that such groups are not intended to introduce any substitution or substitution patterns (e.g., substituted alkyl including optionally substituted cycloalkyl groups, which in turn are defined as including optionally substituted alkyl groups, potentially ad infinitum) that are sterically impractical and/or synthetically non-feasible.

The term "pharmaceutically acceptable ester" refers to esters of the compounds of the present invention, which hydrolyze in vivo and include those that break down readily in the human body to leave the parent compound or a salt thereof. Generally suitable ester groups include, for example, those derived from pharmaceutically acceptable aliphatic carboxylic acids, particularly alkanoic, alkenoic, cycloalkanoic and alkanedioic acids, in which each alkyl or alkenyl moiety advantageously has not more than 6 carbon atoms. Examples of particular esters include, but are not limited to, formates, acetates, propionates, butyrates, acrylates, ethyl succinates, morpholinoethyl esters and the like. Other suitable esters can include sulfate and sulfonate esters. The term "pharmaceutically acceptable salts" as used herein refers to salts that retain the desired biological activity of the compounds the invention and includes pharmaceutically acceptable acid addition salts and base addition salts. Suitable pharmaceutically acceptable acid addition salts of the compounds of Formula I may be prepared from an inorganic acid or from an organic acid, or can be prepared in situ during the final isolation and purification of the compounds of the invention. Examples of such inorganic acids are hydrochloric, sulfuric, and phosphoric acid. Appropriate organic acids may be selected from aliphatic, cycloaliphatic, aromatic, heterocyclic carboxylic and sulfonic classes of organic acids, examples of which are formic, acetic, propionic, succinic, glycolic, gluconic, lactic, malic, tartaric, citric, fumaric, maleic, alkyl sulfonic, arylsulfonic. Suitable pharmaceutically acceptable base addition salts of the compounds of Formula I include metallic salts made from lithium, sodium, potassium, magnesium, calcium, aluminium, and zinc, and organic salts made from organic bases such as choline, diethanolamine, morpholine. Other examples of organic salts are: ammonium salts, quaternary salts such as tetramethylammonium salt; amino acid addition salts such as salts with glycine and arginine. Additional information on pharmaceutically acceptable salts can be found in Remington's Pharmaceutical Sciences, 19th Edition, Mack Publishing Co., Easton, PA 1995. In an embodiment, the pharmaceutically acceptable salt of the compounds of the invention is a sodium salt.

As used herein the terms "subject" or "patient" are well-recognized in the art, and, are used interchangeably herein to refer to a mammal, including dog, cat, rat, mouse, monkey, cow, horse, goat, sheep, pig, camel, and, most preferably, a human, and other, e.g., avian animals, such as a chicken. In preferred embodiments, the terms "subject" or "patient" refer to a human and animals, such as dog, cat, rat, mouse, monkey, cow, horse, goat, sheep, pig, camel, chicken. In some embodiments, the subject is a subject in need of treatment, or a subject being infected by a virus. In other embodiment, a subject can be an animal infected by a virus, such as a chicken. However, in other embodiments, the subject can be a healthy subject or a subject who has already undergone treatment. The term does not denote a particular age or sex. Thus, adult, children and newborn subjects, whether male or female, are intended to be covered. As used herein, the term "sulfate" refers to a group written, interchangeably, as -O-SO ₃ , -O-SO ₃ H, -SO ₄ ² or -SO ₄ H ₂ and includes groups attached to a hydrocarbon linker, for example a Ci- ₅₀ alkyl group as defined herein, to form an "alkylsulfate". C _4-20 alkylsulfates are preferred.

As used herein, the term "sulfonate" refers to a group written, interchangeably, as -SO ₃ or -SO ₃ H and includes groups attached to a hydrocarbon linker, for example a Ci- ₅₀ alkyl group as defined herein, to form an "alkyl sulfonate". C _4-20 alkyl sulfonates are preferred.

As used herein, the term “therapeutically effective amount” refers to an amount of a compound of the invention effective to alter a virus, and to render it inert, in a recipient subject, and/or if its presence results in a detectable change in the physiology of a recipient subject, for example ameliorates at least one symptom associated with a viral infection, prevents or reduces the rate transmission of at least one viral agent.

As used herein, the term “treatment” refers to both therapeutic treatment and prophylactic or preventative measures. Those in need of treatment include those already being infected by a virus, as well as those in which the viral infection is to be prevented or those who are likely to come into contact with a virus. Hence, the mammal, preferably human, to be treated herein may have been diagnosed as being infected by a virus, or may be predisposed or susceptible to be infected by a virus. Treatment includes ameliorating at least one symptom of, curing and/or preventing the development of a disease or condition due to the viral infection. Preventing is meant attenuating or reducing the ability of a virus to cause infection or disease, for example by affecting a post-entry viral event.

As used herein, the term “virucidal” refers to a characterization of antiviral efficacy determined by in vitro testing demonstrating irreversible inhibition of the infectivity of a virus following interaction with an antiviral compound or composition. Even following termination of the interaction (for example, by dilution) and absent any added materials or conditions promoting viral reconstitution, it is essentially impossible for the virus to resume infectivity. Interaction with antiviral compound or composition alters the virus, rendering it inert, and thereby prevents further infections. As used herein, the term “virustatic” refers to a characterization of antiviral efficacy determined by in vitro testing demonstrating reversible inhibition of the infectivity of a virus following interaction with an antiviral compound or composition. Once the interaction terminates (for example, by dilution) and absent any added materials or conditions promoting viral reconstitution, it is possible for the virus to resume infectivity.

Compounds of the Present Invention

A biomimetic strategy has been employed to develop broad-spectrum virucidal drugs. To limit toxicity, it has been decided to stay away from known bio-toxic approaches and to concentrate on mimicking cell-receptors, so to strongly attach to their corresponding viral ligand and generate local viral deformation that would ultimately lead to irreversible viral damage, possibly to viral disassembly. To achieve broad-spectrum efficacy, it was aimed at virus-cell interactions that are common to many viruses. One of these interactions is that between viruses and cell-surface attachment receptors that represent the very first step of the virus replication cycle. Many viruses, including HIV-1, HSV, HCMV, HPV, Respiratory syncytial virus (RSV) and filoviruses, exploit heparan sulfate proteoglycans (HSPGs) as attachment receptors, as HSPGs are expressed on the surface of almost all eukaryotic cell types. The binding between viruses and HSPGs usually occurs via the interaction of stretches of basic amino acids on viral proteins (basic domains) with the negatively charged sulfated groups of heparan sulfate (HS) chains on the glycocalix of the cell surface.

An aspect of the present invention provides a novel class of virucidal compounds designed to mimic cell surface sugars and have very low toxicity as well as broad-spectrum activity against HSV-2 and other HSPG-seeking viruses at nanomolar concentration with a virucidal effect (i.e. the ability of irreversibly inhibiting viral infectivity). The virucidal compounds of the invention have been prepared by partial or complete functionalization of dendritic polyglycerols (dPG), as a soft biocompatible platform, with different sufficiently long ligands having functional groups, such as sulfate and sulfonate. dPG is a highly branched polymer with a flexible polyether backbone and high density of surface hydroxyl groups that can be further modified with different ligands. Another aspect of the present invention provides a compound, pharmaceutically acceptable salt or a pharmaceutically acceptable ester of Formula I wherein dPGC is a dendritic polyglycerol core in which OR represents the free OH groups within and at the periphery of the core, having an average molecular weight from about 5 to about 100 kDa as measured by GPC;

R can be the same or different and is selected from the group comprising: -H, - SO ₃ ^" , -(CH ₂ ) _z -S-(CH ₂ ) _y -CH ₃ , -optionally substituted C ₅ to C ₃₀ alkyl, -C ₃ to C ₃₀ alkenyl, C ₅ to C ₃₀ co-hydroxyalkyl, C ₃ to C ₃₀ ω -hydroxyalkenyl, C ₅ to C ₃₀ co- hydroxyalkylthioalkyl, C ₅ to C ₃₀ co-hydroxyalkoxyalkyl, C ₅ to C ₃₀ w-haloalkyl, and C ₅ to C ₃₀ w-haloalkoxyalkyl, -(optionally substituted C ₅ to C ₃₀ alkyl)-SO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-SO ₃ ^" , -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-OSO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -O-SO ₃ -, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ -, and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ -; y is an integer from about 4 to about 30; z is an integer from about 2 to about 20; y + z is an integer from about 6 to about 30; and having a DF of at least about 30% as measured by 1HNMR.

Those skilled in the art will appreciate that the substituent R as shown in Formula I illustrates that the same or different moieties can be substituted on any of the dPGC free hydroxyl groups.

R is preferably functionalized with a sulfate- or sulfonate-bearing aliphatic chain, more preferably having from about 6 to about 20 carbon atoms, and still more preferably from about 8 to about 15 carbon atoms. The variable "y" is preferably an integer from about 4 to about 20, more preferably from about 8 to about 13 and still more preferably about 11. The variable "z" is preferably an integer from about 2 to about 10, more preferably from about 2 to about 5, and still more preferably about 3. The sum "y + z" is preferably an integer from about 10 to about 16, more preferably about 14. The dendritic polyglycerol core (dPGC) according to the invention has a size from about 4 to 15 nm and a molecular weight from about 5 to 100 kDa. It is composed of repeated units of glycerine with the formula (RO-CH ₂ ) ₂ CH-OR wherein R = H or an adjacent glycerine unit on a multifunctional polyhydroxy starter molecule having a plurality of OH groups, for example 2 to 4 OH groups, such as 2-ethyl-2-(hydroxymethyl)propane-l,3-diol. Figure 1 shows a representative example of a dPG formed from 2-ethyl-2-(hydroxymethyl)propane-l,3-diol polymerized with glycerol monomers. One particular dPGC identified as "(Cl)" has a hydrodynamic diameter (size) of 5.34 ± 0.293 nm and molecular weight => GPC (H ₂ O) M _n = 7.2 kDa, M _w = 10.4 kDa. Figures 21 and 22 show the results of dynamic light scattering (DLS) characterizing the size, and gel permeation chromatography (GPC) characterizing the molecular weight, of dendritic polyglycerol core (Cl). It should be noted that the weight and size of functionalized dPG products of Formula I will be greater than the weight and size of the core (dPGC) that was functionalized.

In one aspect of the invention, the R groups that contribute to DF are selected from the group comprising: -(optionally substituted C ₅ to C ₃₀ alkyl)-SO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-SO ₃ ^" , -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ ^" , -(C ₅ to C ₃₀ alkenyl)-OSO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ ^" , -(CH ₂ ) _z -O-(CH ₂ ) _y -O-SO ₃ -, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ ^" and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ ^' . In another aspect, particularly the compositions of matter, the R groups that contribute to DF are selected from the group comprising: -(C ₅ to C ₃₀ alkenyl)-SO ₃ -, -(C ₅ to C ₃₀ alkenyl)-OSO ₃ -, -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ -, -(CH ₂ ) _z -O-(CH ₂ ) _y -OSO _3- , -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ ^' and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ ^' . In yet another aspect, the R groups that contribute to DF are selected from the group comprising: -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ -, -(CH ₂ ) _z -O-(CH ₂ ) _y -O-SO ₃ -, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ - and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ ^' .

According to an embodiment, the present invention provides a compound of Formula I comprising a dendric polyglycerol core as defined above functionalized with a plurality of same or different R substituents as defined above that are bound to the core, provided however, that not each of said R substituents necessarily comprises a sulfate and/or a sulfonate group, and wherein the degree of functionalization is as defined above. R will comprise a hydrogen or an incompletely reacted precursor group when not fully functionalized with a sulfate- or sulfonatebearing aliphatic chain. Incompletely reacted precursor groups for R can include, without limitation: -SO ₃ ^" , -optionally substituted C ₅ to C ₃₀ alkyl, -C ₃ to C ₃₀ alkenyl, C ₅ to C ₃₀ w-hydroxyalkyl, C ₅ to C ₃₀ w-hydroxyalkenyl, C ₅ to C ₃₀ w-hydroxyalkylthioalkyl, C ₅ to C ₃₀ w-hydroxyalkoxyalkyl, C ₅ to C ₃₀ co-haloalkyl, and C ₅ to C ₃₀ ω -haloalkoxyalkyl.

Another aspect of the present invention provides a compound of Formula II wherein: dPGC is a dendritic polyglycerol core having a size from 4 to 200 nm and a molecular weight from 5 to 100 kDa, each Ri, independently, is optionally substituted alkyl-based ligand selected from the group comprising -(CH ₂ ) _y -CH ₃ , -(CH ₂ ) _y -SO ₃ H, -(CH ₂ ) _y -OSO ₃ H, -(CH ₂ ) ₃ -S-(CH ₂ ) _y -OSO ₃ H. In some embodiments, each Ri, independently, is optionally substituted alkyl-based ligand selected from the group comprising -(CH ₂ ) _y -SO ₃ H, -(CH^-OSChH, -(CH ₂ ) ₃ -S-(CH ₂ ) _y - SO ₃ H, -(CH ₂ ) ₃ -S-(CH ₂ ) _y -OSO ₃ H. In other embodiments, each Ri, independently, is optionally substituted alkyl-based ligand selected from the group comprising -(CH ₂ ) _y. CH ₃ , - (CH ₂ ) ₃ -S-(CH ₂ ) _y -SO ₃ H, -(CH ₂ ) ₃ -S-(CH ₂ ) _y -OSO ₃ H. In further embodiments, at least one Ri is not -(CH ₂ ) _y -SO ₃ H and/or -(CH ₂ ) _y -OSO ₃ H. In some other embodiments, each Ri, independently, is optionally substituted alkyl-based ligand selected from the group comprising -(CH ₂ yCH ₃ , -(CH ₂ ) _y -SO ₃ H, -(CH ₂ )-OSCriH, -(CH ₂ ) ₃ -S-(CH ₂ ) _y -OSO ₃ H, provided that at least one Ri is not -(CH ₂ ) _y -SO ₃ H and/or -(CH ₂ ) _y -OSO ₃ H. each R ₂ , independently, is -H, -SO ₃ H, -CH ₂ -CH=CH ₂ , -(CH ₂ ) ₃ -S-(CH ₂ ) ₂ -CH _3. In some embodiments, each R ₂ , independently, is -H or -SO ₃ H. In other embodiments, each R ₂ , independently is -H, -CH ₂ -CH=CH ₂ , -(CH ₂ ) ₃ -S-(CH ₂ ) ₂ -CH _3. In other embodiments, each R ₂ , independently is -CH ₂ -CH=CH ₂ , -(CH ₂ ) ₃ -S-(CH ₂ ) ₂ -CH _3. In further embodiments, at least one R ₂ is not -H or -SO ₃ H. In some other embodiments, each R ₂ , independently, is -H, -SO ₃ H, -CH ₂ - CH=CH ₂ , -(CH ₂ ) ₃ -S-(CH ₂ ) ₂ -CH ₃ , provided that at least one R ₂ is not -H or -SO ₃ H. y is at least 4, preferably from 4 to 30 or from 8 to 20, most preferably y is 7 to 11 or most preferably y is 11. In other embodiments, y is at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11. In other embodiments, y is at maximum 100, at maximum 70, at maximum 50, at maximum 40, at maximum 30, at maximum 25, at maximum 20, at maximum 15 and having a degree of functionalization (d) that is at least 30%, at least 31%, at least 35%, at least 40%, or at least 45%; at maximum 100%, at maximum 95%, or at maximum 90%; preferably from 30% to 100%, from 31% to 100%, from 35% to 100% from 40% to 100%, from 30% to 90%, 31% to 90%, 35% to 90% or 40% to 90%, or a pharmaceutically acceptable salt thereof.

The compounds of the invention have a virucidal effect in the nanomolar range. The fully organic core overcomes the issue of possible long-term accumulation, that happens with gold nanoparticles or other bio-incompatible materials.

Another aspect of the invention discloses a pharmaceutical composition comprising an effective amount of one or more compounds of the invention and at least one pharmaceutically acceptable excipient, carrier and/or diluent.

Nomenclature

In the present specification, the dendritic polyglycerols are named using the following format: The subscript following “dPG” indicates the weight of the dendritic polyglycerol being named, e.g., dPGio _k o _a names a dendritic polyglycerol having a weight of 10 kDa. The text following such weight indication identifies the functional group(s) and the subscript that follows indicates the degree of functionalization. Thus, dPGiokDa-Cn-sulfate5o%/sulfonate5o% names a dendritic polyglycerol having the structure shown below in Formula RX: which is a compound of Formula 1 where the dPG core has a weight of 10 kDa, where about half of the free hydroxyls are undecane sulfate with a DF of 50%, and the other half are a sulfonate with a DF of 50%.

Synthetic Reaction Parameters

The terms "solvent", "inert organic solvent" or "inert solvent" mean a solvent inert under the conditions of the reaction being described in conjunction therewith [including, for example, benzene, toluene, acetonitrile, tetrahydrofuran ("THF"), dimethylformamide ("DMF"), chloroform, methylene chloride (or dichloromethane), diethyl ether, methanol, pyridine and the like]. Unless specified to the contrary the solvents used in the reactions of the present invention are inert organic solvents. Reactions take place at room temperature and 1 atmosphere of pressure unless otherwise indicated.

Isolation and purification of the compounds and intermediates described herein can be effected, if desired, by any suitable separation or purification procedure such as, for example, filtration, extraction, crystallization, column chromatography, thick-layer chromatography, or a combination of these procedures. Specific illustrations of suitable separation and isolation procedures can be had by reference to the examples hereinbelow. However, other equivalent separation or isolation procedures can, of course, also be used.

Synthesis of the Compounds of Formula I

Syntheses of the compounds of Formula I are described below with reference to the Reaction Schemes.

Starting Materials

The starting materials, such as the glycerol (1) are commercially available or can be readily prepared by those skilled in the art using commonly employed synthetic methodology.

Preparation of dendritic polyglycerol

As illustrated in Reaction Scheme 1, a glycerol such as trimethylolpropane ("TMP") (1) is deprotonated with about 0.4 molar equivalents ("eq.") of potassium methoxide solution (e.g., in methanol). The methanol is evaporated at about 60 °C under vacuum (about 3 mbar). The synthesis reactor is heated to about 100 °C and an excess (e.g., number of branches desired x 3 eq.) of (oxiran-2-yl)methanol ("glycidol") is added slowly, e.g., over a period of about 24 hours, providing ring opening multi-branching polymerization conditions, to afford a dendritic polyglycerol (2), such as Cl, the dPG illustrated in Figure 1 or the dPG illustrated in Figure 1 A, which can be conventionally isolated and purified. The molecular weight of the resulting dPG can be controlled by adjusting the molar ratio of glycidol to TMP and the ring-opening polymerization reaction time accordingly. It will be appreciated by those skilled in the art that the resulting product will be a mixture of dPGs falling within a narrow range of molecular weights, such as 8 kDa to 12 kDa, and will have an average molecular weight such as 10 kDa.

Reaction Scheme 2

Preparation of mercapto-alkyl-sulfonate (6)

As illustrated in Reaction Scheme 2, Step 1, an co-halo-l-alkene of formula 3 where y can be 1 to 27 (especially where y is 6 to 12 - other sizes will entail modification of the conditions) and "Halo" is bromo, chloro, fluoro or iodo, preferably bromo, is contacted with sodium sulfite (about 2 eq.) in a suitable solvent such as methanol and DI water. The mixture is refluxed (at about 102°C) for about 48 hours to afford the corresponding co-sulfonyl-l-alkene (4) which is conventionally isolated and purified (e.g., diethyl ether extraction, evaporation, drying and removal of inorganic salts with pure ethanol and filtration). As illustrated in Reaction Scheme 2, Step 2, the co-sulfonyl-l-alkene (4) obtained in Step 1 is dissolved in a suitable solvent (e.g., methanol) to afford a clear solution, removing any precipitate by filtration (to improve yield). An excess of thioacetic acid (about 2-3 eq.) is added to the solution and it is stirred in front of a UV lamp for about 12 hours. The solution is evaporated until the resulting solid residue becomes colored (e.g., orange-red), after which the solid is washed to remove the colored material (e.g., with diethyl ether) until additional colored material can no longer be removed. The resulting colored solid is dried (e.g., under high vacuum) and dissolved in a suitable solvent (e.g., methanol) to afford a colored (e.g., yellow) solution. A suitable amount of carbon black is added to the solution followed by vigorous mixing, and the mixture is filtered (e.g., through celite in fluted filter paper) to afford a clear solution, from which the solvent is completely evaporated to afford the corresponding sodium acetylthio-alkyl-1 -sulfonate (5).

As illustrated in Reaction Scheme 2, Step 3, the sulfonate (5) obtained in the previous step is refluxed in 1M HC1 for 12 hours, after which the mixture is brought to pH ~3 by addition of 1M NaOH followed by Di-water to create a volume suitable for the scale of the reaction (e.g., 1 L). The resulting solution is kept at 4°C and crystallized over about 12 hours to yield the corresponding mercapto-alkyl sulfonate product (6) that is conventionally isolated and purified (e.g., centrifugation and dried under high vacuum). (Additional product can be extracted from the supernatant of the centrifugation step, by reducing volume and keeping it at 4°C.)

Reaction Scheme 3

Preparation of dPGC-di-allyl (8)

As illustrated in Reaction Scheme 3, a dPG (2) is functionalized in preparation for a subsequent thiol-ene click reaction, by converting the free hydroxyl groups to allyl groups through reaction with an allyl halide (e.g., bromide) (7) where z can be 0 to about 18. The dPG is dissolved in a suitable solvent (e.g., DMF) and the reaction takes place over about 12 hours; it is performed in dry condition in presence of NaH as base for deprotonation of hydroxyl groups. The resulting allyl-functionalized product (8) is conventionally isolated and purified (e.g., solvent removed under vacuum and purification by dialysis in MeOH for 2 days). The degree of functionalization (DF) can be controlled by adjusting the ratio of allyl halide to dPG, the amount of NaH, the reaction time and/or conditions, and is confirmed by 'H NMR. For example, by limiting the equivalents of allyl halide and NaH, the product corresponding to (8) where more of the groups corresponding to R remain hydrogen can be obtained.

Reaction Scheme 4

Synthesis of mercapto alkyl sulfonate functionalized dPG (Formula la)

As illustrated in Reaction Scheme 4, a mercapto alkyl sulfonate (6) where z can be 0 to about 18 is conjugated to a dPG-allyl functionalized core (8) where y can be 1 to 27 by dissolving the reactants in a suitable solvent (e.g., watenmethanol). A catalytic amount of tris(2- carboxyethyljphosphine hydrochloride (TCEP-HC1) is added to reduce disulfide bonds and avoid oxidation of the thiol intermediate. The solution is degassed (e.g., by flushing argon through the reaction mixture for about 10 minutes) and the reaction mixture is stirred and irradiated with UV light (e.g., using a high-pressure UV lamp at room temperature) for about 6 hours. The solution is then dialyzed (MWCO 2 kDa) against watenmethanol for about 2 days. The solvent is evaporated under reduced pressure and the resulting product (Formula la) is conventionally isolated and purified (e.g., by lyophilization). The degree of functionalization can be controlled by adjusting the ratio of mercapto alkyl sulfonate to dPG allyl and/or time of UV irradiation.

Synthesis of dPGC-R-alkanol (10)

As illustrated in Reaction Scheme 5, Step 1 dPG (2) is reacted with an co-halo-alkan-l-ol (9) (about 1.5 eq.) (where p is an integer from 5 to 30) in the presence of NaH (about 2 eq.) (as a base for deprotonation of the dPG hydroxyl groups) to obtain the corresponding dPG-C _p -OH with 50% of degree of functionalization. The reaction mixture is allowed to stir for about 24 hours at about 40 °C and is then quenched by adding methanol and purified by dialysis against methanol to afford the corresponding dPGC-alkanol (10).

Synthesis of dPGC-R-alkyl-sulfate/sulfate (Formula lb)

As illustrated in Reaction Scheme 5, Step 2 both hydroxyl groups of (10) are sulfated, e.g., by contact with pyridine sulfur tri oxide complex or chlorosulfonic acid (about 1.5 eq.), in dry DMF at about 60 °C for about 12 hours. The reactions are quenched with water, and the pH adjusted to 8 by addition of NaOH solution. Solvent is evaporated under reduced pressure, and the product sulfated dPG (Formula lb) is isolated and purified conventionally (e.g., dissolved in brine, dialysed with a NaCl solution, using an ever-decreasing NaCl concentration, until the medium is changed with distilled water). Degree of functionalization is determined by ¹ HNMR. Synthesis of dPGC-R-thioalkanol/alkyl-thio-alkyl

As illustrated in Reaction Scheme 6, Step 1, a dPG-allyl functionalized core (8), about 0.6 eq. of an co-mercapto-l-alkanol (11) and a catalytic amount of 2,2-dimethoxy-2- phenyl acetophenone (DMPA) (as a radical initiator) are dissolved in a suitable solvent (e.g., watenmethanol). A catalytic amount of tris(2-carboxyethyl)phosphine hydrochloride (TCEP- HC1) is added (to avoid oxidation of the thiol intermediate). The solution is degassed (e.g., by flushing argon through the reaction mixture for about 10 minutes). The reaction mixture is stirred and irradiated with UV light using a high-pressure UV lamp at room temperature for about 4 hours. An excess of 1-propanthiol is added to the mixture followed by additional (e.g., about 4 hours) UV irradiation (to quench the remaining allyl group). The solution is then dialyzed (e.g., MWCO 2 kDa) against watenmethanol for about 2 days. The solvent is evaporated (e.g., under reduced pressure) and the resulting intermediate (12) is carried forward in the next step.

Synthesis of dPGC-R-thioalkyl-sulfate/alkyl-thio-alkyl

In the next step the hydroxyl group of (12) is sulfated by reaction with pyridine sulfur trioxide complex; the reaction takes place in a suitable solvent (e.g., dry DMF) at about 60 °C for about 12 hours. The reaction is then quenched with water, and the pH is adjusted to 8 (e.g., by addition of NaOH solution). The solvent is evaporated (e.g., under reduced pressure) and the product is dissolved (e.g., in brine). Dialysis is performed with a NaCl solution, using an ever-decreasing NaCl concentration, until the medium is changed with distilled water. The final product (Formula Ic) is obtained, e.g., after lyophilization.

Synthesis of dPGC-R-alkyl-sulfonate/alkyl-sulfonate

As illustrated in Reaction Scheme 7, dPG (2) is reacted with an w -halo-alkyl- 1 -sulfonate (13) (where p is an integer from 5 to 30) in the presence of NaH. The reaction mixture is allowed to stir for about 24 hours at about 60 °C and the corresponding alkyl sulfonate product of Formula Id is conventionally isolated and purified.

Synthesis of dPGC-Ralkenyl-sulfonate/alkyl-sulfonate or H As illustrated in Reaction Scheme 8, dPG (2) is reacted with an co-bromo-alkenyl-l -sulfonate (14) (where p is an integer from 1 to 28, q is an integer from 1 to 26, and p+q is an integer from 4 to 30) in the presence of NaH. The reaction mixture is allowed to stir for about 24 hours at about 60 °C to afford one or both of the corresponding alkenyl sulfonate products of Formula Ie and Formula If, which are conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography), isolated and purified. Synthesis of hydroxyalkenyl dPGs

As illustrated in Reaction Scheme 9, Step 1, dPG (2) is reacted with an co-bromo-alken-l-ol (15) (where p is an integer from 1 to 28, q is an integer from 1 to 26, and p+q is an integer from 4 to 30) in the presence of NaH. The alkenol (15) is preferably added slowly, dropwise, to minimize the formation of dimers. The reaction mixture is allowed to stir for about 24 hours at about 60 °C to afford one or both of the corresponding hydroxyalkenyl products (16) and/or (17), which can be carried forward together or conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) and carried forward independently. The products can be conventionally isolated and purified.

Synthesis of dPGC-R-alkenyl-sulfate/alkenyl-sulfate or H

As illustrated in Reaction Scheme 9, Step 2, hydroxyalkenyl dPG (16) and/or (17) is sulfated, for example, by reaction with pyridine sulfur trioxide complex; the reaction takes place in a suitable solvent (e.g., dry DMF) at about 60 °C for about 12 hours. The reaction is then quenched with water, and the pH is adjusted to 8 (e.g., by addition of NaOH solution). The solvent is evaporated (e.g., under reduced pressure) and the product(s) is/are dissolved (e.g., in brine). Dialysis is performed with a NaCl solution, using an ever-decreasing NaCl concentration, until the medium is changed with distilled water. The final product(s) (Formula Ig and/or Formula Ih) is/are obtained, e.g., after lyophilization, conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) if necessary, conventionally isolated and purified.

Synthesis of hydroxyalkyl dPGs

As illustrated in Reaction Scheme 10, Step 1, dPG (2) is reacted with an co-halo-alkan-l-ol (18) in the presence of NaH. The alkanol (18) is preferably added slowly, dropwise, to minimize the formation of dimers. The reaction mixture is allowed to stir for about 24 hours at about 60 °C to afford one or both of the corresponding hydroxyalkanol products (19) and/or (20), which can be carried forward together or conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) and carried forward independently. The products can be conventionally isolated and purified.

Synthesis of hydroxyalkoxyalkyl dPGs

As illustrated in Reaction Scheme 10, Step 2, hydroxyalkyl (19) and/or (20) is/are reacted with an ω -halo-alkan-l-ol (21) in the presence of NaH. The alkanol (21) is preferably added slowly, dropwise, to minimize the formation of dimers. The reaction mixture is allowed to stir for about 24 hours at about 60 °C to afford one or all of the corresponding hydroxyalkyl / hydroxyalkoxy products (22), (23) and/or (24), which can be carried forward together or conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) and carried forward independently. The products can be conventionally isolated and purified.

Synthesis of dPGC where R is alkoxy-sulfate or alkoxy-alkyl-sulfate

As illustrated in Reaction Scheme 10, Step 3, dPG (22), (23) and/or (24) is/are sulfated, for example, by reaction with pyridine sulfur tri oxide complex; the reaction takes place in a suitable solvent (e.g., dry DMF) at about 60 °C for about 12 hours. The reaction is then quenched with water, and the pH is adjusted to 8 (e.g., by addition of NaOH solution). The solvent is evaporated (e.g., under reduced pressure) and the product(s) is/are dissolved (e.g., in brine). Dialysis is performed with a NaCl solution, using an ever-decreasing NaCl concentration, until the medium is changed with distilled water. The final product(s) (Formula Ii, Formula Ij and/or Formula Ik) is/are obtained, e.g., after lyophilization, conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) if necessary, conventionally isolated and purified.

Synthesis of haloalkyl and haloalkoxy dPGs

As illustrated in Reaction Scheme 11, Step 1, an hydroxyalkyl / hydroxyalkoxy dPG (22), (23) and/or (24), prepared, e.g., as described with regard to Reaction Scheme 10, is halogenated (e.g., brominated, as described in Zhou et ak, Polym. Chem., 2017, 8, 2189) with stirring in a suitable solvent (e.g., anhydrous CH ₂ CI2) using an excess (e.g., 5 eq. to -OH) of tetrabutylammonium bromide (TBAB). To the stirring solution is added a similar excess of l,8-diazabicyclo(5.4.0)undec-7-ene (DBU) followed by a similar excess of XtalFluor-E (Aldrich), cooling to 0 °C. Stirring is continued at room temperture for about 24 hours. The mixture is precipitated (e.g., into methanol/water) with several drops of hydrochloric acid to afford one or all of the corresponding halogenated products (25), (26) and/or (27) where "halo" is bromo, chloro, fluoro or iodo, preferably bromo, which can be carried forward together or conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) and carried forward independently. The products can be conventionally isolated and purified.

Synthesis of dPGC where R is alkyl-sulfonate or alkoxy-alkyl-sulfonate

As illustrated in Reaction Scheme 11, Step 2, a haloalkyl or haloalkoxy dPG of formulae (25), (26) and/or (27) is contacted with sodium sulfite (about 2 eq.) in a suitable solvent such as methanol and DI water. The mixture is refluxed (at about 102°C) for about 48 hours to afford the corresponding sulfonated dPG products of Formula II, Formula Im and/or Formula In, which can be conventionally separated (e.g., by gel permeation chromatography or size exsclusion chromatography) if necessary, and conventionally isolated and purified (e.g., diethyl ether extraction, evaporation, drying and removal of inorganic salts with pure ethanol and filtration).

Preferred Processes and Last Steps

A mercapto-alkyl sulfonate is conjugated to a dPG-allyl functionalized core in a thiol-ene click reaction under UV light to give the corresponding mercapto alkyl sulfonate functionalized dPG.

Both hydroxyl groups of a dPGC-alkanol are sulfated with pyridine sulfur trioxide complex to give the corresponding dPGC-alkanol sulfate/sulfate.

A dPGC-alkylthioalkanol/alkylthioalkyl is sulfated with pyridine sulfur trioxide complex to give the corresponding dPGC-alkyl-thio-alkyl sulfonate/alkylthioalkyl.

Preferred Compounds

Preferred for the compounds, pharmaceutical formulations, methods of manufacture and use of the present invention are the following combinations and permutations of substituent groups of Formula I (sub-grouped, respectively, in increasing order of preference) where:

• dPGC has a size from about 4 to 10 nm and an average molecular weight from about 5 to 25 kDa o Especially where the dPGC size is about 5 nm and average molecular weight is about 10 kDa o Especially where DF is at least about 40%

^■ Particularly where DF is at least 50%

• Preferably where DF is about 50%. o More preferably where R contributing to DF is selected from the group comprising: -(optionally substituted C ₅ to C ₃₀ alkyl)-SO ₃ H, -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ H, -(CH ₂ ) _z -O-(CH ₂ ) _y -SO ₃ H, -(CH ₂ ) _z -O-

(CH ₂ ) _y -OSO ₃ H, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ H, and -(CH ₂ ) _Z -S- (CH ₂ ) _y -OSO ₃ H.

^■ Still more preferably, R contributing to DF is selected from the group comprising: -(C ₈ to C ₁₅ alkyl)-SO ₃ H, -(C ₈ to C ₁₅ alkyl)-OSO ₃ H, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ H, and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ H. • Even more preferably, R not contributing to DF is selected from the group comprising: -C ₈ to C ₁₅ alkenyl, C ₈ to C ₁₅ ω -hydroxyalkyl, and C ₈ to C ₁₅ ω-hydroxyalkylthioalkyl. o Especially where R not contributing to DF is H,SO ₃ , -CH ₂ -CH=CH ₂ or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH ₃ .

^■ Most preferably, where y is from about 8 to 13, z is from about 2 to 5, and y+z is from about 10 to 16. o Especially where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16.

• Even more preferably where R not contributing to DF isH, SO ₃ , -CH ₂ -CH=CH ₂ or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH _3.

• Even more preferably where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16.

^■ Still more preferably where R not contributing to DF is selected from the group comprising: -C ₈ to C ₁₅ alkenyl, C ₈ to C ₁₅ ω -hydroxyalkyl, and C ₈ to C ₁₅ co -hydroxy alkylthioalkyl .

^■ Still more preferably where R not contributing to DF is H, SO ₃ , -CH ₂ -CH=CH ₂ or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH _3.

^■ Still more preferably where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16. o More preferably where R contributing to DF is selected from the group comprising: -(Cs to C ₁₅ alkyl)-SO ₃ H, -(Cs to C ₁₅ alkyl)-OSO ₃ H, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ H and -(CH ₂ ) _Z -S- (CH ₂ ) _y -OSO ₃ H. o More preferably where R not contributing to DF is selected from the group comprising: -C ₈ to C ₁₅ alkenyl, C ₈ to C ₁₅ w -hydroxy alkyl, and C ₈ to C ₁₅ w-hydroxyalkylthioalkyl. o More preferably where R not contributing to DF is H, , -CH ₂ -CH=CH ₂ or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH ₃ ,. o More preferably where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16.

• R contributing to DF is selected from the group comprising: -(optionally substituted C ₅ to C ₃₀ alkyl)-SO ₃ H, -(optionally substituted C ₅ to C ₃₀ alkyl)-OSO ₃ H, -(CH ₂ ) _z -O-(CH ₂ ) _y - H, -(CH ₂ ) _z -O-(CH ₂ ) _y -OSO ₃ H, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ H, and -(CH ₂ ) _z -S-(CH ₂ ) _y - OSO ₃ H. o Especially where DF is at least about 40% o Especially where DF is at least 50%

^■ Particularly where DF is about 50%. o Especially where R contributing to DF is selected from the group comprising: -(C ₈ to C ₁₅ alkyl)-SO ₃ H, -(C ₈ to C ₁₅ alkyl)-OSO ₃ H, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ H and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ H.

^■ Particularly where R not contributing to DF is selected from the group comprising: -C ₈ to C ₁₅ alkenyl, C ₈ to C ₁₅ w-hydroxyalkyl, and C ₈ to C ₁₅ w -hydroxy alkylthioalkyl .

• Preferably where R not contributing to DF is H, SO ₃ , -CH ₂ -CH=CH ₂ or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH _3. o More preferably, where y is from about 8 to 13, z is from about 2 to 5, and y+z is from about 10 to 16.

^■ Still more preferably where DF is at least 40%

^■ Still more preferably where DF is at least 50%

• Even more preferably where DF is about 50%.

• Prefereably where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16.

^■ Particularly where R not contributing to DF is H, SO ₃ , -CH ₂ -CH=CH ₂ , or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH _3.

^■ Particularly where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16. o Especially where R not contributing to DF is selected from the group comprising: -C ₈ to C ₁₅ alkenyl, C ₈ to C ₁₅ co-hydroxyalkyl, and C ₈ to C ₁₅ ω -hydroxyalkylthioalkyl. o Especially where R not contributing to DF is H,SO ₃ , -CH ₂ -CH=CH ₂ , or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH ₃ . o Especially where y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16.

• R contributing to DF is selected from the group comprising: -(C ₈ to C ₁₅ alkyl)-SO ₃ H,

-(C ₈ to C ₁₅ alkyl)-OSO ₃ H, -(CH ₂ ) _z -S-(CH ₂ ) _y -SO ₃ H, and -(CH ₂ ) _z -S-(CH ₂ ) _y -OSO ₃ H.

• R not contributing to DF is selected from the group comprising: -C ₈ to C ₁₅ alkenyl, C ₈ to C ₁₅ co-hydroxyalkyl, and C ₈ to C ₁₅ ω -hydroxyalkylthioalkyl.

• R not contributing to DF is H,SO ₃ , -CH ₂ -CH=CH ₂ , or -(CH ₂ ) _z -S-(CH ₂ ) _y -CH ₃ .

• y is from about 8 to 13, z is from about 2 to 5, and y + z is from about 10 to 16.

• DF is at least 40%, at least 50%, about 40 to 70%, or about 50%

As illustrated with regard to the group of compounds described in the Examples, the above- described groups and sub-groups are individually preferred and can be combined to describe further preferred aspects of the invention.

Particularly preferred for the compounds, pharmaceutical formulations, methods of manufacture and use of the present invention are the following: RX, R21, R17, R19B and 2.

More preferred for the compounds, pharmaceutical formulations, methods of manufacture and use of the present invention are the following: RX, R21 and R19B.

Presently TRX is most preferred for the compounds, pharmaceutical formulations, methods of manufacture and use of the present invention.

Utility, Testing, Administration and Formulation

General Utility

The compounds of the invention are useful for treating an HSPG dependent virus, i.e., a virus that exploits heparan sulfate proteoglycans (HSPGs) as attachment receptors on eukaryotic cells. In preferred embodiments, the HSPG dependent virus is selected from the group comprising HIV-1, HSV (HSV-1, HSV-2), HCMV, HPV, Respiratory syncytial virus (RSV) and filoviruses. In other embodiments of the invention, the virus is selected from the group comprising HIV-1, HSV (HSV-1, HSV-2), HCMV, HPV, Respiratory syncytial virus (RSV), influenza virus, and filoviruses.

Diseases associated with such viruses are selected from the group comprising respiratory viral diseases, gastrointestinal viral diseases, exanthematous viral disease, hepatic viral diseases, cutaneous viral diseases, hemorrhagic viral diseases, neurologic viral diseases. Typical diseases are, but not limited to, the common cold, flu, warts, viral herpes, hepatitis, rubella, smallpox, HIV/AIDS, Ebola, SARS-CoV-2/COVID-19, polio, viral meningitis, etc...

Another aspect of the invention provides a method of disinfection and/or sterilization of surfaces using one or more compounds of the invention or the virucidal composition of the invention or the pharmaceutical composition of the invention. The disinfection and/or sterilization is preferably done on living surfaces or non-living surfaces. The living surfaces are human or animal skin and/or hair. The non-living surface are, but not limited to, medical equipment, touch screens, textile, clothing, masks, gloves, furniture, and any other surfaces present in rooms, transport means, public spaces such as schools, airports, public transportation and cinemas. In some other embodiments, the non-living surfaces are fabric surfaces (masks, gloves, doctor coats, curtains, bed sheet), metal surfaces (lifts, door handle, nobs, railings, medical equipment and instruments, public transport and places), wood material surfaces (furniture, floors, partition panels), concrete surfaces (hospitals, clinics and isolation wards and walls), and plastic surfaces (medical equipment and instruments, touch screens, switches, kitchen and home appliances).

In a preferred embodiment, the method of disinfection and/or sterilization of surfaces comprises the steps of (i) providing at least one compound of the invention or a virucidal composition of the invention, or pharmaceutical composition of the invention, (ii) contacting a virus- contaminated surface or a surface suspected to be contaminated by a virus with the at least one compound of the invention or a virucidal composition of the invention or pharmaceutical composition of the invention for a time sufficient to obtain virucidal effect. In some embodiments, the virus-contaminated surface is human or animal skin and/or hair. In other embodiments, the virus-contaminated surface is a non-living surface. The non-living surface is, but not limited to, medical equipment, touch screens, textile, clothing, masks, gloves, furniture, and any other surfaces present in rooms, transport means, public spaces such as schools, airports, public transportation and cinemas. In some other embodiments, the non-living surfaces are fabric surfaces (masks, gloves, doctor coats, curtains, bed sheet), metal surfaces (lifts, door handle, nobs, railings, medical equipment and instruments, public transport and places), wood material surfaces (furniture, floors, partition panels), concrete surfaces (hospitals, clinics and isolation wards and walls), and plastic surfaces (medical equipment and instruments, touch screens, switches, kitchen and home appliances).

Another aspect of the invention provides a use of a compound of the invention or a virucidal composition of the invention or a pharmaceutical composition of the invention for sterilization and/or for disinfection. In some embodiments, sterilization and disinfection is for virus- contaminated surfaces or surfaces suspected to be contaminated by a virus. In some preferred embodiments, the surfaces are human or animal skin and/or hair. Thus in some embodiments, the invention provides a use of a compound of the invention or a virucidal composition of the invention or a pharmaceutical composition of the invention for sterilization and/or for disinfection of human or animal skin and/or hair. In other preferred embodiments, the surfaces are non-living surfaces. The non-living surfaces are, but not limited to, medical equipment, touch screens, textile, clothing, masks, gloves, furniture, and any other surfaces present in rooms, transport means, public spaces such as schools, airports, public transportation and cinemas. In some other embodiments, the non-living surfaces are fabric surfaces (masks, gloves, doctor coats, curtains, bed sheet), metal surfaces (lifts, door handle, nobs, railings, medical equipment and instruments, public transport and places), wood material surfaces (furniture, floors, partition panels), concrete surfaces (hospitals, clinics and isolation wards and walls), and plastic surfaces (medical equipment and instruments, touch screens, switches, kitchen and home appliances). In an embodiment, the virucidal composition of the invention or the pharmaceutical composition of the invention is used as virucidal hand disinfectant for frequent use. In another embodiment, the virucidal composition of the invention or the pharmaceutical composition of the invention is applied by spraying. In a further embodiment, the virucidal composition of the invention of the pharmaceutical composition of the invention is applied on a protective mask.

Another aspect of the invention provides a use of the compounds of the invention or the virucidal composition of the invention for manufacturing (producing) virucidal surfaces (i.e. able to inactivate viruses). Such surfaces are, but not limited to, textile, clothing, masks, touch screens, medical equipment, furniture. In some other embodiments, the surfaces are fabric surfaces (masks, gloves, doctor coats, curtains, bed sheet), metal surfaces (lifts, door handle, nobs, railings, medical equipment and instruments, public transport and places), wood material surfaces (furniture, floors, partition panels), concrete surfaces (hospitals, clinics and isolation wards and walls), and plastic surfaces (medical equipment and instruments, touch screens, switches, kitchen and home appliances). In some embodiments, the surfaces can be modified with the one or more compounds of the invention either through chemical modification or physical coating known in the art. Examples of physical coating are spraying or dipping the surface in a solution comprising the one or more compounds of the invention.

Another aspect of the invention provides a method for manufacturing (producing) a virucidal surface, wherein the method comprises coating the surface with the one or more compounds of the invention or the virucidal composition of the invention. The surface is, but not limited to, textile, clothing, masks, touch screens, medical equipment, furniture. In some other embodiments, the surface is fabric surface (masks, gloves, doctor coats, curtains, bed sheet), metal surface (lifts, door handle, nobs, railings, medical equipment and instruments, public transport and places), wood material surface (furniture, floors, partition panels), concrete surface (hospitals, clinics and isolation wards and walls), and plastic surface (medical equipment and instruments, touch screens, switches, kitchen and home appliances). The coating can be done either through chemical modification or physical coating known in the art.

Another aspect of the invention provides a virucidal surface coating composition comprising the one or more compounds of the invention or the virucidal composition of the invention. The virucidal surface coating composition of the invention can be sprayed or painted on surfaces. The surfaces are, but not limited to, medical equipment, touch screens, textile, clothing, masks, gloves, furniture, and any other surfaces present in rooms, transport means, public spaces such as schools, airports, public transportation and cinemas. In some other embodiments, the surfaces are fabric surfaces (masks, gloves, doctor coats, curtains, bed sheet), metal surfaces (lifts, door handle, nobs, railings, medical equipment and instruments, public transport and places), wood material surfaces (furniture, floors, partition panels), concrete surfaces (hospitals, clinics and isolation wards and walls), and plastic surfaces (medical equipment and instruments, touch screens, switches, kitchen and home appliances).

Another aspect of the invention provides a device comprising a surface coated with one or more compounds of the invention or with the virucidal composition of the invention. Such an antiviral coated device can be, but is not limited to, clothing, a mask, a glove, a touch screen, medical equipment, furniture, etc.... In one preferred embodiment, the device is a mask, clothing or medical equipment. In another preferred embodiment, the device is a medical device.

Testing

The weight, size, and degree of functionalization of the compounds of the invention can be determined, for example, by gel permeation chromatography (GPC) using size exclusion chromatography, by dynamic light scattering (DLS) and by nuclear magnetic resonance (NMR) measuring the intensity (integrals) ratio of hydrogens in the dPG core as compared to that of the hydrogens in the linker chain. Such characterization methodology is described, e.g., in Bhatia, et ah, "Linear polysialoside outperforms dendritic analogs for inhibition of influenza virus infection in vitro and in vivo", Biomaterials, Vol. 138, 22-34 (September 2017) ((DOI: 10.1039/c7py01470h) (https://doi.Org/10.1016/j.biomaterials.2017.05.028).

Degree of branching can be calculated using inverse gated (IG) ¹³ C NMR, for example, as described in Haag, R.; Sunder, A.; Stumbe, J.-F. J Am Chem Soc 2000, 122, 2954.

Cytotoxicity is determined by exposing Vero cells to varying concentrations of test drug and measuring the percentage of cells surviving such exposure. Antiviral activity is determined by plaque reduction assays on infected Vero cells, measuring the number of plaques that form in wells exposed to varying concentrations of test drug. Virucidal activity is determined by exposing infected Vero cells to an effective amount of test drug. After incubation, the test drug solution is removed and the cells are incubated again, measuring the plaques that form. The absense of plaques following removal of the test drug is an indication of virucidal activity. These determinations can be carried out, for example, as described in Cagno, et ah, "Broad- spectrum non-toxic antiviral nanoparticles with a virucidal inhibition mechanism," Nature Materials 17, 195-203 (2018).

Administration

The compounds of Formula I are administered at a therapeutically effective dosage, e.g., a dosage sufficient to provide treatment for the disease states previously described. Administration of the compounds of the invention or the pharmaceutically acceptable salts and/or esters thereof can be via any of the accepted modes of administration for agents that serve similar utilities, particularly intravenous, intraperitoneal, subcutaneous, topical and by inhalation. While human dosage levels have yet to be optimized for the compounds of the invention, generally, a daily dose is from about 0.001 to 2.0 mg/kg of body weight/day, preferably about 0.005 to 0.75 mg/kg of body weight/day, and most preferably about 0.01 to 0.5 mg/kg of body weight. Thus, for administration to a 70 kg person, the dosage range would be about 0.07 to 140 mg per day, preferably about 0.35 to 52.5 mg per day, and most preferably about 0.7 to 35 mg per day. Administration can be as a single daily dose or divided into 2 or more doses per day, over a period of treatment lasting from about 1 to about 7 days. The amount of active compound administered will, of course, be dependent on the subject and disease state being treated, the severity of the affliction, the manner and schedule of administration and the judgment of the prescribing physician.

Further aspects of the invention provide a method of treating and/or preventing virus infections and/or diseases associated with viruses, comprising administering to a subject in need thereof, a therapeutically effective amount of one or more compounds of the invention. In some embodiments of the method of treating, the one or more compounds of the invention are administered prophylactically.

Another aspect of the invention provides the compounds of the invention for use in treating and/or preventing virus infections and/or diseases associated with viruses. In some embodiments, the treating is a prophylactic treatment.

Formulation

The compounds of the invention that are used in the methods of the present invention can be incorporated into a variety of formulations and medicaments for therapeutic administration. More particularly, the compounds as provided herein can be formulated into pharmaceutical compositions by combination with appropriate, pharmaceutically acceptable carriers, excipients and/or diluents, and can be formulated into preparations in solid, semi-solid, liquid or gaseous forms, such as tablets, capsules, pills, powders, granules, dragees, gels, slurries, ointments, solutions, suppositories, injections, inhalants and aerosols. As such, administration of the compounds can be achieved in various ways, including oral, buccal, inhalation (pulmonary, nasal), rectal, parenteral, intraperitoneal, intradermal, topical, transdermal, intracranial and/or intratracheal administration. Moreover, the compounds can be administered in a local rather than systemic manner, e.g., in a topical cream or gel, a depot or a sustained release formulation. The compounds can be formulated with common excipients, diluents or carriers, and compressed into tablets, or formulated as elixirs or solutions for convenient oral administration, or administered by the intramuscular or intravenous routes. The compounds can be administered alone, in combination with each other, or they can be used in combination with other known compounds including oither antiviral agents. Suitable formulations for use in the present invention are found in Remington's Pharmaceutical Sciences (Mack Publishing Company (1985) Philadelphia, PA, 17th ed.), which is incorporated herein by reference. Moreover, for a brief review of methods for drug delivery, see, Langer, Science (1990) 249:1527-1533, which is incorporated herein by reference.

As to the appropriate excipients, carriers and diluents, reference may be made to the standard literature describing these, e.g. to chapter 25.2 of Vol. 5 of "Comprehensive Medicinal Chemistry", Pergamon Press 1990, and to "Lexikon der Hilfsstoffe fur Pharmazie, Kosmetik und angrenzende Gebiete", by H.P. Fiedler, Editio Cantor, 2002. The term "pharmaceutically acceptable carrier, excipient and/or diluent" means a carrier, excipient or diluent that is useful in preparing a pharmaceutical composition that is generally safe, and possesses acceptable toxicities. Acceptable carriers, excipients or diluents include those that are acceptable for veterinary use as well as human pharmaceutical use. A "pharmaceutically acceptable carrier, excipient and/or diluent" as used in the specification and claims includes both one and more than one such carrier, excipient and/or diluent.

Sustained-release preparations can be prepared. Suitable examples of sustained-release preparations include semi-permeable matrices of solid hydrophobic polymers containing the compounds of the invention, which matrices are in the form of shaped articles, e.g. films, or microcapsules. Examples of sustained-release matrices include polyesters, hydrogels (for example, poly(2-hydroxy ethyl-methacrylate), or poly(vinylalcohol)), polylactides (U.S. Pat. No. 3,773,919), copolymers of L-glutamic acid and [gamma] ethyl-L-glutamate, non- degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as the LUPRON DEPOT(TM) (injectable microspheres composed of lactic acid-glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3-hydroxybutyric acid.

The compounds of the invention can also be entrapped in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxymethylcellulose or gelatin-microcapsules and poly-(methylmethacylate) microcapsules, respectively, in colloidal drug delivery systems (for example, liposomes, albumin microspheres, microemulsions, nano-particles and nanocapsules) or in macroemulsions. Such techniques are disclosed in Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980).

The pharmaceutical compositions described herein can be manufactured in a manner that is known to those of skill in the art, i.e., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or lyophilizing processes. The following methods and excipients are merely exemplary and are in no way limiting. For injection, a compound of the invention (and optionally another active agent) can be formulated into preparations by dissolving, suspending, or emulsifying them in an aqueous or nonaqueous solvent, such as vegetable or other similar oils, synthetic aliphatic acid glycerides, esters of higher aliphatic acids or propylene glycol; and if desired, with conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifying agents, stabilizers, and preservatives. Preferably, the compounds of the invention can be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks' solution, Ringer's solution, or physiological saline buffer. Fortransmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.

Preferably, pharmaceutical formulations for parenteral administration include aqueous solutions of the compounds of the invention in water-soluble form. Additionally, suspensions of the compounds of the invention can be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions can contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension can also contain suitable stabilizers or agents that increase the solubility of the compounds of the invention to allow for the preparation of highly concentrated solutions.

The amount of a compound of the invention that can be combined with a carrier material to produce a single dosage form will vary depending upon the viral disease treated, the mammalian species, and the particular mode of administration. It will be also understood, that the specific dose level for any particular patient will depend on a variety of factors including the activity of the specific compound employed; the age, body weight, general health, sex and diet of the individual being treated; the time and route of administration; the rate of excretion; other drugs that have previously been administered; and the severity of the particular viral disease undergoing therapy, as is well understood by those of skill in the area.

Another aspect of the invention provides a virucidal composition comprising an effective amount of one or more compounds of the invention and optionally at least one suitable carrier or aerosol carrier. “An effective amount” refers to the amount sufficient for irreversibly inhibiting viruses; i.e. sufficient for obtaining virucidal effect. In an embodiment, the suitable carrier is selected from the group comprising stabilisers, fragrance, colorants, emulsifiers, thickeners, wetting agents, or mixtures thereof. In another embodiment, the virucidal composition can be in the form of a liquid, a gel, a foam, a spray or an emulsion. In a further embodiment, the virucidal composition can be an air freshener, a sterilizing solution or a disinfecting solution.

Another aspect of the invention provides a device (or a product) comprising the virucidal composition of the invention or one or more compounds of the invention and means for applying and/or dispensing thereof (i.e. the compounds of the invention or the virucidal composition). In another embodiment, the means comprise a dispenser, a spray applicator or a solid support soaked with the compounds of the invention. In another embodiment, the support is a woven or non-woven fabric, a textile, a paper towel, cotton wool, an absorbent polymer sheet, or a sponge.

Nasal solutions of the active compound alone or in combination with other pharmaceutically acceptable excipients can also be administered.

Formulations of the active compound or a salt may also be administered to the respiratory tract as an aerosol or solution for a nebulizer, or as a microfme powder for insufflation, alone or in combination with an inert carrier such as lactose. In such a case, the particles of the formulation have diameters of less than 50 microns, preferably less than 10 microns.

Those skilled in the art will appreciate that the invention described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the invention includes all such variations and modifications without departing from the spirit or essential characteristics thereof. The invention also includes all of the steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations or any two or more of said steps or features. The present disclosure is therefore to be considered as in all aspects illustrated and not restrictive, the scope of the invention being indicated by the appended claims, and all changes which come within the meaning and range of equivalency are intended to be embraced therein.

The foregoing description will be more fully understood with reference to the following Examples. Such Examples, are, however, exemplary of methods of practising the present invention and are not intended to limit the application and the scope of the invention.

EXAMPLES

Example 1

Description of the synthesis and functionalization of virucidal compounds

A. Dendritic polyglycerols synthesis (Cl)

A series of compounds have been synthesized through the chemical modification or functionalization of dendritic polyglycerols (dPGs). The dPG core was synthesized through slow monomer addition for ring opening multi-branching polymerization of glycidol (Figure 1). In the first step, trimethylolpropane (TMP) (25 mmol) was deprotonated by potassium methoxide solution (10 mmol KOH in 10 mL methanol). The resulting methanol was evaporated at 60 °C under vacuum (3 mbar). The synthesis-reactor was then heated to 100 °C and glycidol was added slowly over a period of 24h. The resulting dPG, having an average molecular weight of 10 kDa (“dPGio _kDa ” or “Cl”), was used for functionalization. Molecular weight was determined by GPC as shown in Figure 22.

By varying the amount of glycidol and reducing the period of addition, the corresponding dPG, having a molecular weight of 5 kDa ("dPG _5kDa ") was similarly obtained.

B. Synthesis of allyl-functionalized dPG (dPGio _kDa -allyl)

In order to prepare a soft material platform for the further thiol-ene click reaction, the free hydroxyl groups of dPGiokDa were converted to allyl groups through reaction with allyl bromide. The reaction was performed overnight in dry condition in presence of NaH as base for deprotonation of hydroxyl groups (Figure 2). The DMF was removed in vacuum and the functionalized polymer ( dPG _10kDa-allyl ) was purified by dialysis in MeOH for 2 days. The degree of functionalization was confirmed to be 100% by 'H NMR of the pure product correlating the allyl protons at 6.0 -5.1 ppm with the polyglycerol backbone protons (3.7 - 3.4) (Figure 3).

By limiting the equivalents of allyl bromide and NaH, a product, corresponding to the product in Figure 2 where the degree of functionalization for the allyl group was 2%, was obtained.

C. Synthesis of 11-mercapto-l-undecanesulfonate (MUS)

In preparation to functionalize the surface of dPG _10kDa-allyl , 11-mercapto-l-undecanesulfonate (MUS) was synthesized following a reported procedure (see, Figure 4).

Sodium undec-10-enesulfonate: 11-bromo-l-undecene (25 mL, 111.975 mmol), sodium sulfite (NaiSO ₃ ) (28.75 g, 227.92 mmol) and benzyltriethyl-ammonium bromide (10 mg) were added to a mixture of 200 mL methanol and 450 mL Di-water in a 1 L round bottom flask. The mixture was refluxed at 102°C for 48h, extracted with diethyl ether 5 times (5 x 400 ml), and the aqueous phase was evaporated in a rotary evaporator. The resulting white powder was dried under high vacuum, suspended in pure ethanol and filtered. The solution was evaporated, and the process was repeated twice to decrease the amount of inorganic salts. About 33 g of sodium undec-10- enesulfonate was collected as a white, methanol soluble powder.

Sodium 11-acetylthio-undecanesulfonate: Sodium undec-10-enesulfonate (33 g, 147.807 mmol) was dissolved in 500 ml of methanol. (The resulting solution should be clear in order to have high yield, and any precipitate should be removed by filtration.) A 2.6 times excess of thioacetic acid (27.324 mL, 384.3 mmol) was added to the solution and it was stirred in front of a UV lamp overnight (12h). The solution was evaporated in a rotary evaporator until the solid residue turned orange-red. The solid was washed with diethyl ether, until no colored material could be removed. The solid was dried under high vacuum, and then dissolved in methanol producing a yellow solution. About 3 g of carbon black was added to the solution, vigorously mixed, and the mixture was filtered through celite in a fluted filter paper. The filtered solution was clear, the solvent completely evaporated and about 35 g of sodium 11-acetylthio- undecanesulfonate was collected as a white solid. 11 -Mercapto- 1 -undecanesulfonate (MU S) : Sodium 11-acetylthio-undecanesulfonate (35 g, 120.7 mmol) was refluxed in 400 mL of 1M HC1 for 12 h, after which 200mL of 1M NaOH was added to the resulting solution, and an additional 400 mL of Di-water was added to create a 1 L volume. The resulting clear solution was kept at 4°C and crystallized overnight to yield a viscous white product that was centrifuged down in 50 mL falcon tubes, and dried under high vacuum. 12 g of methanol soluble MUS was collected from this purification step. (More material can be extracted from the supernatant of the centrifugation step, by reducing volume and keeping it at 4°C.) The successful synthesis of MUS was proved by ¹ HNMR and ESI-MS analysis (ESI-MS m/z 313.08 (M + Na) and m/z 601.17 (2M+Na) for dimer which is formed due to disulfide bond formation).

D. Synthesis of MUS functionalized dPGio koa (dPGio kDaMUS) (Effect of different functionalization with Cl 1-sulfonate) (2(85%), R17(48%), R18(2%))

As illustrated in Figure 6, the MUS moieties (obtained, for example, in Example 1C) were conjugated to the dPG _10kDa-allyl core (obtained, for example, in Example IB) through the thiol- ene click reaction to obtain different degrees of functionalization. For this aim dPG _10kDa-ally , _l MUS and 2,2-dimethoxy-2-phenylacetophenone (DMPA) as radical initiator were dissolved in a watenmethanol mixture. A catalytic amount of tris(2-carboxyethyl)phosphine hydrochloride (TCEP-HC1) was added to reduce the disulfide bonds and avoid oxidation of the thiol intermediate. The solution was degassed by flushing argon through the reaction mixture for 10 minutes. The reaction mixture was stirred and irradiated with UV light using a high-pressure UV lamp at room temperature for 6 hours. The solution was then dialyzed (MWCO 2 kDa) against watenmethanol for 2 days. The methanol was evaporated under reduced pressure and the samples were lyophilized to obtain the resulting products, _{dPGiokOa-MUSs85/»} (2), dPGiokDa- MUS48% (R17) and dPGiokDa-MUS2% (R 18), as white solids.

These three compounds with different degree of functionalization (85%, 48% and 2%) were synthesized, in the case of (2) and (R17) by varying the amount of MUS employed in the reaction, and in the case of (R18) by starting with a dPGC where degree of functionalization for the allyl group corresponding to R was 2% and the rest remained hydrogen (as discussed in the second paragraph of Example IB). The degree of functionalization was measured by ¹ HNMR (see, Figure 23) and elemental analysis. E. Synthesis of sulfate functionalized dPG (dPG-Cl 1-sulfate) (RX, R21)

In order to investigate the effect of sulfate functional groups on the virucidal activity, dPG was functionalized with undecanesulfate, as illustrated in Figure 8, starting with dPGs _kDa and dPGio _kOa . Each starting dPG (400 mg, 5.4 mmol of OH to be functionalized) was first reacted with 11-bromo-l-undecanol (2 g, 8.1 mmol, 1.5 eq.) in the presence ofNaH (259.17 mg, 10.8 mmol, 2 eq.) as the base for deprotonation of the dPGC's hydroxyl groups to obtain the corresponding dPG-Cn-OH with 50% of degree of functionalization. Each reaction mixture was allowed to stir for 24 hours at 40 °C and then was quenched by adding methanol and purified by dialysis against methanol. In the next step, both types of hydroxyl groups (the 50% that converted to hydroxy alkyl and the 50% that remained OH) were sulfated through the reaction with pyridine sulfur trioxide complex in dry DMF at 60 °C overnight. The reactions were quenched with water, and the pH adjusted to 8 by addition of NaOH solution. Solvent was evaporated under reduced pressure, and the products, dPG5kDa-Cn-sulfate4i%/sulfonate59% (overall weight 16 kDa) (R21) and dPGiokDa-Cn-sulfate5o%/sulfonate5o% (overall weight 30 kDa) (RX) were dissolved in brine. Dialysis was performed with aNaCl solution, using an ever- decreasing NaCl concentration, until the medium was changed with distilled water. Degree of functionalization was roughly 50% determined by ¹ HNMR of the pure products correlating the alkyl chain protons at 1.7 - 0.5 ppm with the polyglycerol backbone protons (4.4-3.1). Size was determined by DLS as 140 nm for RX and 100 nm for R21.

F. Synthesis of sulfate functionalized dPG with Sulfur Bridge (dPG-S-Cl 1-sulfate) (comparison of sulfate and sulfonate) (R19B)

In order to have a compound similar to R17 and compare the sulfate and sulfonate groups in term of virucidal activity, compound R19B was synthesized with similar degree of functionalization, as illustrated in Figure 10 (where R in the starting reactant is allyl). 11- Mercapto-l-undecanol (MUD) moieties were conjugated to a dPG-allyl core through the thiol- ene click reaction aiming for 50% functionalization. The reaction started by dissolving dPG- allyl (50 mg, 0.67 mmol of allyl group), MUD (80 mg, 0.39 mmol) and catalytic amount of 2,2- dimethoxy-2-phenylacetophenone (DMPA) as a radical initiator in a water: methanol mixture. A catalytic amount of tris(2-carboxyethyl)phosphine hydrochloride (TCEP-HC1) was added to avoid oxidation of the thiol intermediate. The solution was degassed by flushing argon through the reaction mixture for 10 minutes. The reaction mixture was stirred and irradiated with UV light using a high-pressure UV lamp at room temperature for 4 hours and then 1 -propanthiol was added to the mixture following by UV irradiation to quench the remaining allyl group ("R" in Figure 10). The solution was then dialyzed (MWCO 2 kDa) against watenmethanol for 2 days. The solvent was evaporated under reduced pressure to obtain a highly viscose white compound. In the next step, the hydroxyl group was sulfated through the reaction with pyridine sulfur trioxide complex in dry DMF at 60 °C overnight. The reaction was quenched with water, and the pH was adjusted to 8 by addition of NaOH solution. The solvent was evaporated under reduced pressure, and the product was dissolved in brine. Dialysis was performed with a NaCl solution, using an ever-decreasing NaCl concentration, until the medium was changed with distilled water. The final product, dPG _10kDa --C _al 3 _ly S _l Cn-sulfate5o _% /C ₃ SC ₃ 50% (R19B) was obtained as white solid compound after lyophilization ( ¹ HNMR spectrum in Figure 11).

G. Synthesis of sulfate and sulfonate functionalized dPG with short alkyl chains (C ₃ , C4) (RP3, RN4)

Figure 12 illustrates a one-step ring opening functionalization. The starting material, dPGio _kDa (100 mg, 1.35 mmol OH to be functionalized) was dried at 60 °C overnight under high vacuum. The dried dPG was dissolved in dry DMF (5 mL). To the stirred solution of dPG in dry DMF at room temperature, NaH (65 mg, 2.7 mmol, 2 eq.) was added. The reaction mixture was allowed to stir for 1 hour at room temperature. At this point, the syntheses diverge for production of the two products, proceeding in separate reaction vessels, to which 1,4-butane sultone (276 pL, 2.7 mmol, 2 eq.) (for RP3) or 1,3-propanediol cyclic sulfate (373.3 mg, 2.7 mmol, 2 eq.) (for RN4) were added and the reactions waere stirred overnight at room temperature. Each mixture was then dialysed against brine using an ever-decreasing NaCl concentration for 2 days, until the medium was changed with distilled water and dialysis continued for 2 more days. The solvent was decreased under reduced pressure and the respective products, dPGiokDa-C ₃ -sulfate2i% (RP3) and dPGiokDa-C4-sulfonate3i% (RN4) were obtained as crystalline powders after lyophilisation.

Example 2 Characterization

A. Gel permeation chromatography (GPC)

GPC measurements were performed using an Agilent 1100 solvent delivery system with a manual injector, isopump, and Agilent 1100 differential refractometer. A Brookhaven BIMwA7-angle light scattering detector was coupled to size exclusion chromatography (SEC) to measure the molecular weight of each fraction of the polymer that was eluted from the SEC columns. For the separation of the polymer samples, three 30 cm columns were used (10 pm PSS Suprema columns with pore sizes of 100 A, 1000 A, and 3000 A). Water was used as the mobile phase; the flow rate was set at 1.0 mL min ^-1 . All columns were held at room temperature. For each measurement, 100 pL of sample with a concentration of 5 mg mL ^-1 was injected. For acquisition of the data from seven scattering angles (detectors), a differential refractometer WinGPC Unity from PSS was used. Molecular weight distributions were determined by comparison with pullulan standards (10 different sizes from 342 to 710 000 g moU ¹ ). Water was used as a solvent with 0.1 M NaNCb.

B. Dynamic light scattering (DLS)

The size of the dendritic polyglycerols was measured in aqueous solution using a Zetasizer Nano ZS analyzer with an integrated 4 mW He-Ne laser at a wavelength of 633 nm with a backscattering detector angle of 173° (Malvern Instruments Ltd, UK) at 25 °C. For DLS experiments, an aqueous solution of dPG with different concentrations was prepared in Milli- Q water and vigorously stirred for 18 hours at room temperature (25 °C). Solutions were filtered via 0.45 pm polytetrafluoroethylene (PTFE) filters and used for dynamic light scattering measurements. Disposable UV-transparent cuvettes (Sarstedt AG & Co, Germany) were used for all the experiments.

C. Nuclear magnetic resonance (NMR)

NMR spectra were recorded on a Jeol ECX 400 or a Jeol Eclipse 700 MHz spectrometer. Proton NMR spectra were recorded at 295 K in ppm and were referenced to the indicated solvents.

Example 3

Other Compounds of Formula I

A. By following the procedures of Example 1 A and adjusting the equivalents of glycidol and reaction time accordingly, there are obtained dPG5 _kDa , dPG _25kDa , dPGsokDa, dPG _75kDa , and dPG _100kDa .

B. By following the procedures of Example IB and substituting allyl bromide with: a) 4-bromobut-l-ene, b) 5-bromopent-l-ene, c) 7-bromo-5-methylhept-l-ene, and d) 10-iodo-5-isopropyl-6-methyldec-l-ene; there are obtained the following compounds, respectively: a) dPGiokDa-but-l-eneioo%, b) dPGiokDa-pent-l-eneioo%, c) dPGiokDa-5-methylhept-l-eneioo%, and d) dPGiokDa-5-isopropyldec-l-eneioo%.

C. By following the procedures of Example 1C and substituting 11-bromo-l-undecene with: a) 8-bromooct-l-ene, b) 1 l-bromo-5,4-dimethylundec-l-ene, c) 14-bromotetradec-l-ene, d) 8-ethyl- 16-iodo-5-methylhexadec-l-ene, and e) 26-bromohexacos-l-ene; there are obtained the following compounds, respectively: a) sodium 8-mercaptooctane-l -sulfonate, b) sodium 1 l-mercapto7,8-dimethylundecane-l -sulfonate, c) sodium 14 mercaptotetradecane-1 -sulfonate, d) sodium 9-ethyl- 16-mercapto-12-methylhexadecand-l -sulfonate, and e) sodium 26-mercaptohexacosand-l -sulfonate.

D. By following the procedures of Example ID and substituting sodium 11-mercapto-l- undecane sulfonate with: a) sodium 8-mercaptooctane-l -sulfonate, b) sodium 1 l-mercapto7,8-dimethylundecane-l -sulfonate, c) sodium 14 mercaptotetradecane-1 -sulfonate, d) sodium 9-ethyl- 16-mercapto-12-methylhexadecane-l -sulfonate, and e) sodium 26-mercaptohexacosand-l -sulfonate; there are obtained the following compounds, respectively: a) dPG10kDa-C ₃ -S-C ₈ SO ₃ -Na ⁺ 85%, b) dPGiokDa-C ₃ -S-C ₁₁ -7,8-di-Me-SO ₃ ^" Na ⁺ 85%, c) dPGl0kDa-C ₃ -S-C ₁₁ SO ₃ -Na ⁺ 48%, d) dPGiokDa-C ₃ -S-C ₁₆ -9-Et-12-Me-SO ₃ ^' Na ⁺ 48%/allyl52%, and e) dPGiokDa-C ₃ -S-C ₂₆ SO ₃ ^' Na ⁺ 85%/allyli5%.

E. By following the procedures of Example IE and substituting 11-bromo-l-undecanol with: a) 5-bromopentan-l-ol, b) 7-bromoheptan-l-ol, c) 7-iodo-2-methylheptan-l-ol, d) 8-bromooctan-l-ol, e) 9-bromononan-l-ol, f) 10-bromodecan-l-ol, g) 1 l-bromo-4,5-dimethylundecan-l-ol, h) 12-chlorododecan-l-ol, i) 13-bromotridecan-l-ol, j) 14-bromotetradecan-l-ol, k) 15-bromopentadecan-l-ol, l) 18-iodooctadecan-l-ol, and m) 31-bromohentriacontan-l-ol; there are obtained the following compounds, respectively: a) dPGiokDa-C ₅ -SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, b) dPGiokDa-C ₇ -SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, c) dPGiokDa-C ₇ -2-Me-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, d) dPGiokDa-C ₈ -SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, e) dPGiokDa-C ₉ -SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, f) dPGiokDa-C ₁₀ -SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, g) dPGiokDa-Cn-4,5-di-Me-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, h) dPGiokDa-Ci2-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, i) dPGiokDa-Ci3-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, j) dPGiokDa-Ci4-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, k) dPGiokDa-Ci5-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, l) dPGiokDa-Ci8-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%, and m) dPGiokDa-C ₃ i-SO ₄ ^' Na ⁺ 5o%/SO ₃ ^' Na ⁺ 5o%.

F. By following the procedures of Example IF and substituting 11-mercapto-l-undecanol with: a) 8-mercaptooctan-l-ol, b) 10-mercaptodecan-l-ol, c) 1-hydroxy-l l-mercaptoundecyl sulfate, d) 12-mercaptododecan-l-ol, and e) 13-mercaptotridecan-l-ol; there are obtained the following compounds, respectively: a) dPGiokD-C ₃ SC ₈ -SO ₄ ^' Na ⁺ 5o%/C ₃ SC ₃ 50%, b) dPGiokDa-C ₃ SC ₁₀ -S0 ₄ -Na ⁺ ₅₀ %/C ₃ SC ₃ 50%, c) dPGiokDa-C ₃ SC ₁₁ -di-SO ₄ ^' Na ⁺ 5o%/C ₃ SC ₃ 50%, d) dPGiokDa-C ₃ SC ₁₂ -SO ₄ -Na ⁺ ₅₀ %/C ₃ SC ₃ 50%, and e) dPGiokDa-C ₃ SC ₁₃ -SO ₄ -Na ⁺ ₅₀ %/C ₃ SC ₃ 50%.

Example 4 Testing

A. METHODS

1. Toxicity assay on Vero Cells

Cytotoxicity of the dendritic polyglycerol was tested on mammalian cells. Vero cells were plated 24h before the experiment in 96-well plates in order to have a confluent layer. Cells were then incubated with different concentrations of the compound being tested at 37°C for 24 h in DMEM w2% FBS. The solution was then removed and the cells washed with DMEM w2%FBS. 100 ul of DMEM w2% FBS were added in each well with 20 ul of MTS (CellTiter 96® AQueous One Solution Cell Proliferation Assay). After 4 hours of incubation at 37°C, the absorbance of each well was measured through a plate-reader (1=490 nm).

A percentage of cytotoxicity was then calculated comparing the absorbance with a reference, in which cells were incubated with just DMEM w2% FBS.

2. Inhibition assay against HSV-2

The virucidal effect of dendritic polyglycerol against HSV-2 was tested by plaque reduction assays on Vero cells. Vero cells were plated 24h before the experiment in 24-well plates at a density of 10 ⁵ cells. A fixed amount of virus (MOI = 0.0005) was pre-incubated for 1 hour with serial dilutions of the compound of interest. The solution was then transferred onto the cells and incubated for 1 hour. Afterwards, the solution was removed and the cells incubated for 24h in DMEM w2% FBS with 0.45w% Methyl-Cellulose. The cells were then stained with crystal violet and the plaques counted.

3. Virucidal assay against HSV-2

The virucidal activity of the dendritic polyglycerol against HSV-2 was tested by virucidal assay. Vero cells were plated 24h before the experiment in 96-well plates in order to have a confluent layer. An effective amount of dendritic polyglycerol (100 mg/ml) were incubated with a fixed amount of viruses (10 ⁵ -10 ⁶ pfu/ml) for 1 hour at 37°C in DMEM - 2%FBS.

A serial dilution of this solution was added in each well and incubated for 1 hour at 37°C. Afterwards, the solution was removed and the cells incubated for 24 h in DMEM w2% FBS with 0.45w% Methyl-Cellulose. The cells were then stained with crystal violet and the plaques counted. The viral titer was evaluated and compared with a reference with no compound.

4. Data Analysis

The EC ₅ 0 values for inhibition curves (dose-response assay) were calculated using GraphPad Prism 8.0 using a 4-parameter. B. RESULTS

Here, we report the results from testing the above-reported dendritic polyglycerols against HSV-2 in Vero Cells.

1. Sulfonated dendritic polyglycerol (2, R17, R18)

Sulfonated dendritic polyglycerols with different degrees of functionalization were tested against HSV-2. As shown in Figure 14, the bare dendritic polyglycerol (Cl) did not show any inhibitory activity in the range tested, as well as the dendritic polyglycerol having a low functionalization (R18, 2%, namely 4 groups per dendritic polyglycerol). Functionalization of at least 50% (2) and (R17) gave inhibitory activity with an IC ₅₀ in the order of hundreds of nanomolar (-300-550 nM). Both the compounds having a DF of at least 50% show virucidal activity , as can be seen in Figure 15, where a decrease of the viral titer of more than 2 orders of magnitude is shown for both the compounds.

2. Alkyl chain length - (RP3, RN4)

Dendritic polyglycerols with shorter alkyl chain (C ₃ and C4), bearing either sulfonate (RP3) or sulfate (RN4) functionality show limited efficacy against HSV-2 as reported in Figure 16. Indeed, neither of them show virucidal activity. This result confirms the importance of the length of the linker to obtain a strong interaction between the dPG and the virus.

3. Size and weight (RX, R21)

The dendritic polyglycerols RS and R21 were tested for inhibition of HSV-2 as described in Example 4A2 and for virucidal activity as described in Example 4A3. This evidences the impact of molecular weight (and size) on activity. The results are shown in Figure 17. The results for RX are also shown in Figure 18.

Both of the compounds showed excellent virucidal inhibitory activity against HSV-2, having IC ₅ 0 in the nanomolar range. Both compounds are virucidal. RX, a slightly larger version, proved to be more effective than the smaller one (R21), having an IC _{5 0} of 4.5 nM.

4. Sulfate / Sulfonate (RX, R19B)

The compound R19B was tested to evaluate the activity of sulfated dPGs. The results are reported in Figure 19. The results show an extremely low IC _{5 0} , but not as low as RX (as shown in the table). The difference in overall zeta-potential may correlate with the higher inhibition activity of RX as compared to R19B.

5. Comparison with CD and AuNPs Gold nanoparticles covered with a binary shell of MUS:OT and modified cyclodextrins bearing MUS have recently been reported as having broad-spectrum antiviral activity with no toxicity and virucidal activity, but these approaches have their own limitations. The main limitation of the gold nanoparticles is that the presence of the gold core raises concerns about bio accumulation. On the other side, the gold nanoparticles have an antiviral activity in the nanomolar range. Conversely, modified cyclodextrins are based on an FDA-approved organic core that overcomes the bio-accumulation issues of the gold nanoparticles, but the modified cyclodextrin's activity is much lower (almost 3 order of magnitude), being in the micromolar range. RX has been compared against these two other compounds. Figure 20 shows the results of testing inhibition activity of the three compounds, e.g., as described in Example 4A2. The data, reported in Table 1, show that RX is superior to both compounds in terms of IC ₅₀ (both in molarity and in mass concentration), meaning that RX has a stronger affinity. Thus, RX has an equivalent effect on HSV-2 as compared to AuNPs and MUS-CD, respectively, using 1/10 and 1/200 of the material. 6. Summary

Table 2 summarizes the characteristics and activities of the compounds made and tested as reported above. Example 5

Lip BalmFormulation As disclosed in US 2013/0150312, polyethylene 1450 and 300 are melted at 50° C with stirring. The dPGRX, 2-deoxy-D-glucose, silica gel and stevioside are triturated together. The triturated powders are slowly sifted into the melted PEGs with stirring. The flavoring is added, followed by thorough mixing. The mixture is poured into applicator tubes and allowed to cool to room temperature.

Example 6

Aqueous Cream Formulation As disclosed in US 2013/0150312, apart of the dPGRX is dissolved in water with the 2-deoxy-

D-glucose and propylene glycol at ambient temperature to produce an aqueous solution. The paraffins and emulsifiers (cetostearyl alcohol and sodium lauryl sulphate) are mixed together, heated to 60° C, and emulsified with the aqueous solution, also at 60° C. The remaining dPG RX is added, the mixture dispersed, allowed to cool, and filled into lacquered aluminum tubes.

Example 7

Nebulizer Formulation

A nebuliser, for example, as disclosed in US 9,364,618 B2 or EP 3,517,117 A1 is provided comprising the following pharmaceutical composition in its fluid reservoir: 14.0 mg dPG RX, 0.9% w/v NaCl dissolved in sterile deionised water. The nebuliser is used to deliver the composition by inhalation as an aerosol to the lower respiratory tract of a patient suffering from influenza. While the present invention has been described with reference to the specific embodiments thereof, it should be understood by those skilled in the art that various changes may be made and equivalents may be substituted without departing from the true spirit and scope of the invention. In addition, many modifications may be made to adapt a particular situation, material, composition of matter, process, process step or steps, to the objective, spirit and scope of the present invention. All such modifications are intended to be within the scope of the claims appended hereto. All patents and publications cited above are hereby incorporated by reference.