Title:  Detoxication of active pharmaceutical substances using cyclodextrine oligomers

United States Patent:  6,642,214

Issued:  November 4, 2003

Inventors:  Moser; Joerg G (Duesseldorf, DE)

Assignee:  CeramOptec Industries, Inc. (East Longmeadow, MA)

Appl. No.:  554223

Filed:  August 3, 2000

PCT Filed:  November 11, 1998

PCT NO:  PCT/EP98/07229

PCT PUB.NO.:  WO99/24474

PCT PUB. Date:  May 20, 1999

Abstract

A cyclodextrin oligomer comprising two cyclodextrins connected through a spacer at the secondary side, characterized in that said spacer comprises the unit B which is a rigid and preferably hydrophilic structural element.

Description of the Invention

The present invention relates to spacer-bridged cyclodextrin oligomers, and to complexes of such cyclodextrin oligomers with pharmaceutically active substances.

Cyclodextrins are circular glucose polymers which are referred to as .alpha.-, .beta.- or .gamma.-cyclodextrins, depending on the number of glucose units (6 to 8, respectively). A lipophilic cavity exists inside the oligoglucose ring. It is known that lipophilic substances can be enclosed within this cavity. Cyclodextrins are used, inter alia, for converting compounds having low solubility to a soluble complex by complex formation with cyclodextrin.

In Tetrahedron, Vol. 51, 2 (1995), p. 377-388, R. Breslow et al. describe cyclodextrin dimers which are capable of binding substrates having the correct geometry in aqueous solutions.

In the Journal of Inclusion Phenomena and Molecular Recognition in Chemistry, 27 (1997), p. 69-84, A. Ruebner et al. describe dimeric cyclodextrins having high binding affinities for porphyrinoid photosensitizers as a carrier system for the application of drugs in photodynamical cancer therapy.

In Tetrahedron Letters 25 (1984), p. 5533-5536, K. Fujita et al. describe the preparation of ditosylates of .beta.-cyclodextrin and their purification by reversed-phase chromatography.

In Tetrahedron Letters 18 (1977), p. 1527-1530, I. Tabushi et al. describe the specific bifunctionalization of cyclodextrin.

In Arch. Microbiol. 165 (1996), p. 206-212, R. Feederle et al. describe the purification and characterization of the enzyme cyclodextrinase from Klebsiella oxytoca.

In an electronic publication which is accessible under http://antas.agraria. uniss.it/electronic_papers/eccc3/bcd/welcome.htm, B. Manunza et al. describe a molecular dynamics study of the structure and internal movement of solvated .beta.-cyclodextrin.

In the Journal of Pharmaceutical Sciences 84 (1995), p. 1223-1230, U. S. Sharma et al. describe the pharmaceutical and physical properties of paclitaxel (taxol) complexes with cyclodextrins.

In SPIE Biomedical Optics 3191 (1997), p. 343-353, A. P. Savitsky et al. describe an avidin-biotin system for the selected transport of photosensitizers and other cytotoxic agents into tumor tissue.

In Eur. J. Nucl. Med. 20 (1993), p. 1138-1140, F. Fazio & G. Paganelli describe that such biotin-avidin systems enable a highly specific labeling of tumors.

It has been the object of the present invention to provide cyclodextrin oligomers which are suitable for the inclusion of pharmaceutically active substances due to their geometry.

The cyclodextrin oligomers according to the invention are two cyclodextrins connected through a spacer at the secondary side, the spacer comprising the unit B which is a rigid and preferably hydrophilic structural element. In one embodiment, the cyclodextrin oligomers have the general formula

CD--X--A--X--B--X--A--X--CD

where

each X is independently selected from --NH--, --O--, --S--, --CO-- or a covalent bond;

each A is an aliphatic C₂ to C₄ residue or a covalent bond;

B is a rigid and preferably hydrophilic structural element; and

each CD represents a cyclodextrin bound through its secondary side.

Preferred compounds for the unit B include melamine, trimesic acid, alizarintetracarboxylic acid or tetraaminoalizarin, and cyclodextrins, such as .alpha.-cyclodextrin, .beta.-cyclodextrin or .gamma.-cyclodextrin, the cyclodextrins being preferably bound through their primary sides.

As examples of the cyclodextrin oligomers according to the invention, there may be mentioned the compounds:

di-.beta.-CD(--NH--(C₈ H₁₆)--NH--);

di-.beta.-CD(terephthaldiamide);

di-.beta.-CD(--NHCO(trimesyl)CONH--);

di-.beta.-CD(--NH--(C₃ H₆)--NHCO--(C₃ H₆)--CONH--(C₃ H₆)--NH--);

di-.beta.-CD(--NH--(C₂ H_{4)--NHCO(trimesyl)CONH--(C2 H.}sub.4)--NH--);

di-.beta.-CD(--NH--(C₂ H_{4)--SH--[6-.alpha.-CD-6]--SH--(C2 H.}sub.4)--NH--);

di-.beta.-CD(--NH--(C₄ H₈)--NHCO--(C₃ H₆)--COHN--(C₄ H₈)--NH--);

di-.beta.-CD(--NH--(C₂ H_{4)--SH-[6-(.beta.,.gamma.)-CD-6]-SH--(C2 H.}sub.4)--NH--); or

di-.beta.-CD(--NH--(C₃ H_{6)--NHCO(trimesyl)COHN--(C3 H.}sub.6)--NH--).

The cyclodextrin oligomers according to the invention can be obtained by the tosylation of the OH groups on the secondary side of cyclodextrins, followed by reaction with short bifunctional spacer groups. The thus obtained functionalized cyclodextrins can be converted to dimers with bifunctional reagents. The corresponding synthetic protocols can be found in A. Ruebner et al. (loc. cit.). Alternatively, carboxylic acid functions may also be introduced into the cyclodextrins by reacting the regiospecific tosylates with amino- or mercaptocarboxylic acids, such as 3-mercaptopropionic acid or 4-aminobutyric acid.

Binding constants can be determined in competition with 6-(p-toluidino)-2-naphthalenesulfonic acid (TNS). The measurement is performed by fluorescence spectroscopy according to Ruebner et al. (loc. cit.).

Preferably, the cyclodextrin oligomer additionally carries at least one and preferably two affinity groups which can interact with molecular target structures.

In principle, said at least one affinity group can have binding capability for a tissue-specific antigen as a molecular target structure. However, according to the polyphasic application route sought by Fazio & Paganelli (loc. cit.), an indirect tissue-specific binding is preferred: The affinity group recognizes a target structure which was previously attached at the desired site of action in a tissue-specific way.

For example, a possible site of action is a tumor. Thus, a tissue- or tumor-specific antibody which carries the target structure for the affinity group of the substance according to the invention can first be introduced into the organism once or several times to become enriched at the site of action, in terms of a polyphasic tumor therapy. Subsequently, a selective enrichment of the pharmaceutically active substance can be achieved at the site of action by the administration of a complex of a pharmaceutically active substance and the substance according to the invention having the affinity group. Then, the pharmaceutically active substance can be released at the site of action. This is effected, for example, by disrupting the cyclodextrin unit(s) at the site of action. Poly- or monoclonal antibodies, but also antibody fragments, such as Fab or F(ab)₂ fragments, and artificial antibodies such as scFv fragments, can be used as said antibodies. Suitable affinity groups and target structures include a wide variety of binding members, such as biotin/avidin, biotin/streptavidin or enzyme/inhibitor systems. Preferred affinity groups include biotinyl residues and digoxin/digoxigenin residues.

Therefore, the invention further relates to a complex of a pharmaceutically active substance and the cyclodextrin oligomers according to the invention. These physical inclusion complexes, which are highly hydrophilic externally, serve to prevent the uptake of the complexed pharmaceutically active substance into body cells for the purpose of reducing or excluding the side-effects of tumor therapeutics, for example. Preferably, the pharmaceutically active substance has a high potential for side-effects and can therefore be employed in its free form only in a limited way. Suitable pharmaceutically active substances include mitotic inhibitors, tumor therapeutics and photo-chemotherapeutics, especially taxol, a taxol derivative, coichicine, colchemide, 3^1,81 -(di-t-butylphenoxy)porphyrin, chlorotrianisene, tamoxifene, vinblastin, vincristin, docetaxel.

If the cyclodextrin oligomers carry an additional affinity group, they can serve for the selective polyphasic application of a pharmaceutically active substance. Surprisingly, the side-effects of a therapy can be reduced by this method. In addition, a considerable dosage reduction of up to a factor of 10,000 is possible.

The invention also relates to medicaments which contain at least one cyclodextrin oligomer according to the invention or at least one complex according to the invention. Such medicaments are useful, in particular, for the treatment of tumor diseases, such as tumors of the bladder, carcinomas of the breast or uterus, esophageal cancers, gastric carcinomas, cephalo -cervical carcinomas, nasopharynx carcinoma, hepatocellular carcinomas, pulmonary carcinomas, lymphomas, melanomas, ovarial carcinomas and prostatic carcinomas.

In a preferred embodiment, the cyclodextrin oligomer is di-.beta.-CD(--NH--(C₄ H₈)--NHCO--(C₃ H₆)--CONH--(C₄ H₈)--NH--), and the pharmaceutically active substance is taxol or a taxol derivative.

The indication, dosage and successes of treatment in tumor diseases are described in: Proceedings ASCO, Vol. 16 (1997) (Denver, Colo., USA).

Due to the cyclodextrins being bonded on the secondary side, the cavities of the cyclodextrins are oriented towards each other. The cyclodextrins are preferably .beta.-cyclodextrins. The distance between the cyclodextrin units is determined by selecting the spacer. Depending on the size of the compound to be enclosed, the spacer is selected such that the distance between the at least two cavities approximately matches the distance between two hydro-phobic groups of the pharmaceutically active substance. Preferred spacings are within a range of about 0.6 to 2 nm, more preferably in a range of about 0.8, 1.0, 1.2, 1.4, 1.6 and 1.8 nm. Such compounds are shown in the following Table.

The spacer between the two cyclodextrin units is rigid, i.e., the spacer has only a small number of bonds which can freely rotate about the binding axis. Preferably, the spacer contains an additional cyclodextrin unit which is preferably bound to the complexing cyclodextrins through its primary side and through short aliphatic spacers.

Such rigid spacers can also be achieved by the use of appropriately functionalized (hetero)aromatics. For example, rigid spacers prevent too high a conformational flexibility, such as twisting of the two cyclodextrins involved in the binding. This results in comparably high binding constants as stated in the above Table. By the use of rigid spacers, affinity constants within a range of greater than 10⁶, preferably greater than 10⁷ and more preferably greater than 10⁸ l/mol are achieved. Thus, rigid spacers are those which achieve correspondingly high stability constants of the complexes together with cyclodextrins involved in the binding and appropriate guest compounds.

The complexes according to the invention can be disrupted, for example, by the action of cyclodextrinase from Klebsiella oxytoca to release the complexed pharmaceutically active substance. Alternatively, functional groups can be contained in the spacers which facilitate the breaking of the bond at the target site.

Claim 1 of 12 Claims

What is claimed is:

1. A cyclodextrin oligomer having the general formula

CD--X--A--X--B--X--A--X--CD

wherein X--A--B--X--A--X is a bifunctional spacer in which:

each X is selected from the group consisting of --NH--, --O--, --S--, --C(=O)-- and a covalent bond;

each A is selected from the group consisting of a C₂ to C₄ alkylidenyl residues and a covalent bond;

B is selected from the group consisting of diradical linkers derived from melamine, 1,3,5-benzenetricarboxylic acid (aka trimesic acid), a tetracarboxylic acid derivative of 1,2-dihydroxyanthraquinone, a tetraamino derivative of 1,2-anthraquinone, .alpha.-cyclodextrin, .beta.-cyclodextrin and .gamma.-cyclodextrin; wherein each said cyclodextrin is bound through two primary carbons; and

each terminal CD is bound through a secondary carbon; and

wherein said cyclodextrin oligomer is optionally substituted with at least one affinity group which interacts with a molecular target structure.

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