RU2243236C2 - Linear copolymers of cyclodextrin - Google Patents

Abstract

FIELD: chemistry of polymers, medicine, pharmacy.

SUBSTANCE: invention relates to linear copolymers of cyclodextrin and oxidized cyclodextrin that can be used as carriers in delivery of different therapeutic agents. Water-soluble linear copolymer of cyclodextrin is prepared by a method that involves: a) preparing at least one precursor of cyclodextrin monomer allowing carrying out the polymerization reaction at two positions only; b) interaction of precursor of cyclodextrin monomer with compound comprising two functional groups through that the binding precursors of cyclodextrin monomer can be carried out and resulting to preparing water-soluble linear cyclodextrin copolymer comprising repeated elementary links of the formula

. The cyclodextrin copolymer comprising cyclodextrin fragments as a component of the main chain of polymer and comprising repeated elementary links of the formula (Ia) and (Ib). The therapeutic composition comprises above said copolymer and effective dose of the therapeutic agent. The composition comprises: a) the first copolymer wherein at least one cyclodextrin fragment or monomer is oxidized, and b) the second cyclodextrin copolymer comprising cyclodextrin fragments as component of the main polymeric chain. The therapeutic composition comprises above said cyclodextrin composition and effective dose of therapeutic agent. Method for delivery of therapeutic agent to patient comprises administration to patient therapeutically effective dose of above said therapeutic compositions. Invention provides preparing copolymers that can be used as a carrier in delivery of medicinal agent and can release this agent in response to specific biological processes of catabolism.

EFFECT: improved preparing method, valuable medicinal and pharmaceutical properties of copolymers.

50 cl, 33 ex

Description

Field of Invention

The invention relates to linear cyclodextrin copolymers and linear oxidized cyclodextrin copolymers. These copolymers respectively contain a cyclodextrin fragment, unoxidized or oxidized, as a monomer unit that is embedded in the main chain of the copolymer. The invention also relates to methods for producing linear copolymers of cyclodextrin and linear copolymers of oxidized cyclodextrin. Such cyclodextrin copolymers can be used as a delivery vehicle for various therapeutic agents.

Description of the prior art

Cyclodextrins are cyclic polysaccharides containing elementary units of the natural D (+) glucopyranose in the α- (1,4) compound. The most common cyclodextrins are alpha (α) cyclodextrins, beta (β) cyclodextrins, and gamma (γ) cyclodextrins, which contain six, seven, or eight glucopyranose units, respectively. The cyclic structure of cyclodextrin is similar in shape to a torus or toroidal ring having an internal non-polar or hydrophobic cavity, and secondary hydroxyl groups are located on one side of the cyclodextrin torus and primary hydroxyl groups are located on the other side. Thus, using (β) -cyclodextrin as a sample, cyclodextrin is often represented schematically as follows:

The side on which the secondary hydroxyl groups are located has a larger diameter than the side on which the primary hydroxyl groups are located. The hydrophobic nature of the internal cavity of cyclodextrin makes it possible to include various compounds in it. (Comprehensive Supramolecular Chemistry, Volume 3, JL Atwood et al., Eds., Pergamon Press (1996); T. Cserhati, Analytical Biochemistry, 225: 328-332 (1995); Husain et al., Applied Spectroscopy, 46: 652 -658 (1992); FR 2665169).

Cyclodextrins have previously been used as a delivery vehicle for various therapeutic compounds through the formation of inclusion complexes with various drugs that can fit into the hydrophobic cavity of the cyclodextrin, or through the formation of non-covalent associated complexes with other biologically active molecules, such as oligonucleotides and their derivatives. For example, US Pat. No. 4,727,064 describes pharmaceutical preparations consisting of a drug with a substantially low solubility in water and an amorphous, water-soluble cyclodextrin-based mixture. The drug forms an inclusion complex with cyclodextrins of the mixture. US Pat. No. 5,691,316 describes a cyclodextrin system for delivering oligonucleotides to cells. In such a system, an oligonucleotide is non-covalently linked to a cyclodextrin in a complex, or, alternatively, an oligonucleotide can be covalently linked to adamantine, which in turn is non-covalently associated with cyclodextrin.

Various polymers containing cyclodextrin and methods for their preparation are also known. (Comprehensive Supramolecular Chemistry, Volume 3, J. L. Atwood et al., Eds., Pergamon Press (1996)). A method for producing a polymer containing immobilized cyclodextrin is described in US Pat. No. 5,608,015. According to this method, a cyclodextrin derivative is reacted with α, β-unsaturated acid monomeric acid chloride or its derivative, or with α, β-unsaturated acid or its derivative containing isocyanate at the end a group or derivative of an isocyanate group. A cyclodextrin derivative is prepared by reacting cyclodextrin with compounds such as carboxylic acid halides and acid anhydrides. The resulting polymer contains cyclodextrin elementary units in the form of side chains extending from the main chain of the linear polymer macromolecule.

US 5,276,088 describes a process for synthesizing polymers containing cyclodextrin by reacting polyvinyl alcohol or cellulose or its derivatives with cyclodextrin derivatives or by co-polymerizing a cyclodextrin derivative with vinyl acetate or methyl methacrylate. Again, the resulting polymer contains a cyclodextrin fragment in the form of a side fragment extending from the polymer backbone.

Biologically degradable polymer aggregates with a supramolecular structure are described in WO 96/09073 A1. The unit includes a number of cyclic compounds that carry the drug, which are obtained by binding the drug to α-, β- or γ-cyclodextrin and then “stringing” the compounds of the drug / cyclodextrin composition along a linear polymer chain containing biodegradable fragments attached to both its ends. Such an aggregate has been reported to be able to release a drug in response to a specific biological degradation occurring in a disease. These aggregates are commonly referred to as “necklace” cyclodextrin polymers.

However, there is still a need for linear polycyclodextrins in which the cyclodextrin fragment is part of the main chain and not the side fragment emanating from the main chain, and a process for their preparation.

SUMMARY OF THE INVENTION

The present invention satisfies this need and provides linear cyclodextrin copolymers. Such linear cyclodextrin copolymers contain a repeating unit of formulas Ia, Ib, or a combination thereof:

The invention also provides methods for preparing linear cyclodextrin copolymers. One such method involves co-polymerizing a starting cyclodextrin monomer disubstituted with the same or different leaving groups and a starting comonomer A capable of replacing a leaving group. Another such method involves the iodination of the starting cyclodextrin monomer to obtain the starting diiodated cyclodextrin monomer and subsequent co-polymerization of the starting diiodated cyclodextrin monomer with the starting comonomer A to produce a linear cyclodextrin copolymer. Another method involves the iodination of the starting cyclodextrin monomer to form the starting diiodinated cyclodextrin monomer, the amination of the starting diiodinated monomer to produce the starting diaminated cyclodextrin monomer and the subsequent co-polymerization of the starting diaminated cyclodextrin monomer with the starting comonomer A to form a linear copolymer of cyclodextrin. Another method involves reducing a linear oxidized cyclodextrin copolymer to a linear cyclodextrin copolymer.

The invention also provides a linear oxidized cyclodextrin copolymer. A linear oxidized cyclodextrin copolymer is a linear cyclodextrin copolymer that includes at least one oxidized cyclodextrin fragment of formula VIa or VIb:

Each cyclodextrin fragment of the linear cyclodextrin copolymer of the invention can be oxidized so as to produce a linear oxidized cyclodextrin copolymer containing a repeating unit of formula VIa, VIb, or a combination thereof.

The invention also provides a method for producing a linear oxidized cyclodextrin copolymer. One such method involves oxidizing a linear cyclodextrin copolymer such that at least one cyclodextrin monomer is oxidized. Other methods include co-polymerizing the starting oxidized cyclodextrin monomer with the starting comonomer A.

The invention further provides a linear copolymer of cyclodextrin or a linear copolymer of oxidized cyclodextrin grafted onto a substrate, and a method for producing it. The invention also provides a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer crosslinked with another polymer, and a method for producing it. A method for producing crosslinked cyclodextrin polymers involves reacting a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer with a polymer in the presence of a crosslinking agent.

The invention provides a linear cyclodextrin copolymer or a linear oxidized dextrin copolymer containing at least one ligand bound to a cyclodextrin copolymer. The ligand can be associated either with a cyclodextrin fragment or with a fragment of comonomer A.

The invention also provides a cyclodextrin composition comprising at least one linear cyclodextrin copolymer of the present invention and at least one linear oxidized cyclodextrin copolymer of the present invention. The invention also provides therapeutic compositions comprising a therapeutic agent and a linear cyclodextrin copolymer and / or the linear oxidized cyclodextrin copolymer of the present invention. A method of treatment by administering a therapeutically effective amount of a therapeutic composition of the invention is also described.

DETAILED DESCRIPTION OF THE INVENTION

One embodiment of the invention is a linear cyclodextrin copolymer. A linear cyclodextrin copolymer is a polymer containing cyclodextrin fragments as part of the backbone of a polymer macromolecule. Previously, cyclodextrin fragments were not parts of the main chain of the polymer macromolecule, but departed from the main chain of the polymer macromolecule as side fragments. According to this invention, a linear cyclodextrin copolymer contains a repeating unit of formula Ia, Ib, or a combination thereof:

In the formulas Ia and Ib, C represents a substituted or unsubstituted cyclodextrin monomer, A represents a comonomer linked, for example, covalently linked to cyclodextrin C. The polymerization of the starting cyclodextrin monomer C with the starting comonomer A leads to a linear cyclodextrin copolymer of the present invention. Within one linear cyclodextrin copolymer of the present invention, the unit of cyclodextrin monomer C may be the same or different, and similarly, comonomer A may be the same or different.

The cyclodextrin monomer precursor may be any cyclodextrin or derivative thereof known in the art. As described above, cyclodextrin is defined as a cyclic polysaccharide, in most cases containing from six to eight elementary units of the naturally occurring D (+) - glucopyranose in the α- (1,4) bond. Preferably, the cyclodextrin monomer precursor is cyclodextrin containing six, seven and eight glucose units, i.e., alpha (α) -cyclodextrin, beta (β) -cyclodextrin and gamma (γ) -cyclodextrin, respectively. The cyclodextrin derivative may be any known substituted cyclodextrin in which the substituent does not affect co-polymerization with the comonomer A precursor, as described above. According to the invention, the cyclodextrin derivative may be neutral, cationic or anionic. Examples of suitable substituents include, but are not limited to hydroxyalkyl groups, such as, for example, hydroxypropyl, hydroxyethyl; ether groups such as, for example, dihydroxypropyl ether, methylhydroxyethyl ether, ethylhydroxyethyl ether and ethylhydroxypropyl ether groups; alkyl groups, such as, for example, methyl; saccharides, such as, for example, glucosyl and maltosyl; acid groups, such as, for example, carboxylic acid, phosphoric acid, phosphinic acid, phosphonic acid, phosphoric acid, thiophosphonic acid, thiophosphonic acid and sulfonic acid groups; imidazole groups and sulfate groups.

In addition, the cyclodextrin monomer precursor may be chemically modified (e.g., halogenated, aminated) to promote or influence the polymerization of the cyclodextrin monomer precursor with the comonomer A precursor, as described below. Chemical modification of the cyclodextrin monomer precursor allows polymerization in only two positions on each cyclodextrin fragment, that is, to obtain a bifunctional cyclodextrin fragment. The C1-C6 numbering scheme of the positions of each glucopyranose ring is as follows:

In a preferred embodiment, the polymerization is carried out at any two of the C2, C3 or C6 positions of the cyclodextrin fragment, including combinations thereof. For example, one cyclodextrin monomer precursor can polymerize at two C6 positions, while the other cyclodextrin monomer precursor can polymerize at positions C2 and C6 of the cyclodextrin fragment. In the case of β-cyclodextrin, the recording scheme for the relative position of each glucopyranose ring in cyclodextrin will be as follows:

In a preferred embodiment of the linear cyclodextrin copolymer of the present invention, the cyclodextrin monomer C has the following general formula (II):

In formula (II), n and m are integers that, in total with two glucopyranose rings, determine the total number of glucopyranose unit units in the cyclodextrin monomer. Formula (II) represents a cyclodextrin monomer which is capable of undergoing polymerization at two C6 positions on a cyclodextrin unit. Examples of cyclodextrin monomers of formula (II) include, but are not limited to, 6 ^A , 6 ^B- deoxy-α-cyclodextrin (n = 0, m = 4), 6 ^A , 6 ^C- deoxy-α-cyclodextrin (n = 1, m = 3), 6 ^A , 6 ^D- deoxy-α-cyclodextrin (n = 2, m = 2), 6 ^A , 6 ^B- deoxy-β-cyclodextrin (n = 0, m = 5), 6 ^A , 6 ^C- deoxy-β-cyclodextrin (n = 1, m = 4), 6 ^A , 6 ^D- deoxy-β-cyclodextrin (n = 2, m = 3), 6 ^A , 6 ^B- deoxy-γ-cyclodextrin (n = 0, m = 6), 6 ^A , 6 ^C- deoxy-γ-cyclodextrin (n = 1, m = 5), 6 ^A , 6 ^D- deoxy-γ-cyclodextrin (n = 2, m = 4 ), 6 ^A , 6 ^E- deoxy-γ-cyclodextrin (n = 3, m = 3). In another preferred embodiment of the linear cyclodextrin copolymer of the present invention, the cyclodextrin unit of monomer C has the following general formula (III):

where p = 5-7. In the formula (III), one of the D (+) elementary units, the glucopyranose of the cyclodextrin monomer, undergoes a ring opening to polymerize to the C2 and C3 positions of the cyclodextrin element. Cyclodextrin monomers of formula (III) are commercially available from Carbomer of Westborough, MA. Examples of cyclodextrin monomers of formula (III) include, but are not limited to, 2 ^A , 3 ^A- deoxy-2 ^A , 3 ^A- dihydro-α-cyclodextrin, 2 ^A , 3 ^A- deoxy-2 ^A , 3 ^A- dihydro-β -cyclodextrin, 2 ^A , 3 ^A -deoxy-2 ^A , 3 ^A -dihydro-γ-cyclodextrin, which are commonly called 2,3-deoxy-α-cyclodextrin, 2,3-deoxy-β-cyclodextrin and 2,3, respectively -deoxy-γ-cyclodextrin.

The comonomer precursor A may be any straight or branched chain compound, symmetric or asymmetric, which, as described above, combines two cyclodextrin monomers by reaction with the cyclodextrin monomer precursor. Preferably, the comonomer A precursor is a compound containing at least two functional groups through which the reaction and, therefore, the binding of cyclodextrin monomers can be carried out. Examples of possible functional groups, which may be the same or different, located at the end or middle of the molecule of each starting comonomer A, include, but are not limited to, an amino group, an acid group, an ester group, an imidazole group or an acyl halide group and their derivatives. In a preferred embodiment, the two functional groups are the same and terminal. By copolymerizing the comonomer precursor A with the cyclodextrin monomer precursor, two cyclodextrin monomers can bind together by combining the primary hydroxyl groups of one cyclodextrin monomer, by combining the secondary hydroxyl groups of one cyclodextrin monomer or another with a second primary hydras ksilnyh groups of one cyclodextrin monomer with the secondary groups giroksilnymi another cyclodextrin monomer. Accordingly, combinations of such chemical bonds may exist in the final copolymer.

Both the precursor of comonomer A and comonomer A of the final copolymer can be neutral, cationic (for example, containing protonated groups, such as, for example, Quaternary ammonium groups) or anionic (for example, containing deprotonated groups, such as, for example, sulfate, phosphate or carboxylate anionic groups). The charge of the comonomer A of the copolymer can be controlled by adjusting the pH. Examples of suitable comonomer A precursors include, but are not limited to, cystamine, 1,6-diaminohexane, diimidazole, dithioimidazole, spermine, dithiospermine, digistidine, dithiohistidine, succinimide (e.g., dithiobis (succinimidyl propionate) (DSP) and disuccide) (e.g. dimethyl 3,3'-dithiobispropionimidate (DTBP)). Copolymerization of the comonomer A precursor with the cyclodextrin monomer precursor results in a linear cyclodextrin copolymer of the present invention containing bonds of comonomer A of the following general formulas:

-HNC (O) (CH ₂ ) _x C (O) NH-, -HNC (O) (CH ₂ ) _x SS (CH ₂ ) _x C (O) NH-,

- ⁺ H ₂ N (CH ₂ ) _x SS (CH ₂ ) _x NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ , -HNC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NH-,

-HNNHC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NHNH-,

- ⁺ H ₂ NCH ₂ (CH ₂ CH ₂ O) _x CH ₂ CH ₂ CH ₂ NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ -,

-NC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ SS (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NH-,

-HNC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NH-,

-SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-,

-SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ ) _x SS (CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-,

-SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) CH ₂ CH ₂ (OCH ₂ CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ CH ₂ S-

In the above formulas, x = 1-50 and y + z = x. Preferably x = 1-30. More preferably x = 1-20. In a preferred embodiment, comonomer A is biodegradable or acid labile. In addition, in a preferred embodiment, the starting comonomer A and, therefore, comonomer A can be selectively selected for the intended use. For example, for the delivery of therapeutic agents with small molecules, the presence of a charge in the polymer may not be necessary, and comonomer A may be a polyethylene glycol group.

The linear cyclodextrin copolymer of the present invention may be modified with at least one ligand attached to the cyclodextrin copolymer. The ligand can be attached to the cyclodextrin copolymer via cyclodextrin monomer C or comonomer A. Preferably, the ligand is attached to at least one cyclodextrin fragment of the linear cyclodextrin copolymer. Preferably, the ligand allows the linear cyclodextrin copolymer to reach and bind to the cell. If several ligands are attached to the linear cyclodextrin copolymer of the invention, which may be the same or different, the additional ligand or ligands can be attached to the same or different cyclodextrin fragments or to the same or different comonomer A of the copolymer.

Examples of suitable ligands include, but are not limited to, vitamins (e.g., folic acid), proteins (e.g., transferrin and monoclonal antibodies) and polysaccharides. The ligand will vary depending on the type of target delivery. For example, receptor-mediated delivery can be achieved by, but not limited to, the use of a folic acid ligand, while delivery of antisense oligonucleotides can be achieved by, but not limited to, a transferrin ligand. The ligand can be attached to the copolymer of the invention by known methods.

Another embodiment of the invention is a method for producing a linear cyclodextrin copolymer. According to the invention, the linear cyclodextrin copolymer of the invention can be prepared by co-polymerizing a cyclodextrin monomer precursor disubstituted with suitable leaving groups with a comonomer A precursor which is capable of replacing leaving groups. The leaving groups, which may be the same or different, can be any leaving group known in the art that can be substituted during co-polymerization with the precursor of comonomer A. In a preferred embodiment, a linear cyclodextrin copolymer can be prepared by iodinating a cyclodextrin monomer precursor to obtain diiodinated cyclodextrin monomer precursor and co-polymerization of diiodated cyclodextrin precursor Vågå monomer with a comonomer A precursor to obtain a linear cyclodextrin copolymer having repeating unit of formula Ia, Ib or a combination thereof, as described above. In a preferred embodiment of the process for preparing the linear cyclodextrin copolymer of the present invention, the iodination of the cyclodextrin monomer precursor as described above is carried out to obtain a diiodinated cyclodextrin monomer precursor of formula IVa, IVb, IVc, or a mixture thereof:

Diiodinated cyclodextrin can be obtained by any known method. (Tabushi et al., J. Am. Chem. 106, 5267-5270 (1984); Tabushi et al., J. Am. Chem. 106, 4580-4584 (1984)). For example, β-cyclodextrin can be reacted with biphenyl-4,4'-disulfonyl chloride in the presence of anhydrous pyridine to form β-cyclodextrin blocked by biphenyl-4,4'-disulfonyl chloride, which can then be reacted with potassium iodide to give diiodo-β β-cyclodextrin. The cyclodextrin monomer precursor is iodinated in only two positions. By copolymerizing the diiodinated cyclodextrin monomer precursor with the comonomer A precursor as described above, a linear cyclodextrin polymer containing a repeating unit of formula Ia, Ib or a combination thereof, as also described above, can be obtained. If appropriate, then the iodine or iodine groups may be replaced with other known leaving groups.

In addition, according to the invention, iodine groups or another suitable leaving group can (optionally) be replaced by a group that allows interaction with the precursor of comonomer A, as described above. For example, a diiodinated cyclodextrin monomer precursor of the formula IVa, IVb, IVc or a mixture thereof may undergo an amination reaction to form a diaminated cyclodextrin monomer precursor of the formula Va, Vb, Vc or a mixture thereof:

The diaminated cyclodextrin monomer precursor can be prepared by any known method. (Tabushi et al. Tetrahedron Lett. 18: 1527-1530 (1977); Mungall et al., J. Org. Chem. 1659-1662 (1975)). For example, diiodo-β-cyclodextrin can be reacted with sodium azide and then reduced to form diamino-β-cyclodextrin.

The cyclodextrin monomer precursor is aminated in only two positions. The diaminated cyclodextrin monomer precursor can then be copolymerized with the monomer precursor A as described above to obtain a linear cyclodextrin copolymer containing a repeating unit of formula Ia, Ib or a combination thereof, as also described above. However, the amino functionality of the diaminated cyclodextrin monomer precursor is optionally attached directly to the cyclodextrin fragment. Alternatively, amino functionality may be introduced by replacing the iodine atom or other suitable leaving groups of the cyclodextrin monomer precursor with amino group fragments, such as, for example, -SCH ₂ CH ₂ NH ₂ , to form the diaminated cyclodextrin monomer precursor of formula Vd, Ve, Vf, or a mixture thereof .

The linear cyclodextrin copolymer of the present invention can also be obtained by reducing the linear oxidized cyclodextrin copolymer of the present invention, as described below. This method can be carried out if comonomer A does not contain a fragment capable of undergoing reduction or a group, such as, for example, a disulfide bridge.

According to the present invention, the linear cyclodextrin copolymer of the present invention can be oxidized so that at least one oxidized cyclodextrin monomer is introduced into the copolymer so that the oxidized cyclodextrin monomer is an integral part of the backbone of the polymer macromolecule. A linear cyclodextrin copolymer that contains at least one oxidized cyclodextrin monomer is called a linear oxidized cyclodextrin copolymer. The cyclodextrin monomer can be oxidized either on the side of the secondary hydroxyl groups or on the side of the primary hydroxyl groups of the cyclodextrin fragment.

If several oxidized cyclodextrin monomers are present in the linear oxidized cyclodextrin copolymer of the invention, the same or different cyclodextrin monomers may be present, oxidized on the side of the primary hydroxyl groups or secondary hydroxyl groups, or on both sides. For example, a linear copolymer of oxidized cyclodextrin with oxidized secondary hydroxyl groups contains, for example, at least one elementary unit of formula VIa or VIb:

In formulas VIa and VIb, C is a substituted or unsubstituted monomer of oxidized cyclodextrin, and A is a comonomer joined, for example, by covalent bonding with oxidized cyclodextrin C. In addition, in formulas VIa and VIb, oxidation of the secondary hydroxyl groups leads to the opening of the cyclodextrin fragment ring and the formation of aldehyde groups.

A linear oxidized cyclodextrin copolymer can be prepared by oxidizing a linear cyclodextrin copolymer as described above. The oxidation of a linear cyclodextrin copolymer can be carried out using known oxidation methods. (Hisamatsu et al., Starch 44: 188-191 (1992)). An oxidizing agent such as, for example, sodium periodate is preferably used. It will be appreciated by those skilled in the art that, under standard oxidation conditions, the degree of oxidation based on the polymer may or may be changed. Thus, in one embodiment of the invention, the linear oxidized copolymer of the present invention may contain one oxidized cyclodextrin monomer. In another embodiment, substantially all or all of the cyclodextrin monomers of the copolymer will be oxidized.

Another method for preparing a linear oxidized cyclodextrin copolymer of the invention involves oxidizing the starting diiodated or diaminated cyclodextrin monomer as described above to form the starting oxidized diiodated or diaminated cyclodextrin monomer and copolymerizing the oxidized diiodinated or diaminated cyclodextrin monomer precursor with the precursor oxidized precursor cyclodextrin new monomer of the formula VIIa, VIIb, VIIc or a mixture thereof:

can be obtained by (a) oxidizing a diiodinated cyclodextrin monomer precursor of the formulas IVa, IVb, IVc, or a mixture thereof, as described above. In another preferred embodiment, the oxidized diaminated cyclodextrin monomer precursor of formulas VIIIa, VIIIb, VIIIc, or a mixture thereof:

can be obtained (a) by amination of a precursor of an oxidized diiodinated cyclodextrin monomer of formulas VIIa, VI Ib, VIIc, or a mixture thereof, as described above. In another preferred embodiment, the oxidized diaminated cyclodextrin monomer precursor of formulas IXa, IXb, IXc, or a mixture thereof:

can be prepared by (a) substituting iodine or other suitable leaving groups for an oxidized cyclodextrin monomer precursor disubstituted with iodine or another suitable leaving group with an amino group fragment, for example, —SCH ₂ CH ₂ NH ₂ .

Alternatively, the oxidized diiodinated or diaminated cyclodextrin monomer precursor as described above can be prepared by oxidizing the cyclodextrin monomer precursor to form the oxidized cyclodextrin monomer precursor and then diiodating and / or diaminating the oxidized cyclodextrin monomer as described above. As described above, the cyclodextrin fragment can be modified by other leaving groups other than the iodine group and other functionalities containing an amino group. The oxidized diiodinated or diaminated cyclodextrin monomer precursor can then be copolymerized with the comonomer A precursor as described above to form a linear oxidized cyclodextrin copolymer of the present invention.

The linear oxidized cyclodextrin copolymer may also be further modified by the addition of at least one ligand to the copolymer. The ligand is described above.

In a preferred embodiment of the invention, the linear cyclodextrin copolymer or the linear oxidized cyclodextrin copolymer comprises at least one comonomer A precursor or a hydrolyzed comonomer A precursor product as described above at the end. As a result of the cyclodextrin copolymer terminating with at least one precursor of copolymer A, at least one free functional group, as described above, is located on a linear cyclodextrin copolymer or on a linear oxidized cyclodextrin copolymer. For example, the functional group may be an acidic group or a group that can undergo hydrolysis to give an acidic group. According to this invention, the functional group may undergo further chemical modification, if necessary, to improve the properties of the cyclodextrin copolymer, such as, for example, colloidal stability or transfection efficiency. For example, a functional group can be modified by reacting with PEG to form a cyclodextrin copolymer with a terminal PEG group to increase colloidal stability or with histidine to obtain a cyclodextrin copolymer with an imidazole terminal group to increase intracellular and transfection efficiency.

Additional chemical modification of the cyclodextrin copolymer can be carried out using a modified functional group. For example, a modified functional group can be used to build up the polymer chain by attaching a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer, as described herein, to the same or another cyclodextrin copolymer or to a non-cyclodextrin polymer. In a preferred embodiment of the invention, the polymer to be attached is the same or a different linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer, which may also contain at least one comonomer A precursor at the end for further modification, as described above.

Alternatively, at least two identical or different linear cyclodextrin copolymers or linear oxidized cyclodextrin copolymers containing at the end a functional group or a modified functional group, as described above, can interact and join together through a functional or modified functional group. Preferably, during the interaction of the functional or modified functional groups, a fragment is formed which is capable of decomposing, such as, for example, a disulfide bridge. For example, modification of the terminal functional group with cysteine can be used to produce a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer containing at least one free thiol group. Interaction with the same or another cyclodextrin copolymer, also containing at least one free thiol group, will lead to the formation of a disulfide bridge between the two copolymers. In a preferred embodiment of the invention, the functional or modified functional groups may be selected to provide bridges exhibiting different degradation rates (e.g., enzymatic degradation), and thereby provide a system with a long release time for the therapeutic agent if necessary. The resulting polymer may be crosslinked as described in this invention. The therapeutic agent described in this invention may be added before crosslinking or after crosslinking of the polymer. The ligand, which is described above, can also be attached via a modified functional group.

According to the invention, a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer can be crosslinked or grafted onto the substrate. The substrate can be any substrate known in the art. In another preferred embodiment of the invention, the linear cyclodextrin copolymer or the linear oxidized cyclodextrin copolymer may be crosslinked with the polymer to form a suitably crosslinked cyclodextrin copolymer or crosslinked oxidized cyclodextrin copolymer. The polymer may be any polymer capable of crosslinking with a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer of the present invention (e.g., polyethylene glycol (PEG), polyethylene). The polymer may also be the same or a different linear cyclodextrin copolymer or a linear oxidized dextrin copolymer. Thus, for example, a linear cyclodextrin copolymer can be crosslinked with any polymer, including, but not limited to, this copolymer itself, another linear cyclodextrin copolymer and a linear oxidized cyclodextrin copolymer. A crosslinked linear cyclodextrin copolymer of the present invention can be prepared by reacting a linear cyclodextrin copolymer with a polymer in the presence of a crosslinking agent. A crosslinked linear oxidized cyclodextrin copolymer of the present invention can be prepared by reacting a linear oxidized cyclodextrin copolymer with a polymer in the presence of a suitable crosslinking agent. The crosslinking agent may be any crosslinking agent known in the art. Examples of crosslinking agents include dihydrazides and disulfides. In a preferred embodiment, the crosslinking agent is a mobile group, so that the crosslinked copolymer can be converted to an uncrosslinked, if necessary.

The linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer of the present invention may be characterized by any methods known in the art. Such characterization methods or methods include, but are not limited to, gel permeation chromatography (GPC), matrix assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF Mass spec), ¹ H and ¹³ C NMR, light scattering and titration.

The invention also relates to a cyclodextrin composition comprising at least one linear cyclodextrin copolymer and at least one linear oxidized cyclodextrin copolymer of the present invention, as described above. Accordingly, either one of a linear copolymer of cyclodextrin and a linear copolymer of oxidized cyclodextrin, or both of these copolymers can be crosslinked with another polymer and / or connected to a ligand, as described above. The therapeutic compositions of this invention comprise a therapeutic agent and a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer, including crosslinked copolymers of the present invention. A linear cyclodextrin copolymer, a linear oxidized cyclodextrin copolymer and derivatives thereof are described above. The therapeutic agent may be any synthetic or natural biologically active therapeutic agent, including known therapeutic agents. Examples of suitable therapeutic agents include, but are not limited to, antibiotics, steroids, polynucleotides (e.g., genomic DNA, cDNA, mRNA and antisense oligonucleotides), plasmids, peptides, peptide fragments, small molecules (e.g., doxorubicin) and other biologically active macromolecules, such such as proteins and enzymes.

The therapeutic composition of the invention can be obtained using known methods. In a preferred embodiment, the copolymer of the invention is mixed with a therapeutic agent as described above for spontaneous aggregation. According to the invention, the therapeutic agent and the linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer are associated with each other so that the copolymer acts as a delivery vehicle for the therapeutic agent. The therapeutic agent and the cyclodextrin copolymer may be associated through causes known to the skilled person, for example through electrostatic interaction and hydrophobic interaction. The degree of association can be determined by known methods, including, for example, fluorescence studies, DNA motility studies, light scattering, electron microscopy, and will vary depending on the therapeutic agent.

For example, a therapeutic composition of the present invention containing a copolymer of the present invention and DNA can be used as a delivery method to facilitate transfection, that is, the introduction of DNA into a cell of a living organism (e.g., human). (Boussif, O. Proceedings of the National Academy of Sciences, 92: 7297-7301 (1995); Zanta et al. Bloconjugate Chemistry, 8: 839-844 (1997)).

The therapeutic composition of the invention may be, for example, a solid, liquid, suspension or emulsion. Preferably, the therapeutic composition of the invention is a preparation that can be administered intravenously. Other methods of administering the therapeutic composition of the invention include, depending on the physical state of the therapeutic composition, known methods such as, but not limited to, oral administration, topical administration, parenteral, intravenous, intranasal, intraocular, intracranial or intraperional injection.

Depending on the type of therapeutic agent used, the therapeutic composition of the present invention can be used in various therapeutic methods (e.g., DNA vaccines, antibiotics, antiviral drugs) for the treatment of hereditary or acquired disorders, such as, for example, cystic fibrosis, Gaucher disease, muscular dystrophy, AIDS, cancers (e.g., multiple myeloma, leukemia, melanoma, and ovarian carcinoma), cardiovascular disorders (e.g., progressive heart failure accuracy, restenosis and hemophilia) and neurological disorders (e.g., brain injury). According to the invention, a treatment method comprises administering a therapeutically effective amount of a therapeutic composition of the invention. It is clear to one skilled in the art that a therapeutically effective amount will be determined for each case. Factors to be considered include, but are not limited to, the disease to be treated and the physical characteristics of the patient suffering from the disease.

Another embodiment of the invention is a composition comprising at least one biologically active compound for agricultural use, and a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer of the present invention. For example, suitable agricultural biologically active compounds include, but are not limited to, fungicides, herbicides, insecticides, and plant disease control agents.

The following examples are given to illustrate the invention. However, it should be understood that the invention is not limited to the specific conditions or certain details described in these examples.

EXAMPLES

Materials: β-cyclodextrin (Cerestar USA, Inc. of Hammond, IN) was dried in vacuo (<0.1 mTorr) at 120 ° C. before use. Diphenyl-4,4'-disulfonyl chloride (Aldrich Chemical Comranu, Inc. of Milwaukee, WI) was recrystallized from chloroform / hexanes. Potassium iodide is ground into powder using a mortar and pestle and dried in an oven at 200 ° C. All other reagents are obtained from commercial sources and used without further purification. Polymer samples were analyzed by HPLC using a Hitachi HPLC system equipped with an Anspec RI detector and a Progel-TSK G3000 _pwxL column using water with an _ejection volumetric flow rate of 1.0 ml / min ^-1 .

Example 1: β-Cyclodextrin blocked at positions A and D by biphenyl-4,4'-disulfonyl, 1 (Tabushi et al. J. Am. Chem. Soc. 106, 5267-5270 (1984))

A 500 ml round bottom flask equipped with a magnetic stirrer, Schlenk adapter and membrane was charged with 7.92 g (6.98 mmol) of dry β-cyclodextrin and 250 ml of anhydrous pyridine (Aldrich Chemical Company, Inc). The resulting solution was stirred at 50 ° C. under nitrogen atmosphere, during which 2.204 g (6.28 mmol) of biphenyl-4,4′-disulfonyl chloride was added in four equal portions at 15 minute intervals. After stirring at 50 ° C. for an additional 3 hours, the solvent was removed in vacuo and the residue was purified by reverse phase column chromatography using gradient elution (0-40% acetonitrile in water). Fractions are analyzed by high performance liquid chromatography (HPLC) and the appropriate fractions are combined. After removal of acetonitrile on a rotary evaporator, the resulting aqueous suspension is lyophilized to dryness. The result is 3.39 g (38%) of 1 as a colorless solid.

Example 2: 6 ^A , 6 ^D- Diiod-6 ^A , 6 ^D- deoxy-β-cyclodextrin, 2 (Tabushi et al. J. Am. Chem. Soc. 106, 4580-4584 (1984))

In a 40 ml centrifugation tube equipped with a magnetic stirrer, a Schlenk adapter and a membrane, 1.02 g (7.2 mmol) of 1, 3.54 g (21.3 mmol) of dry powdered potassium iodide (Aldrich) and 15 ml are loaded anhydrous N, N-dimethylformamide (DMF) (Aldrich). The resulting suspension was stirred at 80 ° C under nitrogen atmosphere for 2 hours. After cooling to room temperature, solids were isolated by centrifugation, and the supernatant was decanted. The solid precipitate was washed with a second portion of DMF and the supernatants were combined and concentrated in vacuo. After that, the residue was dissolved in 14 ml of water, cooled in an ice bath and then 0.75 ml (7.3 mmol) of tetrachlorethylene (Aldrich) was added with vigorous stirring. The precipitate containing the complex is filtered on a glass frit medium, washed with a small amount of acetone and dried in vacuum over P ₂ O ₅ for 14 hours. The result is 0.90 g (92%) of 2 as a white solid.

Example 3: 6 ^A , 6 ^D- Diazo-6 ^A , 6 ^D- deosi-β-cyclodextrin, 3 (Tabushi et al. Tetrahedron Lett. 18, 1527-1530 (1977))

1.704 g (1.25 mmol) of β-cyclodextrin diiodide, 0.49 g (7.53 mmol) of sodium azide (EM Science of Gibbstown, NJ) are charged into a 100 ml round bottom flask equipped with a magnetic stirrer, Schlenk adapter and membrane. and 10 ml of anhydrous N, N-dimethylformamide (DMF). The resulting suspension was stirred at 60 ° C under nitrogen atmosphere for 14 hours. After that, the solvent is removed in vacuo. The resulting residue was dissolved in enough water to obtain a 0.2 M salt solution and then passed through 11.3 g of Biorad AG501-X8 (D) resin to remove residual salts. After this, the eluate is lyophilized to dryness, resulting in 1.232 g (83%) 3 as a white solid amorphous substance, which is used in the next step without further purification.

Example 4: 6 ^A , 6 ^D- Diamino-6 ^A , 6 ^D- deoxy-β-cyclodextrin, 4 (Mungall et al., J. Org. Chem. 1659-1662 (1975))

Into a 250 ml round bottom flask equipped with a magnetic stirrer and membrane, 1.232 g (1.04 mmol) of β-cyclodextrin bis-azide and 50 ml of anhydrous pyridine (Aldrich) are charged. 0.898 g (3.42 mmol) of triphenylphosphine are added to this suspension with stirring. The resulting suspension was stirred for one hour at room temperature, and then 10 ml of concentrated aqueous ammonia was added. The addition of ammonia is accompanied by a rapid evolution of gas, and the solution becomes homogeneous. After 14 hours, the solvent was removed in vacuo and the residue was triturated with 50 ml of water. The solids were filtered off, the filtrate was acidified (pH <4) with 10% HCl, and then the resulting solution was loaded onto an ion exchange column containing Toyopearl SP-650M resin (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{4}}$ the form). Product 4 was eluted with ammonium bicarbonate (eluent with a gradient of 0-0.5 M ammonium bicarbonate). The appropriate fractions are combined and lyophilized to give 0.832 g (71%) of product 4 as a bis (bicarbonate) salt.

Example 5: a copolymer of b-cyclodextrin and DSP, 5

A solution of 92.6 mg (7.65 × 10 ^-5 mol) of bis (bicarbonate) salt 4 in 1 ml of water is loaded into a 20 ml scintillation tube. The pH of the solution was adjusted to 10 with a 1M NaOH solution and then a solution of 30.9 mg (7.65 × 10 ^-5 mol) of dithiobis (succinimidyl propionate) (DSP, Pierce Chemical Co. of Rockford, IL) in 1 ml of chloroform was added. . The resulting biphasic mixture was stirred with a Vortex mixer for 0.5 hours. After that, the aqueous layer is decanted and extracted with freshly distilled chloroform. The aqueous polymer solution is then analyzed by gel permeation chromatography (GPC) on Toypearl HW-40F resin using water as the eluent. Fractions were analyzed by GPC and the corresponding fractions were lyophilized to give 85 mg (85%) of the product as a colorless amorphous powder.

Example 6: a copolymer of β-cyclodextrin and DSS, 6

The β-cyclodextrin copolymer and DSS 6 are synthesized in a similar manner to the preparation of β-cyclodextrin copolymer and DSP 5, with the difference that DSP is replaced by disuccinimidyl suberate (DSS, Pierce Chemical Co. of Rockford, IL). Compound 6 was obtained in 67% yield.

Example 7: a copolymer of β-cyclodextrin and DTBP, 7

A solution of 91.2 mg (7.26 x 10 ^-5 mol) of bis (bicarbonate) salt 4 in 1 ml of water is charged into a 20 ml scintillation flask. The pH of the solution was adjusted to 10 with a 1 M NaOH solution and 22.4 mg (7.26 × 10 ^-5 mol) of dimethyl-3,3'-dithiobis (propionimidate) 2CH1 (DTBP, Prierce Chemical Co. Of Rockford, IL). The resulting homogeneous solution was stirred with a Vortex mixer for 0.5 hours. The aqueous polymer solution is chromatographed by gel permeation chromatography (GPC) on Toypearl HW-40F resin using water as an eluent. Fractions were analyzed by GPC and the corresponding fractions lyophilized to give 67 mg (67%) of the product as a colorless amorphous powder.

Example 8: Copolymer of b-cyclodextrin and cystamine, 8

To a solution of 166.2 mg (7.38 × 10 ^-5 mol) of cystamine dihydrochloride (Aldrich) in 15 ml of a 0.1 N NaOH solution, 100 mg (7.26 × 10 ^-5 mol) of 2 and 5 ml of acetonitrile are added. The resulting homogeneous solution was heated to 80 ° C, maintained at this temperature for 2 hours and chromatographed using gel permeation chromatography (GPC) on Toyopearl HW-40F resin (eluent: water). Fractions were analyzed by GPC and the corresponding fractions were lyophilized to give 17.2 mg (19%) of the product as a colorless amorphous powder.

Example 9: Polyethylene glycol 6000 dihydrazide, 9

1.82 g (3.0 mmol) of polyethylene glycol 600 (Fluka Chemical Corp. of Milwaukee, WI), 40 ml of absolute ethanol (Quantum Chemicals Pty Ltd of Tuscola, IL) are charged into a 100-volume round bottom flask equipped with a magnetic stirrer and reflux condenser. ) and a few drops of sulfuric acid. The resulting solution was refluxed for 14 hours. Solid sodium carbonate is added to terminate the reaction, and a PEG diester solution is transferred to the dropping funnel under a nitrogen atmosphere. This solution was added dropwise to a solution of 0.6 ml (9.0 mmol) of hydrazine hydrate (Aldrich) in 10 ml of absolute ethanol. A small amount of cloudy precipitate forms. The resulting solution was refluxed for 1 hour, filtered and concentrated. GPC reveals contaminants with a higher molecular weight in the product. Gel permeation chromatography on Toyopearl HW-40F resin allows you to partially purify the product to obtain the target compound with a purity of 85%.

Example 10: Oxidation of a copolymer of β-cyclodextrin and DSS, 10 (Husamatsu et al., Starch 44, 188-191 (1992))

The copolymer of β-cyclodextrin and DSS 6 (92.8 mg, 7.3 × 10 ^-5 mol) is dissolved in 1.0 ml of water, cooled in an ice bath and 14.8 mg (7.38 × 10 ^-5 mol) is added sodium periodate. The solution immediately acquires a bright yellow color, and it is stirred in the dark at 0 ° C for 14 hours. The solution is then chromatographed using gel permeation chromatography (GPC) on Toyopearl HW-40F resin (eluent: water). Fractions analyze GPC. The appropriate fractions were combined and lyophilized to dryness, resulting in 84.2 mg (91%) of the product as a bright yellow amorphous powder.

Example 11: Polyethylene glycol (PEG) 600 diacid chloride, 11

A 50 ml round bottom flask equipped with a magnetic stirrer and reflux condenser was charged with 5.07 g (ca. 8.4 mmol) of 600 polyethylene glycol diacid (Fluka Chemical Corp. of Milwaukee, WI) and 10 ml of anhydrous chloroform (Aldrich). To this solution, 3.9 ml (53.4 mmol) of thionyl chloride (Aldrich) was added with stirring, and the resulting solution was refluxed for 1 hour, during which gas evolution was observed. The resulting solution was allowed to cool to room temperature, and the solvent and excess thionyl chloride were removed in vacuo. The resulting oil is stored in a dry box and used without purification.

Example 12: a copolymer of β-cyclodextrin and PEG 600, 12

A solution of 112.5 mg (8.95 × 10 ^-5 mol) of bis (bicarbonate) salt of 6 ^A , 6 ^D- diamino-6 ^A , 6 ^D- deoxy-β-cyclodextrin, 50 μl (50 μl) is loaded into a 20 ml scintillation tube 3.6 x 10 ^-4 mol) of triethylamine (Aldrich) and 5 ml of anhydrous N, N-dimethylacetamide (DMAc, Aldrich). The resulting suspension is treated with 58 mg (9.1 × 10 ^-5 mol) of polyethylene glycol 600, dichloride, 11. The resulting solution is stirred with a Vortex mixer for 5 minutes and then left at 25 ° C for 1 hour, during which the solution becomes homogeneous. The solvent was removed in vacuo and the residue was chromatographed by gel permeation chromatography on Toyopearl HW-40F resin (eluent: water). Fractions were analyzed by GPC and the corresponding fractions were lyophilized to dryness to give 115 mg (75%) of the product as a colorless amorphous powder.

Example 13: a copolymer of β-cyclodextrin and DSP, 13

A solution of 102.3 mg (8.80 x 10 ^-5 mol) of 2 ^A , 2 ^A- diamino-2 ^A , 3 ^A- deoxy-β-cyclodextrin in 1 ml of water is loaded into an 8 ml tube. The pH of the solution was adjusted to 10 with a 1M NaOH solution and a solution of 36.4 mg (8.80 x 10 ^-5 mol) of dithiobis (succinimidyl propionate) (DSP, Pierce Chemical Co. of Rockford, II) in 1 ml of chloroform was added. The resulting two-phase mixture was stirred with a Vortex mixer for 0.5 hours. The aqueous layer was decanted and extracted with freshly distilled chloroform (3 × 1 ml). The aqueous polymer solution is chromatographed using gel permeation chromatography.

Example 14: 6 ^A , 6 ^D- Bis- (2-amino-ethylthio) -6 ^A , 6 ^D- deoxy-β-cyclodextrin, 14 (Tabushi, I: Shimokawa, K; Fugita, K. Tetrahedron Lett. 1977, 1527- 1530)

Into a 25 ml Schlenk flask equipped with a magnetic stirrer and membrane, 0.91 ml (7.37 mmol) of a 0.81 M solution of sodium 2-aminoethylthiolate in ethanol are charged. (Fieser, LF; Fieser M. Reagents for Organic Synthesis, Wiley: New York, 1967; Vol. 3, pp. 265-266). The solution was evaporated to dryness and the solid residue was redissolved in 5 ml of anhydrous DMF (Aldrich). 6 ^A , 6 ^D- diiodo-6 ^A , 6 ^D- deoxy-β-cyclodextrin (100 mg, 7.38 × 10 ^-5 mol) was added and the resulting suspension was stirred at 60 ° C under nitrogen atmosphere for 2 hours. After cooling to room temperature, the solution was concentrated in vacuo and the residue was redissolved in water. After acidification with a 0.1 N HCl solution, the solution is transferred to a Toyopearl SP-650M (NH) ion exchange resin column $\genfrac{}{}{0ex}{}{\text{+}}{\text{4}}$ form) and elute (sodium bicarbonate solution, eluent with a gradient of 0-0.4 M). The appropriate fractions are combined and lyophilized to dryness. The result is 80 mg (79%) of 14 as a white powder.

Example 15: copolymer of β-cyclodextrin (cystamine) and DTBP, 15

A solution of 19.67 mg (1.42 · 10 ^-5 mol) of bis (bicarbonate) salt 14 in 0.1 M NaHCO ₃ solution is loaded into a 4 ml test tube. The solution was cooled in an ice bath and 4.4 mg (1.4 · 10 ^-5 mol) of dimethyl-3,3'-dithiobispropionimidate-2HCl (DTBP, Pierce) was added. The resulting solution was stirred with a Vortex mixer and left for 1 hour at 0 ° C. The reaction was quenched with a 1M Tris-HCl solution, acidified to pH 4 with 0.1 n HCl. The aqueous polymer solution was subjected to gel permeation chromatography on Toyopearl HW-40F resin. Fractions were analyzed by GPC and suitable fractions were lyophilized to dryness. The result is 21.3 mg (100%) 15 in the form of a white powder.

Example 16: a copolymer of β-cyclodextrin (cystamine) and DSM, 16

In a 10 ml Schlenk flask equipped with a magnetic stirrer and membrane, 200 mg (1.60 × 10 ^-4 mol) of 14.44 μl (3.2 ml × 10 ^-4 mol) of triethylamine (Aldrich Chemical Co., Milwaukee, WI), 43.6 mg (1.60 × 10 ^-4 ) of dimethylsuberimidate-2HCl (DMS, Pierse) and 3 ml of anhydrous DMF (Aldrich Chemical Co., Milwaukee, WI). The resulting suspension is heated to 80 ° C and maintained at this temperature for 18 hours under a steady stream of nitrogen, during which most of the solvent evaporates. The resulting residue was redissolved in 10 ml of water and the resulting solution was acidified with 10% Hcl to pH 4. Then, the resulting solution was passed through an Amicon Centricon Plus-20 5000 NMWL centrifugal filter. After washing with water (2 × 10 ml), the polymer solution was lyophilized to dryness, resulting in 41.4 mg (18%) of the product as a yellow amorphous solid.

Example 17: Attachment of a folate ligand to a cyclodextrin polymer 1. Addition of a resin:

50 mg of FMOC-PEG _3400- NHS (Shearwater Polymers, Inc. of Huntsville, AL) was dissolved in 1 ml of anhydrous N, N-dimethylformamide (DMF) and added to 10 equivalents of hydrazide-2-chlorotrityl resin (Novobiochem USA of La Jolla, CA) swollen in DMF. The mixture is stirred at 60 ° C until the entire polymer is attached to the resin, as determined by a GPC system equipped with an ultraviolet detector. The resin-polymer compound is then transferred to a sintered glass column for further interaction.

2. Blocking the functional groups of the resin:

Unreacted hydrazide groups on the resins are blocked with acetic anhydride and products derived from acetic acid are neutralized with diisopropylene ethylamine.

3. Removal of the protective group:

The FMOC protecting group is removed by two washes with a 20% solution of piperidine in DMF (total volume 1 ml). Then the resin is washed 10 times with DMF (each time 1 ml) and 5 times with H ₂ O (each time 1 ml).

4. Blocking functional groups with folic acid:

10 equivalents of folic acid and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide (EDC) are added to the resin along with 1.5 ml of H ₂ O. 1N is added to the reaction mixture. NaOH solution until folic acid dissolves (approximately pH 10). Then the glass column is placed on a rotator and stirring is continued overnight. The resin is washed with 10 × 1 ml NaOH (1 N), 10 × 1 ml 50 mm sodium bicarbonate and then 5 times with water, THF and dichloromethane.

5. Isolation from resin:

1% trifluoroacetic acid (TFA) in 1 ml of DCM is added twice to the resin each time for a minute. The supernatant was collected and the DCM was evaporated. The resulting oily film was rehydrated in H ₂ O and lyophilized, resulting in a light yellow powder. NMR analysis confirms the presence of the PEG polymer.

6. Binding to the polymer:

The coupling agent-folic acid compound is reacted with 6 equivalents of a cyclodextrin copolymer (oxidized as in Example 10) by mixing in 50 mmoles of borate (pH 8.5). The reaction mixture is analyzed and the presence of the conjugated polymer is confirmed by a GPC system with an ultraviolet detector at 285 nm. (See Scheme 1 at the end of the description).

Example 18: Attachment of a Folate Ligand to a Cyclodextrin Polymer

1. Binding:

36 mg of tert-butylcarbazate was dissolved in 240 μl of a DCM / ethyl acetate (1: 1) mixture, added to 260 mg of FMOC-PEG _3400- NHS (Shearwater Polymer) and stirred at room temperature for 2 hours. The product is precipitated twice from a mixture of ethyl acetate / ether (1: 1).

2. Removal of the protective group:

The FMOC protecting group is removed with 20% piperidine in DMF. The solvent was removed in vacuo and the product was redissolved in 1.3 ml of DMSO.

3. Blocking functional groups with folic acid:

One drop of pyridine is added to 1.2 equivalents of folic acid and DCC, and the resulting solution is stirred in the dark at room temperature for 6 hours. DMSO is removed in vacuo and the presence of folic acid conjugation is confirmed by GPC with UV monitoring at 285 nm.

4. Removal of the protective group of hydrazide:

Finally, the hydrazide group is removed by stirring in 4M HCl in dioxane for one hour, followed by removal of the solvent in vacuo. The final product is purified using Toyopearl HW-40F column chromatography.

5. Binding to the polymer:

The folic acid-binding agent compound is reacted with 6 equivalents of a cyclodextrin copolymer (oxidized as in Example 10) by mixing in 50 mmoles of borate (pH 8.5). The reaction mixture was analyzed and the presence of the conjugated polymer was confirmed using a GPC system with an ultraviolet detector at 285 nm. (See Scheme 2 at the end of the description).

Example 19: attaching a transferrin ligand to a cyclodextrin polymer

1. The oxidation of transferrin

50 mg of iron-free human transferrin (Sigma of St. Louis, MO) is dissolved in 30 mM sodium acetate buffer and cooled to 0 ° C. To this solution was added 20 mg of sodium periodate dissolved in 4 μl of a 30 mM sodium acetate solution. The mixture was stirred at 0 ° C. overnight. Then add 1 g of resin AG501-X8 (Biorad) to remove salts and the solution is lyophilized.

2. Binding to the resin:

20 mg FMOC-PEG _3400- NHS (Snearwater Polymers, Inc. of Huntsville, AL) was dissolved in 0.5 ml of anhydrous N, N-dimethylformamide (DMF) and 10 equivalents of 2-chlorotrityl resin (Novabiochem USA of La Jolla) was added , CA) swollen in DMF. The mixture is stirred at 60 ° C until the entire polymer is bound to the resin, as determined by a GPC system equipped with an ultraviolet detector. The resin-polymer compound is transferred to an agglomerated glass column for all additional reactions.

3. Blocking the functional groups of the resin:

Unreacted hydrazide groups on the resins are blocked with acetic anhydride and products derived from acetic acid are neutralized with diisopropylethylamine.

4. Removal of the protective group:

The FMOC protecting group is removed by washing twice with 20% piperidine in DMF (total volume 1 ml). The resin is then washed 10 times (1 ml each time) with DMF and five times (1 ml each time) H ₂ O.

5. Linking with transferrin:

1.2 equivalents of transferrin dissolved in a 0.05 M sodium carbonate solution and 0.1 M sodium citrate buffer, pH 9.5, are added to the resin. Then, 5 M cyanoborohydride in 1N was added to the solution. NaOH solution. The glass column is placed on a rotator and stirred for 2 hours. The resin is then washed 15 times with water and 5 times each time with tetrahydrofuran (THF) and DCM.

6. Isolation from resin:

1% trifluoroacetic acid (TFA) in 1 ml of DCM is added twice to the resin, each time for a minute. The supernatant was collected and the DCM was evaporated. The resulting oily film was rehydrated in H ₂ O and lyophilized.

7. Binding to the polymer:

A compound of the binding agent-transferrin composition is reacted with 6 equivalents of a cyclodextrin copolymer by reductive amination with sodium cyanoborohydride: first, the copolymer is added to the transferrin binding agent dissolved in a 0.05 M sodium carbonate solution and 0.1 M sodium citrate buffer. Add 5 M cyanoborohydride in 1 N. NaOH solution and the reaction mixture was stirred for two hours at room temperature. Unreacted aldehyde sites are blocked by adding ethanolamine and reacting for 15 minutes at room temperature. The resulting conjugate was purified by dialysis. (See diagram 3 at the end of the description).

Example 20: General Procedure for the Preparation of Small Molecule Cyclodextrin Complexes

A cyclodextrin-based copolymer (CD polymer) is dissolved in water, a buffer, or an organic solvent to an appropriate concentration. The small molecule compound is dissolved in a solvent miscible with the solvent of the CD polymer solution and added to the CD polymer solution. The mixture was then stirred for _^1/2 hour and allowed to equilibrate overnight.

Example 21: Formation of a complex of a copolymer of cyclodextrin with doxorubicin.

Doxorubicin and the CD polymer are dissolved at various concentrations in PBS (phosphate buffered saline, pH 7.2). The association constant between CD and doxorubicin is determined by measuring the degree of increase in fluorescence of doxorubicin as a result of coplex formation with CD. (The hydrophobic interaction of CD and doxorubicin increases the fluorescence intensity). The association constant is approximately 200 M ^-1 at pH 7.1. The addition of β-CD significantly increases the fluorescence of doxorubicin, which is an indicator of the complexation between the CD polymer and doxorubicin. In the publication of Husain et al., Applied Spectroscopy Vol. 46, No. 4, 652-658 (1992), it is shown that the association constant between β-CD and doxorubicin is 210 M ^-1 at pH 7.1.

Example 22: Delivery of small molecules to cultured cells

For cultured cell lines, a medium containing doxorubicin and doxorubicin / CD polymer complexes at various concentrations is used. After 5 hours, the medium is removed and replaced with fresh medium. The effect of doxorubicin on cell survival is determined using an MTT toxicity assay ([3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl-2H-tetrazolium). (R. Ian Feshney, “Culture of animal cells”, 3 ^rd ed., Wiley-Liss: New York (1994)). The results are presented in the table below. A copolymer of 15 or 16 (138 μM equivalent of CD monomer) is non-toxic to KB or KB-VI (a derivative of a multi-drug resistant KB) cell lines in the absence of doxorubicin. For receptor-based delivery, a ligand, such as a folate covalently attached to a CD polymer, is used for complexation with doxorubicin.

Example 23: obtaining a complex of a copolymer with a fixed constant charge and plasmids

Typically, equal volumes of a fixed charge CD polymer and plasmid DNA are mixed at appropriate polymer / plasmid charge ratios. The mixture is then allowed to reach equilibrium and left overnight for spontaneous aggregation at room temperature. The progress of complexation is controlled by transferring a small aliquot of the mixture into a 0.6% agarose gel and checking the DNA mobility. Free DNA moves under the influence of the applied voltage, while complexed DNA slows down in the reservoir.

1 μg of DNA with a concentration of 0.2 μg / μl in distilled water is mixed with 10 μl of copolymer 15 with a polymer amine: DNA phosphate charge ratio of 2, 4, 6, 12, 24, 36, 60 and 120. The solution is mixed manually with using a micropipette and mix gently overnight on a laboratory rotator. The next morning, 1 μg / μl of loading buffer (40% sucrose, 0.25% bromophenol blue, 20 mM Ttis acetate buffer containing 5 mM EDTA (Gao et al., Biochemistry 35: 1027-1036 (1996) )). Each sample was loaded onto a 0.6% agarose electrophoresis gel containing 6 μg EtBr / 100 ml in 1 × TAE buffer (40 mm Tris-acetate / 1 mm EDTA) and a voltage of 40 V was applied to the gel for 1 hour The degree of complexation of the DNA / polymer shows a slowdown of the DNA in the migration pattern in the gel. The polymer (15) slows down the DNA at a charge ratio of 6 or higher, which indicates a complexation ovany in these conditions.

Example 24: Complex formation of a crosslinked polymer with a plasmid Copolymer 15 or copolymer 16 is oxidized, as in Example 10. The oxidized copolymer 15 or 16 is then complexed with a DNA plasmid, as in Examples 23 and 26. A crosslinking agent is then added (for example, PEG ₆₀₀ - dihydrazide) for encapsulating DNA. The implementation of encapsulation is determined by scattering of light and using an electron microscope.

Example 25: Complexation of a copolymer with a variable (pH-sensitive) charge with a plasmid

Equal volumes of solutions of the CD polymer and plasmid DNA in water are mixed at the appropriate polymer charge / plasmid charge ratios. The mixture is left for 30 minutes to achieve equilibrium and spontaneous aggregation. A crosslinking agent (eg, REG _600- dihydrazide) is then added to encapsulate the DNA. Then a concentrated buffer solution was added to adjust the pH, and thus a neutral CD polymer was obtained. The implementation of the encapsulation is determined by scattering of light and visually using an electron microscope.

Example 26: Study of transfection with plasmids encoding the Luciferase reporter gene

BHK-21 cells are placed in 24-cell plates with a density of 60,000 cells / well 24 hours before transfection. Plasmids encoding the luciferase gene are encapsulated using a CD polymer, as described in examples 23 or 25, so that complexes are formed at a DNA / polymer charge ratio of 6, 12, 24, 36 and 60, as described in example 23 DNA binding studies. The medium containing the DNA / polymer complexes is added to the cultured cells and replaced with fresh medium after 5 hours of incubation at 37 ° C. Cells are lysed 48 hours after transfection. Suitable substrates for light evaluation of luciferase are added to the cell lysate. The luciferase activity measured with the resulting light units is counted using a luminometer. DNA / polymer complexes successfully transfect BHK-21 cells with charge ratios of 6, 12, and 24. Cell lysate is also used to determine cell viability using Lauri protein estimation. (Lowry et al., Journal of Biological Chemistry, Vol. 193, 265-275 (1951)). The maximum toxicity is observed at polymer amine: DNA phosphate charge ratios of 36 and 60 at 91% cell survival.

Example 27: Study of transfection with plasmids encoding the Lusiferase reporter gene

BHK-21 cells are placed in 24-cell plates with a density of 60,000 cells / well 24 hours before transfection. Plasmids encoding the luciferase gene are encapsulated using a CD polymer as described in Example 23, with the difference that polymer 15 is replaced with polymer 16 and that DNA / polymer complexes successfully transfect BHK-21 cells at charge ratios of 10, 20, 30 and 40 with maximum transfection with a polymer amine: DNA phosphate charge ratio of 20. A medium containing DNA / polymer complexes is added to the cultured cells and replaced with fresh medium after 24 hours of incubation at 37 ° C. Cells are lysed 48 hours after transfection. Suitable substrates for light evaluation of luciferase are added to the cell lysate. The luciferase activity measured with the resulting light units is counted using a luminometer. The results are presented below. DNA / polymer complexes successfully transfect BHK-21 cells with charge ratios of 6, 12, and 24. Cell lysate is also used to determine cell viability using Lauri protein estimation. (Lowry et al., Journal of Biological Chemistry, Vol. 193, 265-275 (1951)). The results are presented below. The maximum toxicity is observed at polymer amine: DNA phosphate charge ratios of 40 and 50 at 33% cell survival.

Example 28: Study of transfection with plasmids encoding the GFP reporter gene

Plasmids encoding a green fluorescent protein are encapsulated using a CD polymer as described in Examples 23 or 25. A medium solution containing DNA / polymer complexes is added to the cultured cells and replaced with fresh medium after incubation at 37 ° C. Cells are separated from the trypsin surface, washed and resuspended in a balanced Hanks saline solution with propidium iodide. After that, the cells are analyzed by sorting fluorescently activated cells (FACS). Cell viability is determined by the size of the cells and the exclusion of propidium iodide, and the successful implementation of transfection by GFP protein fluorescence.

Example 29: Formation of Polymer Complexes with Oliogens

Complexation with desensitizing oligogens is carried out in accordance with the methods for plasmid complexation of examples 23 and 25.

Example 30: Study of transfection with oligogens.

Desensitizing oligogens directed against the luciferase gene are encapsulated using a CD polymer as described in Example 29. A medium solution containing oligogen / polymer complexes is added to HeLa X1 / 5 cells (HeLa cells that stably express the luciferase gene provided by CLONTECH) and replaced with fresh medium after incubation for 5 hours at 37 ° C. Cells are lysed 48 hours after transfection and suitable substrates are added for quantitative analysis of luciferase. The luciferase activity, measured in the form of the obtained units of light, is calculated using a luminometer. Successful transferase activity is determined by capturing (by knockout) luciferase activity.

Example 31: Toxicity of a β-cyclodextrin (cystamine) copolymer and DTBP, 15

The active toxicity of copolymer 15 is investigated using a Swiss-Webster “white mouse”. A total of 48 mice were used, as described in table 2 below. Each mouse is subjected to a single intravenous (i.v.) or intraperitoneal (i.p.) infection with sterile saline or copolymer solution 15. After five days, the animals are sacrificed and completely opened. Mortality and toxicity are not observed.

Example 32: Study of transfection with plasmids encoding the Luciferase reporter gene

Plasmids encoding the luciferase gene are encapsulated using a CD polymer as described in Example 23, with the difference that copolymer 16 is used instead of copolymer 15. DNA / polymer complexes are used to successfully transfect BHK-21 or CHO-K1 cells, each placed on 24 cell plates with a cell density of 60,000 cells / cell 24 hours before transfection at different charge ratios in a medium containing 10% serum or serum free, in accordance with the procedure described in example 27. Cells are lysed 48 hours after transfection and. Appropriate substrates for quantitative light determination of luciferase are added to the cell lysate. The luciferase activity, measured from the calculation of the resulting light units (i.e. relative light units - relative light units (RLU)), is calculated using a luminometer. A cell lysate is also used to determine cell viability by Lauri protein quantification. (Lowry et al., Journal of Biological Chemistry, Vol. 193, 265-275 (1951)). Toxicity is measured by determining the total cellular protein in the cells 48 hours after transfection. The results of transfection and cell survival in 10% serum and serum-free medium are given below.

The activity of luciferase protein in BHK-21 cells transfected in serum-free medium reaches a stable maximum at 30 +/- 5 × 10 ⁷ RLU. The presence of 10% serum in a transfection medium reduces luciferase activity at all charge ratios, with the exception of 70 +/-. For CHO-K1 cells, an increase in the charge ratio also leads to an increase in transfection for all test conditions. Additionally, serum transfection reduces the number of light units by an order of magnitude.

Copolymer 16 shows toxicity only to BHK-21 cells for transfection in the absence of serum. Toxicity is reduced to a minimum in the presence of 10% serum during transfection. Noticeable toxicity was not observed with transfections on CHO-K1 cells.

) и выживание слеток (

) в ВНК-21 клетках. Результаты трансфекции в 10% сыворотке и среде, не содержащей сыворотки, представлены соответственно пунктирной и сплошной линиями. Данные представлены в виде среднего значения трех образцов +/- ст. отклонение. Данные токсичности представлены в виде линий наилучшего приближения (best fit lines).The effect of the ratio of the charges of the copolymer 16 / DNA and the presence of serum on the efficiency of transfection (

) and the survival of fights (

) in BHK-21 cells. The results of transfection in 10% serum and serum-free medium are represented by dashed and solid lines, respectively. The data are presented as the average of three samples +/- st. deviation. Toxicity data are presented as best fit lines.

) и выживание клеток (

) в СНО-К1 клетках. Результаты трансфекции в 10% сыворотке и в среде, не содержащей сыворотки, представлены соответственно пунктирной и сплошной линиями. Данные представлены в виде среднего значения трех образцов +/- ст. отклонение. Данные токсичности представлены в виде линий наилучшего приближения (best fit lines).The effect of the ratio of the charges of the copolymer 16 / DNA and the presence of serum on the efficiency of transfection (

) and cell survival (

) in CHO-K1 cells. The results of transfection in 10% serum and in serum-free medium are shown by dashed and solid lines, respectively. The data are presented as the average of three samples +/- st. deviation. Toxicity data are presented as best fit lines.

Comparative Example 1: Transfection Study with Plasmids Encoding the Luciferase Reporter Gene

In accordance with the procedure of example 32, the transfection efficiency and toxicity of various non-viral vectors with BHK-21 and CHO-K1 cells are studied and the results are compared with the results obtained with DNA / polymer complexes 16. BHK-21 and CHO-K1 cells are transfected in the range of charge ratios and initial densities for all vectors in a serum-free medium. The results are shown below and they represent the optimal transfection conditions established for each vector.

It should be understood that the above discussion and examples provide only a detailed description of certain preferred embodiments. It is clear to a skilled person that various modifications and equivalents can be made without isolating from the scope and scope of the invention. All patents, journal articles, and other documents discussed or cited above are incorporated herein by reference.

Claims (50)

1. A water-soluble linear cyclodextrin copolymer obtained by a process comprising a) producing at least one cyclodextrin monomer precursor allowing polymerization in only two positions; b) the interaction of the cyclodextrin monomer precursor with a compound containing two functional groups through which the cyclodextrin monomer precursors can be coupled, resulting in a water-soluble linear cyclodextrin copolymer comprising repeating unit units of formula Ia, Ib, or both:

where C is a substituted or unsubstituted cyclodextrin monomer; A is a comonomer associated with cyclodextrin C. 2. A water-soluble linear cyclodextrin copolymer containing cyclodextrin fragments as part of its main chain of the copolymer, including repeating elementary units of the formula Ia, Ib or both:

where C is a substituted or unsubstituted cyclodextrin monomer; A is a comonomer associated with cyclodextrin C. 3. The copolymer according to any one of claims 1 to 2, where the cyclodextrin fragments or cyclodextrin monomers are the same throughout the polymer. 4. The copolymer according to any one of claims 1 to 2, where the cyclodextrin fragments or cyclodextrin monomers contain at least two different cyclodextrin fragments. 5. The copolymer according to claim 4, where at least several cyclodextrin fragments or cyclodextrin monomers are substituted. 6. The copolymer according to claim 5, where at least one cyclodextrin monomer or fragment is replaced by a ligand capable of binding the copolymer to the cell. 7. The copolymer according to claim 1 or 2, where the cyclodextrin fragments or cyclodextrin monomers are α-, β-, γ-cyclodextrins or a combination thereof. 8. The copolymer according to claim 7, where cyclodextrin fragments or cyclodextrin monomers are independently selected from the group consisting of 6 ^A , 6 ^B- deoxy-α-cyclodextrin, 6 ^A , 6 ^C- deoxy-α-cyclodextrin, 6 ^A , 6 ^D- deoxy-α-cyclodextrin, 6 ^A , 6 ^B- deoxy-β-cyclodextrin, 6 ^A , 6 ^C -deoxy-β-cyclodextrin 6 ^A , 6 ^D- deoxy-β-cyclodextrin, 6 ^A , 6 ^B- deoxy-γ-cyclodextrin, 6 ^A , 6 ^{C -} deoxy-γ-cyclodextrin, 6 ^A , 6 ^D- deoxy-γ-cyclodextrin, 6 ^A , 6 ^E- deoxy-γ-cyclodextrin. 9. The copolymer according to claim 1 or 2, where these cyclodextrin fragments or cyclodextrin monomers contain oxidized cyclodextrin monomers having the general formula (III)

where p = 5-7. 10. The cyclodextrin copolymer according to claim 9, wherein said oxidized cyclodextrin fragments or cyclodextrin monomers include fragments selected from 2 ^A , 3 ^A- deoxy-2 ^A , 3 ^A- dihydro-α-cyclodextrin, 2 ^A , 3 ^A- deoxy- 2 ^A , 3 ^A- dihydro-β-cyclodextrin, 2 ^A , 3 ^A -deoxy-2 ^A , 3 ^A- dihydro-γ-cyclodextrin. 11. The copolymer according to claim 1, where the comonomer is selected from: -HNC (O) (CH ₂ ) _x C (O) NH-, -HNC (O) (CH ₂ ) _x SS (CH ₂ ) _x C (O) NH-, - ⁺ H ₂ N (CH ₂ ) _x SS (CH ₂ ) _x NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ -, -HNC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NH-, -HNNHC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NHNH-, - ⁺ H ₂ NCH ₂ (CH ₂ CH ₂ O) _x CH ₂ CH ₂ CH ₂ NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ -, -HNC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ SS (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NH-, -HNC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NH-, -SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-, -SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ ) _x SS (CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-, -SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) CH ₂ CH ₂ (OCH ₂ CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-,

and

where x = 1-50 and y + z = x. 12. The copolymer according to claim 1, where the comonomers contain biodegradable or acid-labile binders. 13. The copolymer according to claim 1 or 2, where the copolymer is crosslinked with another polymer. 14. The copolymer according to item 13, where at least one ligand is associated with a copolymer. 15. The copolymer according to claim 1 or 2, where at least one ligand is associated with the copolymer. 16. The copolymer according to claim 1 or 2, where at least one cyclodextrin fragment or monomer is oxidized. 17. The copolymer according to clause 16, where the cyclodextrin fragment is an α-, β-, γ-cyclodextrin or a combination thereof. 18. The copolymer according to clause 16, where the copolymer is crosslinked with another polymer. 19. The copolymer according to claim 18, where at least one ligand is associated with an oxidized copolymer. 20. The copolymer according to claim 16, where at least one ligand is associated with a copolymer. 21. The copolymer according to claim 1 or 2, where basically all of the cyclodextrin fragments are oxidized. 22. The copolymer according to claim 1 or 2, where all cyclodextrin fragments are oxidized. 23. A therapeutic composition comprising the copolymer according to any one of the preceding paragraphs and an effective amount of a therapeutic agent. 24. A composition comprising a) a first copolymer according to claim 16 and (b) a second water-soluble linear cyclodextrin copolymer comprising cyclodextrin fragments as an integral part of its main copolymer chain. 25. The composition of claim 24, wherein at least one of said first copolymer and said second copolymer is crosslinked with another polymer. 26. The composition according A.25, where at least one ligand is associated with at least one of these first and second copolymers. 27. The composition according to paragraph 24, where at least one ligand is associated with at least one of these first and second copolymers. 28. A therapeutic composition comprising a cyclodextrin composition according to any one of paragraphs.24-27 and an effective amount of a therapeutic agent. 29. A method of obtaining a water-soluble linear cyclodextrin copolymer comprising the following steps: a) producing at least one cyclodextrin monomer precursor that allows polymerization in only two positions; b) the interaction of the cyclodextrin monomer precursor with a compound containing two functional groups, through which the cyclodextrin monomer precursors are coupled, as a result of which a water-soluble linear cyclodextrin copolymer is obtained, including repeating elementary units of formula Ia, Ib or both:

where C is a substituted or unsubstituted cyclodextrin monomer; A is a comonomer associated with cyclodextrin C. 30. The method of claim 29, wherein said cyclodextrin monomer precursor is a diiodinated cyclodextrin monomer precursor of formula IVa, IVb, IVc, or a mixture thereof:

31. The method according to clause 29, where the specified cyclodextrin monomer precursor is α-, β-, γ-cyclodextrin or a combination thereof. 32. The method according to clause 29, wherein said cyclodextrin monomer precursor is selected from the group consisting of: 6 ^A , 6 ^B- deoxy-α-cyclodextrin, 6 ^A , 6 ^C- deoxy-α-cyclodextrin, 6 ^A , 6 ^D- deoxy -α-cyclodextrin, 6 ^A , 6 ^B -deoxy-β-cyclodextrin, 6 ^A , 6 ^C -deoxy-β-cyclodextrin, 6 ^A , 6 ^D -deoxy-β-cyclodextrin, 6 ^A , 6 ^B -deoxy-γ -cyclodextrin, 6 ^A , 6 ^C- deoxy-γ-cyclodextrin, 6 ^A , 6 ^D- deoxy-γ-cyclodextrin, 6 ^A , 6 ^E- deoxy-γ-cyclodextrin. 33. The method according to clause 29, where the specified cyclodextrin monomer precursor has the General formula (III)

where p = 5. 34. The method according to claim 33, wherein said cyclodextrin monomer precursor is selected from the group consisting of 2 ^A , 3 ^A- deoxy-2 ^A , 3 ^A- dihydro-α-cyclodextrin, 2 ^A , 3 ^A- deoxy-2 ^A , 3 ^A- dihydro-β-cyclodextrin, 2 ^A , 3 ^A- deoxy-2 ^A , 3 ^A- dihydro-γ-cyclodextrin. 35. The method according to clause 29, where the compound containing two functional groups is selected from the group including -HNC (O) (CH ₂ ) _x C (O) NH-, -HNC (O) (CH ₂ ) _x SS (CH ₂ ) _x C (O) NH-, - ⁺ H ₂ N (CH ₂ ) _x SS (CH ₂ ) _x NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ , -HNC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NH-, -HNNHC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NHNH-, - ⁺ H ₂ NCH ₂ (CH ₂ CH ₂ O) _x CH ₂ CH ₂ CH ₂ NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ -, -NC (O) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ SS (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (O) NH-, -HNC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ CH ₂ O) _x CH ₂ CH ₂ C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NH-, -SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-, -SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) (CH ₂ ) _x SS (CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ S-, -SCH ₂ CH ₂ NHC (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) CH ₂ CH ₂ (OCH ₂ CH ₂ ) _x C (NH $\genfrac{}{}{0ex}{}{\text{+}}{\text{2}}$ ) NHCH ₂ CH ₂ CH ₂ S-

or

where x = 1-50 and y + z = x. 36. The method according to clause 29, comprising the additional step of reacting said copolymer with a ligand to form a linear copolymer containing at least one ligand bound to the copolymer. 37. The method according to p. 30, comprising the additional stage of the amination of the specified precursor diiodinated cyclodextrin monomer with the formation of the precursor diaminated cyclodextrin monomer and the polymerization of the precursor diaminated cyclodextrin monomer with the formation of the specified copolymer. 38. The method according to clause 37, where the specified precursor to a diaminated cyclodextrin monomer is a monomer having the formula Va, Vb, or Vc, or a mixture thereof:

39. The method according to clause 37, where the specified predecessor of the diaminated cyclodextrin monomer is a monomer of the formula Vd, Ve, or Vf or a mixture thereof:

40. A method of producing a copolymer, comprising the step of reducing a linear oxidized cyclodextrin copolymer, provided that said linear oxidized cyclodextrin copolymer does not contain reducible comonomer A. 41. A method of producing a copolymer, comprising the step of oxidizing a copolymer according to claim 1 or 2. 42. The method according to paragraph 41, comprising the additional step of reacting said copolymer with a ligand to form a copolymer containing at least one ligand bound to the copolymer. 43. A method of obtaining a copolymer, comprising the following stages: (a) iodination of the oxidized cyclodextrin monomer precursor to form the oxidized diiodinated cyclodextrin monomer precursor of formula VIIa, VIIb, VIIc, or a mixture thereof:

(b) reacting said oxidized diiodinated cyclodextrin monomer precursor with comonomer A precursor to form a linear oxidized cyclodextrin copolymer. 44. The method according to item 43, comprising the additional step of reacting said linear oxidized cyclodextrin copolymer with a ligand to form a linear oxidized cyclodextrin copolymer containing at least one ligand bound to the copolymer. 45. A method of producing a copolymer comprising the steps of (a) aminating an oxidized diiodinated cyclodextrin monomer precursor to form an oxidized diaminated cyclodextrin monomer precursor of formula VIIIa, VIIIb, VIIIc, or a mixture thereof:

(b) reacting said oxidized diaminated cyclodextrin monomer precursor with comonomer A precursor to form a linear oxidized cyclodextrin copolymer. 46. A method of producing a copolymer comprising the steps of (a) aminating an oxidized diiodinated cyclodextrin monomer precursor to obtain an oxidized diaminated cyclodextrin monomer precursor of formula IXa, IXb, IXc, or a mixture thereof:

(b) reacting said oxidized diaminated cyclodextrin monomer precursor with comonomer A precursor to form a linear oxidized cyclodextrin copolymer. 47. A method of obtaining a crosslinked cyclodextrin copolymer, comprising reacting the copolymer according to claim 1 or 2 with a second copolymer in the presence of a crosslinking agent. 48. The method according to clause 47, where the specified second copolymer is a linear copolymer of cyclodextrin or a linear copolymer of oxidized cyclodextrin. 49. A method for delivering a therapeutic agent to a patient, comprising administering to the patient a therapeutically effective amount of a therapeutic composition according to claim 23. 50. A method for delivering a therapeutic agent to a patient, comprising administering to the patient a therapeutically effective amount of a therapeutic composition according to claim 28.