WO2004037275A1

WO2004037275A1 - Bile acid conjugates and particles formed therewith

Info

Publication number: WO2004037275A1
Application number: PCT/EP2003/010580
Authority: WO
Inventors: Gerhard PÜTZ
Original assignee: Wieland, Heinrich
Priority date: 2002-10-25
Filing date: 2003-09-23
Publication date: 2004-05-06
Also published as: AU2003275978A1; DE10249890A1

Abstract

Disclosed is a conjugate comprising a structural lipid unit, a structural bile acid unit or bile salt unit, and an optional structural linker unit, the structural lipid unit and the structural bile acid unit or bile salt unit being covalently linked, optionally via the intermediate structural linker unit. Such conjugates allow very useful particles, particularly liposomes or similar vesicular particles, to be formed along with additional components as desired. Such conjugates and particles comprising or containing said conjugates in a bonded manner are particularly suitable for therapeutic applications, above all for target-specific and especially liver-specific therapy.

Description

Bile acid conjugates and particles formed with them

The present invention relates to conjugates containing a bile acid or bile acid salt component. The invention further relates to particles, in particular liposomes or similar vesicular particles, which comprise such conjugates or which are formed, inter alia, with these conjugates. Such conjugates and particles which comprise or contain the conjugates are particularly well suited for therapeutic applications, in particular for target-specific and especially liver-specific therapy.

In the prior art, target- or cell type-specific liposomes have been constructed in which an internalizing receptor serves as a cellular target protein for the binding of the liposomes on the cell surface. This approach enables the liposomes bound in this way to enter the cell via receptor-mediated endocytosis. Geho et. al. in US Pat. No. 4,603,044 describe such a targeted drug delivery system (trageted drug delivery) which is to be bound to hepatobiliary receptors. A vesicle is linked to a conjugate consisting of a connector molecule, a bridge molecule and a target molecule. The target molecule is said to have an intended receptor molecule Hepatocytes interact. An iminodicarboxyl acid residue as a connector molecule, a “bridging ion”, especially a chromium complex as a bridge molecule, and a target molecule that is identical to the connector molecule are proposed for the structure, wherein other target molecules for binding to the desired receptor are also conceivable.

In U.S. Patent 6,063,400 Geho et al. such a conjugate for liposomes, which contain a biogenic amine as therapeutic agent and also a so-called sequestering agent (especially nucleotide derivatives) to enable targeted binding to a receptor.

The object of the present invention is to provide an improved system for targeted therapy.

The object is achieved according to the invention by a conjugate according to claim 1, a particle according to claim 12, a use according to claim 23 and a kit according to claim 24. Preferred embodiments are specified in the subclaims.

The conjugate according to the invention has the following components:

(a) a lipid structural unit and

(b) a bile acid or bile acid salt structural unit,

where components (a) and (b) are covalent, optionally via a

(c) linker structural unit.

Without the linker component, the conjugate consequently has the structure (a) - (b), with this the preferred structure (a) - (c) - (b). According to the invention, it was found that such a conjugate with components (a) and (b) and optionally (c) is capable of hepatocyte-specific proteins transporting bile salt, such as Na ^+, which mainly transports cholyltaurine (CT) / Cholyltaurine co-transporting polypeptide (Ntcp) and various organic anion transporting polypeptides (Oatp 1-x), which was confirmed by inhibition of bile acid transport by the special conjugate according to the invention. The observed inhibition of bile salt uptake is incomplete even when the hepatocytes are covered with the appropriate vesicles, which creates an important, favorable prerequisite for the applicability of the conjugate according to the invention for therapy, since complete inhibition of the transport of the bile acid or the bile salt increases a cholea stasis and thus damage the patient.

The invention is explained in more detail below on the basis of preferred embodiments and with reference to the accompanying drawings.

1 shows an example of a conjugate according to an embodiment of the invention, in which a lipid structural unit is linked via a linker structural unit to a structural unit of a bile acid salt;

2A schematically shows the possible structure of a bile salt-bearing liposome according to an embodiment of the invention, and

FIG. 2B shows an enlarged detail in area X of FIG. 2A;

FIG. 3 shows the inhibition of the uptake of natural bile salts in hepatocytes by particles carrying bile salts using the example of liposomes.

FIG. 4 shows a typical organ distribution when using particles carrying the conjugate according to the invention using the example of liposomes in comparison to liposomes without the conjugate; 5A to C show the binding of the inventive contact jugate-bearing particles using the example of liposomes, in which a fluorescent model substance had been incorporated, on hepatocyte primary cultures, and FIG. 5D shows a comparison with liposomes without the conjugate, in each case as a fluorescence-microscopic image of the hepatocytes.

The term "lipid" of the structural unit (a) of the conjugate means any fat molecule or a fat-like substance which is preferably found in natural cells. With lipid structure units occurring in natural cells, immunological defense reactions can advantageously be suppressed better. Phospholipids, especially phosphoglycerides, are particularly advantageous. A phospho-diester linkage can conveniently be used for the covalent connection to the structural unit (b) or - in the preferred case the intermediate linker - (c). The use of phosphoglycerides as a lipid structural unit in the conjugate is very advantageous, since when the particles described in more detail below, in particular with liposomes, are built up, the conjugate is stably anchored via a phosphoglyceride with long-chain acyl residues. Consequently, the lipid structural unit (a) is preferably constructed as a diacyl phosphoglyceride, where the acyl residue can be, for example, a saturated fatty acid residue such as lauryl, myristyl, palmitinyl, stearinyl, arachinyl or an unsaturated fatty acid residue such as palmitoleyl, oleyl, linolyl, lindenyl and arachidonyl. For better integration into preferred liposome particles, C12 to C18 acyl residues, in particular C16 to C18 acyl residues, are particularly suitable. The acyl radicals can be selected independently of one another.

A bile acid or a bile acid salt serves as the structural unit (b), the salt form being, for example, the Na, the K, the ammonium salt, the salt of an amino acid or an organic amine or the like. Unless otherwise stated, the terms bile acid and bile acid salt are used synonymously below. The structural unit (b) has the steroid characteristic of bile acid Basic structure, for example in the form of a 5β-cholanic acid derivative. The cholanic acid derivative is preferably modified on the 24C carboxylate with nitrogen-containing bases such as glycine or taurine. Modification with taurine is particularly advantageous because the -S0 ₃ H group forms a permanently charged sulfate group at the end of the bile salt opposite the lipid structure unit after dissociation and thereby an undesired insertion of the hydrophobic cholane framework area into a particle, especially into a liposome membrane - bran, difficult or prevented. Moreover, the non-hepatocyte-specific insertion of the hydrophobic framework region into the membranes of cells can be made more difficult or prevented by the negative charge.

In principle, a functional group in the 24C radical and one or more hydroxyl groups in the 3-, 7- or 12-position of the cholane skeleton can be considered for linking the structural units (a) and (b). A suitable functionalization can also be introduced at other points for linking. In view of a stronger specific binding to hepatocytes, it is preferred to form the link to the lipid structural unit (a) or the linker (c) structurally as far as possible from the 24C residue of the bile acid steroid structure. The linkage is therefore preferably via the 3-hydroxy position of the steroid structure of bile acid. The 7- and 12-positions of the steroid skeleton of bile acid may or may not be hydroxylated, or they may be modified by other structural units. Particularly suitable bile acids are taurocholic acid and especially lithotaurocholic acid and their salts.

It was also found that the specific binding to hepatocytes was significantly stronger when the linkage to the 3-position of the steroid structure of the bile acid or salt was in the β configuration.

The structural unit (b) can be directly covalently linked to the structural unit (a) may be connected, for example via an ester linkage such as a phosphorus diester linkage with the phosphoglyceride described above. For easier synthesis and for the introduction of a molecular spacer so that the structural units (a) and (b) can better perform their respective functions, there is preferably a linker structural unit (c) between the structural units (a) and (b). This linker molecule can itself consist of one or more structural units. Suitable structural units are, for example, alone or in combination, a polyalkylene glycol group with n polarization units, where n is an integer such as di-, tri- or tetramethylene glycol or polyethylene glycol, a dicarboxylate group such as succinyl and the like.

A specific example of a conjugate according to the invention is shown in FIG. 1 in the form of a schematic structure. The respective structural units (a) - (c) - (b) are indicated. The structural representation only serves to illustrate a possible structure, but is not to be understood as restrictive, since according to the invention any variations are possible within the scope of the respective structural unit (a), (b) and, if appropriate, (c).

The above-described conjugate according to the invention with the essential structural units (a) and (b), which are preferably linked to the linker (c), are particularly suitable for use in the medical field, in particular for therapeutic processes.

In a further object of the present invention, a particle comprises the special conjugate described above. Suitable particles are microparticulate carriers or transport vehicles of suitable size, which are preferably designed such that therapeutic agents are bound or included on and / or in the particle. The particle can be of natural or artificial origin or an artificial modification of natural vehicles. In this regard, can be referred to particles that are known from conventional drug delivery systems. Suitable are, for example, liposomes, microspheres, nanoparticles, niosomes, polymer particles, lipoproteins, virus particles (viruses, virus envelopes and other, modified virus particles, the virulence of which has been removed or modified in some other way) and certain cell types such as blood cell subtypes, such as Erythrocytes and lymphocytes. The functional component (b) is easily integrated into the hydrophobic areas of the particle via the lipid component (a) and anchored there in a stable manner.

The particle composition suitably contains 0.1-50% by weight, preferably 0.5-5% by weight and in particular 0.75-2% by weight, based on the total amount of the particle components, e.g. the liposome mixture, the conjugate according to the invention.

Particularly preferred particle carriers for the conjugate according to the invention are liposomes. In combination with the lipid-containing conjugate of the invention, in particular in combination with the preferred diacylphosphoglycerides described above, a particularly stable anchoring of the functional component (b) to the liposomes is possible. Furthermore, the production of such liposome particles is particularly simple since the conjugate according to the invention can easily be used as a starting material and only needs to be mixed with other lipid components suitable for liposomes, such as normal phosphoglycerides, cholesterol, lecithin or the like.

In the case of the liposome particles, in addition to the conjugate according to the invention, which is suitably used in a proportion of 0.1 to 50 mol%, preferably 0.5 to 5 mol%, the liposome mixture can contain up to 99.9 mol% of normal phospholipids and if desired, contain 0.1 to 80 mol% of cholesterol and optionally further constituents. Suitable normal phospholipids can be selected from the following group: Phospholipids with long-chain saturated fatty acid residues of C16 and above, in particular C16-C18 such as DPPC and DSPC, preferably in a proportion of 20 to 40 mol%, or of long-chain unsaturated fatty acid residues such as POPC and DNPC, preferably in a proportion of 10 to 30 mol%, since these favor the storage of the liposomes at 4 ° C without transition into the liquid crystalline phase; and phospholipids with medium saturated chain lengths (C8-C14) such as DMPC, preferably in a proportion of 20 to 40 mol%, or with medium unsaturated chain lengths such as D yPC, preferably in a proportion of 20 to 90 mol% and in particular from 40 to 80 mol% because these favor a fusion with cell membranes. The addition of cholesterol to the liposome mixture on the one hand reduces the fluidity of the membrane and on the other hand increases the width of the phase transition between the liquid-crystalline and the gel phase. Cholesterol further reduces the permeability of the liposome membrane and increases the length of time that the hydrophilic compounds present in the interior of the liposomes are dissolved in the liposomes. Therefore, the liposome mixture preferably contains from 10 to 90 mol%, more preferably from 20 to 60 mol%, of cholesterol. While the main components of the liposome mixture are neutral in total, such as the uncharged cholesterol or the zwitterionic diacylphosphocholine, one or more negatively charged phospholipids can additionally be added to the liposome mixture. As a result of the Coulomb repulsion against the negatively charged bile acid residue, in particular the taurine-modified bile acid, the structural unit (b) of the conjugate contained in the liposome can protrude outwards and bind better to a transport protein. A negative total charge of the liposome also leads to a slower opsonization in vivo and thus to an increased length of time of the liposomes in the blood. Suitable negatively charged lipids are, for example, phosphatidylglycerol, DSPG, DMPG and glutyryl-DSPG, since it ensures a particularly long half-life of the liposomes in the blood. The negatively charged lipid component of the liposomes can preferably be present in a proportion of 1 to 50 mol%, more preferably 4 to 15 Mol% are added.

FIG. 2A shows a schematic representation of the possible structure of a liposome carrying bile salt, and FIG. 2B shows section X of FIG. 2A enlarged. The hydrophilic residues in the bile salt structural unit (b) of the conjugate according to the invention (3-litho-cholyltaurine-distearoyl-phosphoethanolamine) point, for example, inwards and outwards of the double-walled liposome membrane, while the lipid structural unit (a) of the conjugate is stable in the liposomes - membrane is integrated and thus anchored. In practical use, it is preferred that the bile salt structural unit (b) points essentially outwards.

The size of the particle according to the invention can generally be in the nanometer range up to the micrometer range. The average particle size is suitably in the range from 10 nm to 10 μm, more preferably from 40 to 200 nm and in particular from 100 to 150 nm.

In addition to the conjugate according to the invention, the particle preferably further comprises a fusion peptide known per se, e.g. those described in WO 99/39742 A, such as the 25-amino acid N-terminal sequence of the surfactant associated protein B, in order to promote the fusion with the cellular membrane after the binding of the particle to the transporter of the hepatocyte and thus the content of the particle easier to discharge into the cell plasma. The fusion peptide can be tailored via tailored amino acid sequences, e.g. via hydrophobic amino acids, or suitable modifications, e.g. via covalent links, firmly connected to the particle or anchored there, e.g. in WO 99/39742 A and by Pecheur et al. (Biochemistry 36, 3773-3781 (1997) and Biochemistry 37, 2361-2371 (1998).

In addition to the conjugate according to the invention and optionally the fusion peptide, the particles, in particular the liposomes, can preferably also be a sterically stabilized, ie a so-called Have “stealth component”, for example lipid-bound polyethylene glycol (PEG) or lipid-bound glutaryl, in order to prolong the half-life of the particles against the immune system in circulation in vivo.

The particle according to the invention can preferably comprise a therapeutic agent or a pharmaceutical composition or a diagnostic agent for use in the medical field. Suitable therapeutic agents are, for example, cytostatics, antibiotics, antivirals, antimycotics, gene therapy substances such as DNAs or antisense molecules, antibodies, cytokines, interferons, radionuclides and the like. Suitable diagnostic agents are, for example, contrast agents, radioactive isotropes, NMR imaging agents and the like.

The conjugate according to the invention and the particle according to the invention comprising it are therefore particularly well suited for use in therapy and diagnostics in medicine, in particular in hepatocyte-related diseases such as hepatitis or liver cell carcinoma.

In a further object of the invention, the conjugate according to the invention and a particle and / or a therapeutic or diagnostic agent are combined in separate complements of a kit. The kit preferably comprises, in separate compliments, a liposome preparation in which the conjugate according to the invention is enclosed in the liposomes, and separately the therapeutic or diagnostic agent. The liposome preparation can then be loaded with the therapeutic or diagnostic agent before use.

The invention is explained in more detail below on the basis of examples which are not to be understood as limiting. Examples

Abbreviations:

CT cholyltaurine

DCCI dicyclohexylcarbodiimide

DMPG l, 2-0-dimyristoyl-sn-glycero-3-phospho-rac- (1-glycerol)

DMPC 1, 2-0-dimyristoyl-sn-glycero-3-phosphocholine

DmyPC 1, 2-0-dimyristoleoyl-sn-glycero-3-phosphocholine

DNPC l, 2-0-dinervonoyl-sn-glycero-3-phosphocholine

DPPC 1, 2-O-dipalmitoyl-sn-glycero-3-phosphocholine

DSPE 1, 2-0-distearoyl-sn-glycero-3-phosphoethanolamine

DSPG l, 2-0-distearoyl-sn-glycero-3-phospho-rac- (1-glycerol)

Glutaryl-DSPE N-glutaryl-1, 2-O-distearoyl-sn-glycero-3 - phosphoethanolamine

LCT lithocholyl taurine

LCT-DSPE disodium-3- (1, 2-0-distearoyl-sn-glycero-3-phospho-

2'-ethanolamidosuccinyl) ethoxy-cholan-24-oyl-2-aminoethanesulfonate

Ph ₂ -DiOC18 3,3'-dioctadecyl-5, 5'-diphenyloxacarbocyanine chloride

PC phosphatidylcholine

POPC 1-O-palmitoyl-2-O-oleoyl-sn-glycero-1-phosphocholine

UDCT Ursodeoxycholyltaurine

Example 1: Preparation of a conjugate according to the invention

(1) Preparation of 3 - (2-hydroxyethoxy) -5ß-cholan-24-acid Literature: G. Wess et al. , Tetrahedron Letters 33, 195-198 (1992)

(1-1) Synthesis of 3α-hydroxy-5ß-cholan-24-acidic acid methyl ester

15 g of 3α-hydroxy-5β-cholan-24-acid (39.8 mmol) were dissolved in 150 ml of absolute methanol and mixed with 70 μl of concentrated hydrochloric acid. After heating under reflux for 4 hours, the solvent was removed in vacuo and the crude product obtained was purified by recrystallization from MeOH / water. Yield: 4.3 g (36.6 mmol, 92%).

(1-2) Synthesis of 3α- (2-propenoxy) -5ß-cholan-24-acidic acid methyl ester

Under an argon atmosphere, 5 g (12.8 mmol) of 3α-hydroxy-5β-cholan-24-acidic acid methyl ester and 6.5 ml of diisopropyl-ethyl-amine dissolved in 15 ml of anhydrous DMF and heated to boiling temperature. 15 ml of allyl bromide (173 mmol) was added dropwise over a period of 8 h. The excess allyl bromide, HBr formed and the diisopropyl-ethyl-amine were then removed by distillation at atmospheric pressure, and the solvent was removed by distillation under high vacuum. The cooled residue was taken up in ether and purified by column chromatography in the mobile phase cyclohexane / ethyl acetate 30/1 (v / v). Yield: 4.2 g (9.8 mmol, 77%).

(1-3) Synthesis of 3oc- (2-oxoethoxy) -5ß-cholan-24-acidic acid methyl ester

To a solution of 3.5 g of 3α- (2-propene) -5ß-cholan-24-acidic acid methyl ester (8.1 mmol) and 56 mg of osmium tetraoxide (0.21 mmol) in 50 ml of dioxane and 10 ml of water were added within 4.5 g of sodium periodate (21 mmol) were added in small portions for 45 min. After stirring for 10 hours at room temperature, the salt which had hitherto precipitated was filtered off and the solvent was removed in vacuo. The cleaning was carried out chromatographically in the mobile phase mixture cyclohexane / ethyl acetate 5/1 (v / v). Yield: 2.8 g (6.5 mmol, 80%).

(1-4) Synthesis of 3α- (2-hydroxyethoxy) -5ß-cholan-24-acidic acid methyl ester

3.5 g of methyl 3α-hydroxyethoxy-5β-cholan-24-acid (8.1 mmol) were dissolved in 40 ml of a 4/1 (v / v) THF / water mixture and mixed with 250 mg of sodium borohydride ( 6.61 mmol) added. After stirring for 14 hours, 0.1 N HCl was added until no more hydrogen evolution was observed. The organic components of the solution were extracted with ether, the combined ether phases were dried over sodium sulfate and the solvent was then removed on a rotary evaporator. The crude product obtained was flash-chromatographed in the eluent Cy- clohexane / ethyl acetate 4: 1 (v / v) cleaned. Yield: 2.2 g (5.1 mmol, 63%).

(1-5) Synthesis of 3α- (2-hydroxyethoxy) -5ß-cholan-24-acid

The saponification of 1 g of methyl 3α- (2-hydroxyethoxy) -5ß-cholan-24-acid (2.3 mmol) was carried out by refluxing for four hours in a solution of 260 mg of potassium hydroxide (4.6 mmol) in 20 ml Methanol and 5 ml water. After cooling, this solution was mixed with 250 ml of dilute hydrochloric acid and extracted 3 times with 200 ml of ether each time. The combined organic phases were dried over sodium sulfate and the solvent was removed in vacuo. To purify the product obtained, chromatography was carried out using a mixture of cyclohexane / ethyl acetate / acetic acid in a ratio of 70/30/1 (v / v / v) as the eluent. Yield: 0.89 g (2.1 mmol, 92%).

(2) Synthesis of 3ß- (2-hydroxyethoxy) -5ß-cholan-24-acid

2.6 ml of methanesulfonic acid chloride (32 mmol) were slowly added dropwise to a solution of 10 g of 3α-hydroxy-5β-cholan-24-acid (26.6 mmol) in 10 ml of dried and freshly distilled pyridine under an argon atmosphere. During this time and for a further 30 minutes, the reaction mixture was cooled to 0 ° C. The mixture was then stirred at room temperature for a further 2 h. The methanesulfonic acid ester formed was dissolved in a mixture of 40 ml of anhydrous glycol and 10 ml of freshly distilled pyridine without purification and stirred at 100 ° C. for 2 hours. After cooling, this solution was poured into 500 ml of 0.1 N HCl and the organic components were extracted three times with 500 ml of ether each time. The combined organic phases were dried over sodium sulfate and the solvent was removed on a rotary evaporator. The purification of the crude product was carried out using the chromatographic method Mobile solvent cyclohexane / ethyl acetate / acetic acid 70/30/1 (v / v / v) carried out. Yield: 3.8 g (9.0 mmol, 38%).

(3) Preparation of sodium 3- (2-hydroxyethoxy) -5ß-cholan-24-oyl) -2 'aminoethanesulfonate

(3-1) Synthesis of 3- (2-acetyl-ethoxy) -5ß-cholan-24-acids

4 ml of acetic anhydride were added to a solution of 1 g of 3- (2-hydroxyethoxy) -5β-cholan-24-acid (2.4 mmol) in 15 ml of dried and freshly distilled pyridine and the reaction mixture was stirred at room temperature for 1 hour , In order to hydrolyze excess acetic anhydride and any mixed anhydride formed, 8 ml of water were added. Pyridine, water and acetic acid were then removed azeotropically with toluene in vacuo. The product was purified by chromatography under elevated pressure in the mobile phase cyclohexane / ethyl acetate / acetic acid 80/20/1 (v / v / v).

3 - (2-acetyl-ethoxy) -5ß-cholan-24-acid:

Yield: 870 mg (1.9 mmol, 79%); Melting point: 107 ° C

3ß- (2-acetylethoxy) -5ß-cholan-24-acid:

Yield: 980 mg (2.1 mmol, 88%); Melting point: 109 ° C

(3-2) Synthesis of sodium 3- (2-hydroxyethoxy) -5ß-cholan-24-oyl-2 'aminoethanesulfonates

660 mg (1.4 mmol) of 3- (2-acetylethoxy) -5ß-cholan-24-acid, 430 mg (2.1 mmol) of DCCI and 240 mg (2.1 mmol) of N were dissolved in 10 ml of dried dioxane under a protective gas atmosphere -Hydroxysuccinimide dissolved, and the reaction mixture was stirred at room temperature for 12 h. The N, N-dicyclohexane urea which precipitated out during the reaction was filtered off and the clear filtrate was mixed with a solution of 263 mg of taurine (2.1 mmol) and 177 mg of sodium hydrogen carbonate (2.1 mmol) in 5 ml of water. The- This reaction mixture was stirred again at room temperature for 12 h. The N, N-dicyclohexane urea which had precipitated out during the reaction was again filtered off, and dioxane and water were removed under reduced pressure. The acetyl protective group was split off by heating under reflux in 50 ml of 1N NaOH for 2 hours. The product was determined by adsorption chromatography on Serdolit® PAD

1 removed from the reaction mixture and purified by column chromatography at elevated pressure using CHCl ₃ / MeOH 3/1 (v / v).

Sodium (3α- (2-hydroxyethoxy) -5ß-cholan-24-oyl) -2 ^{^} - aminoethanesulfonate:

Yield: 550 mg (1.0 mmol, 72%); Melting point: 223 ° C.

Sodium (3ß- (2-hydroxyethoxy) -5ß-cholan-24-oyl) -2-aminoethanesulfonate:

Yield: 650 mg (1.2 mmol, 85%); Melting point: 214 ° C.

(4) Preparation of the dinatrrum-3- (1, 2-O-distearoyl-sn-glycero-3-phospho-2 ^' ' -ethanolamidosuccinyl) -ethoxy-5β-cholan-24-oyl-2-aminoethanesulfonate

(4-1) Synthesis of sodium 3-succinylethoxy-5β-cholan-24-oyl

2 ^{^} aminoethanesulfonates

To a solution of 200 mg (0.38 mmol) of sodium 3- (2-hydroxyethoxy) -5β-cholan-24-oyl-2 ^{^} -aminoethanesulfonate in 5 ml of dried and freshly distilled pyridine was added 120 mg (1.2 mmol ) Succinic anhydride and stirred at room temperature for 12 h. Excess succinic anhydride and any mixed anhydride formed were hydrolyzed by stirring for 1 hour after adding 2 ml of water. After the reaction, pyridine and water were removed azeotropically with toluene on a rotary evaporator. The crude product obtained was purified by column chromatography in CHCl ₃ / MeOH 2/1 (v / v). Sodium 3α- (succinyl-ethoxy) -5ß-cholan-24-oyl-2 ^{^} - aminoethanesulfonate: Yield: 170 mg (0.27 mmol, 71%); Melting point: 178 ° C.

Sodium 3ß- (succinyl-ethoxy) -5ß-cholan-24-oyl-2-aminoethanesulfonate:

Yield: 190 mg (0.30 mmol, 79%); Melting point: 176 ° C.

(4-2) Synthesis of disodium 3- (1,2-0-distearoyl-sn-glycero-3-phospho-2 "-ethanolamidosuccinyl) -ethoxy-5β-cholan-24-oyl-2-aminoethanesulfonate (Fig. 1)

A solution of 250 mg (0.39 mmol) of sodium 3- (2-ethoxy) -5ß-cholan-24-oyl-2 ^{^} -aminoethanesulfonate in 3 ml of anhydrous DMF was treated under an argon atmosphere with 116 mg of DCCI (0.58 mmol ) and 65 mg (0.58 mmol) of N-hydroxysuccinimide were added and the mixture was stirred at room temperature for 16 h. The precipitate was filtered off and washed with 2 ml of cold DMF. The activated bile salt derivative was precipitated by adding 50 ml of ether, the precipitate was separated from the supernatant after a short centrifugation and dissolved in a mixture of 10 ml of CHCl ₃ , 5 ml of MeOH and 200 μl of water.

300 mg of DSPE (0.4 mmol) and 400 μl of 1N NaOH (0.4 mmol) were added to this solution and the reaction mixture was then stirred at room temperature for 2 h. After the reaction had ended, the solvent was removed in vacuo and the product obtained was purified by means of column chromatography with the mobile phase CHCl ₃ / MeOH / H ₂ 0 9/3 / 0.1 (v / v / v).

Disodium 3α- (1, 2-0-distearoyl-sn-glycero-3-phospho-2 - ethanolamidosuccinyl) -ethoxy-5ß-cholan-24-oyl-2 "- aminoethanesulfonate:

Yield: 170 mg (0.12 mmol, 31%); Melting point: 211 ° C. R _f values: 0.08 CHCl ₃ / MeOH / H ₂ 0 9/3 / 0.1 (v / v / v)

0.10 CHCl ₃ / MeOH 2/1 (v / v) ^ - MR: δ = 0.69 (s, 3H, CH ₃ -18); 0.90 (t, J = 8.6H, 2 x H ₃ C-CH ₂ -); 0.95 (s, 3H, CH ₃ -19); 0.95 (d, J = 7 Hz, 3H, CH ₃ - 21); 2.32 (dd, J = 7.4H, 2 X -CH ₂ -CH ₂ -CH ₂ -0-CO-); 2.56 (t, J = 7, 2H, -CO-CH ₂ -CH ₂ -CO-0-); 2.69 (t, J = 7.2H, -NH-CO-CH ₂ -CH ₂ -C0-); 3.01 (t, J = 7 Hz, 2H, -CH ₂ -S0 ₃ ^~ ); 3.35

(m, 1H, CH-3); 3.44 (m, 2H, -0-CH ₂ -CH ₂ -NH-); 3.61 (t, J = 7 Hz, 2H, -HN-CH ₂ -CH ₂ -S0 ₃ Na); 3.70 (t, 2H, J = 6 Hz, -CH ₂ -0-CH-); 3.81 - 4.18 (mb, 6H); 4.21 (t, 2H, J = 6 Hz, -CO-0-CH ₂ -); 5.23 (m, -0-CH ₂ -HCOH-CH ₂ -0-);

(Solvent: CDCl ₃ / CD ₃ OD / D ₂ 0 2/1 / 0.1 (v / v / v))

Mass spectrum: m / z = 1424 (M + Na) ⁺ , 1402 (M + H) ⁺ , 1380 (M- Na + 2H) ⁺ , 1378 (M-Na) ^" , 677 (M-2Na) ^2"

Disodium-3ß- (1, 2-0-distearoyl-sn-glycero-3-phospho-2 '- ethanolamidosuccinyl) -ethoxy-5ß-cholan-24-oyl-2 - aminoethanesulfonate:

Yield: 220 mg (0.16 mmol, 41%); Melting point: 206 ° C.

R _f values: 0.08 CHCl ₃ / MeOH / H ₂ 0 9/3 / 0.1 (v / v / v) 0.03 CHCl ₃ / MeOH / H ₂ 0 8/2 / 0.1 (v / v / v)

^ Η NMR: 5 = 0.68 (s, 3H, CH ₃ -I8); 0.92 (t, J = 8.6H, 2 XH ₃ C-CH ₂ -); 0.96 (d, J = 7 Hz, 3H, CH ₃ -21); 0.97 (s, 3H, CH ₃ -

19); 2.35 (dd, J = 7.4H, 2 X -CH ₂ -CH ₂ -CH ₂ -0-C0-); 2.56 (t, J = 7, 2H, -C0-CH ₂ -CH ₂ -C0-0-); 2.68 (t, J = 7, 2H, -NH-CO-CH ₂ -CH ₂ -CO-); 3.01 (t, J = 7 Hz, 2H, -CH ₂ -S0 ₃ ^~ ); 3.44 (m, 2H, -0-CH ₂ -CH ₂ -NH-); 3.58 (t, J = 7 Hz, 2H, -HN-CH ₂ -CH ₂ -S0 ₃ Na); 3.62 (t, 2H, J = 6 Hz, -CH ₂ -0-CH-); 3.67 (m, 1H, CH-3); 3.92 - 4.21 (mb, 6H); 4.21 (t, 2H, J = 6 Hz, -CO-O-CH ₂ -); 5.27 (m, -0-CH ₂ -HCOH-CH ₂ -0-); (Solvent: CDC1 ₃ / CD ₃ 0D / D ₂ 0 2/1 / 0.1 (v / v / v))

Mass spectrum: m / z = 1424 (M + Na) ⁺ , 1402 (M + H) ⁺ , 1380 (M-

Na + 2H) ⁺ , 1378 (M-Na) ^" , 677 (M-2Na) ^2"

Example 2:

Production of particles according to the invention using the example of liposomes

Liposomes were produced in a modified form using the extrusion method of MacDonald et al. , Biochim. Biophys. Acta. 1061, 297-303. (1991).

The individual lipids, which should be contained in the liposome membrane, were in a defined concentration in chloroform (POPC, DPPC, DNPC, DMPC, DMyPC, cholesterol) or in a mixture of chloroform, methanol and dist. Dissolved water in the ratio 1/1 / 0.02 (v / v / v) (DSPG, DMPG, 3-LCT-DSPE). These stock solutions were stored at -20 ° C. For the production of liposomes of the desired lipid composition, the corresponding aliquots of the lipid stock solutions were transferred to a 25 ml round-bottomed flask. The lipid composition of the liposomes used in mol% is given in Table 1.

Table 1. Lipid composition of the liposomes used in molar

The resulting lipid mixture was freed from the solvent in vacuo and then dried in an oil pump vacuum for 45 min. The following buffer (pH 7.4) was used for liposome preparation (liposome buffer):

NaCl 118 mM

KC1 4.74 rtiM

KH ₂ P0 ₄ 0.59 mM

Na ₂ HP0 ₄ 0.59 mM

HEPES 10mM NaHCO "15mM

MgCl ₂ 1.18 mM

The composition of the buffer used corresponded to that of the perfusion buffer used for hepatocyte isolation. No glucose was used to increase the shelf life of the liposomes. Before use, the liposome buffer was filtered through a membrane with a pore diameter of 220 μm. In order to resuspend the lipids, the desired amount of liposome buffer was added to the dried lipid mixture and the mixture was stirred for 15 minutes at 40 ° C. and 50 rpm using a rotary evaporator. By adding 10-20 glass beads (710 - 1018 microns) the resuspen- lipids are accelerated considerably. The amount of liposome buffer added was chosen so that the suspension obtained contained 10-40 mg lipid / ml. The lipid suspension was centrifuged at 1000 xg for 30 seconds and then left at room temperature for 2 hours. Finally, the resulting multilamellar vesicles were pressed 13 times through a polycarbonate membrane (LFM-200, Milsch equipment) with a pore diameter of 200 nm using a LiposoFast ^® extrusion apparatus (Milsch Equipment, Laudenbach, Germany) and then 21 times through two superimposed polycarbonate membranes (LFM-100, milk equipment) with a pore diameter of 100 nm. The liposomes were stored at 4 ° C and used no longer than 7 days after their preparation.

The size of the liposomes produced was determined by means of PCS (photon correlation spectroscopy) (C. Washington, 1992, in M.H. Rubinstein, ed., Ellis Horwood, Ltd., London, England.). The results of mean diameters (measured from 9 individual preparations each, the mean values and the standard deviation are given) for the bile salt-bearing liposomes and the control liposomes are shown in Table 2.

Table 2 ..

The liposomes produced with an average diameter of approximately 120 nm can be used very well both in vitro and in vivo for the investigation of the binding of bile salt-bearing liposomes to hepatocytes. The possible structure for the bile salt-bearing liposome is shown in FIG. 2A to illustrate the size relationships (the size shown should correspond to a liposome diameter of 100 nm). The liposome membrane 1 has so-called bile salt layers 2. 2B shows a modeled, enlarged section of the membrane of such a bile salt-bearing liposome.

Example 3:

Inhibition of bile salt transport in hepatocytes by the particles carrying the conjugate according to the invention

For this investigation, the uptake of radioactively labeled substances in freshly isolated hepatocytes was determined using a centrifugal filtration technique through a layer of silicone oil (Klingenberg, M. & Pfaff, E., Methods Enzymol. 10, 680-684). Control liposomes or liposomes carrying 3-ß-LCT-DSPE in concentrations of 0.5, 2 and 5 mM liposomal PC were used to determine the uptake rate in the presence of liposomes. Three series of measurements were carried out for each liposome type and each liposome concentration. In the experiments with liposomes, appropriate amounts of liposome stock solution were added to the CT solution and the incubation batches containing CT and liposomes were heated to 37 ° C. for at least 15 minutes before the cells were added. By adding 400 μl each of the cell stock solution, which has a concentration of approximately 2.5 × 10 ⁶ cells / ml

(5 mg / ml), the measurement of the CT recording was started. For a period of 60 s, 100 μl were removed from the incubation batch every 10 s and transferred to a centrifuge tube which had previously been filled with 10 μl 0.77 M perchloric acid and 150 μl of a silicone oil mixture (AR 20: AR 200 = 1: 1

(v / v) was filled without bubbles. The recording was made by Centrifugal filtration stopped (Microfuge® B, Beckman Instruments, Munich, Germany), the cells being transferred from the aqueous medium through the silicone oil layer into the perchloric acid solution and denatured there. After completion of the measurement, the centrifuge tubes were frozen in liquid nitrogen and severed at the phase boundary between perchloric acid and silicone oil. The tips, in which the denatured cells were located, were transferred individually into a scintillation vial and with 250 μl of a 1: 2-

Mix (v / v) of Biolute® S (Zinsser Analytics) and xylene added.

To determine the total radioactivity used, 50 μl were taken twice from each batch and transferred to liquid scintillation vessels. To determine the radioactivity, the samples were treated in exactly the same way as the tips which contained the denatured cells. The protein concentration in the uptake tests was determined by determining the protein concentration in the cell stock solution six times.

The measured values were evaluated according to the following equation:

B D

In this equation:

J: flow rate [nmoles / (min-mg protein)]

A: slope of the regression line [dpm / min]; from 100 μl sample

B: total radioactivity in approach [dpm]

C: amount of substrate [nmoles]

D: Amount of protein in 100 μl of the incubation batch [mg / 100 μl]

At a concentration of 2 mM liposomal PC and simultaneous addition of liposomes and CT, a significantly significant bile salt was mediated in freshly isolated hepatocytes Inhibition of CT transport can be determined. In freshly isolated hepatocytes, liposomes bearing 3-ß-LCT-DSPE showed significantly more pronounced interactions with the hepatocytes than liposomes bearing 3-ß-LCT-DSPE. A typical result is shown in FIG. 3. Both the uptake of radioactive CT in the absence and in the presence of liposomes increases linearly with time in the observed interval. Control liposomes have practically no influence on the transport speed of CT, while liposomes bearing 3-α-LCT-DSPE slightly inhibit the absorption of CT, liposomes bearing 3-β-LCT-DSPE significantly inhibit the absorption of CT.

The results illustrate a specific binding of the bile salt-bearing liposomes to bile salt-transporting proteins of the hepatocytes. Uptake of the liposomes modified according to the invention via a receptor-mediated endocytosis was practically not observed. A lysosomal pathway can thus be prevented or at least strongly suppressed.

Example 4:

Long-lasting particles carrying bile salt using the example of liposomes

(1) Manufacturing

The long-lived liposomes were prepared as described in Example 2, but using the lipid composition given in Table 3.

Table 3 Lipid composition of long-lived liposomes in mole

a: available from Avanti Polar Lipids, Ine Alabaster,

Alabama, USA b: obtained according to Example 1

(2) Investigation of organ distribution of long-lived liposomes in vivo

Liposomes labeled with [ ³ H] cholesteryl hexadecyl ether were used to study the organ distribution of long-lived liposomes. Male albino rats of the Wistar S 300 strain (approx. 200 g) were anesthetized by intraperitoneal injection of a 2% Nembutal ^® solution (1.5 ml / kg body weight). The amount of long-lived liposomes injected was calculated so that the PC concentration in the serum of the rats was 2 mM liposomal PC. The corresponding amount of stock liposome solution was mixed with 1% by volume of a sterile 125 mM CaCl ₂ solution and injected into the vein of the hind leg over a period of about 1 min using a cannula. After 90 minutes, the rats' abdominal cavity was opened and the kidneys and spleen were removed. A blood sample was taken from the heart and placed in a prepared 1.5 ml reaction vessel containing EDTA to prevent coagulation. To separate the erythrocytes, the blood sample was centrifuged at 2000 xg for 2 min. 3 aliquots of 200 μl each were transferred from the supernatant to 20 ml liquid scintillation vials. The liver was perfused with 10 ml of the cell buffer indicated below and then removed. Kidneys and spleen were each treated with 10 ml of ice-cold cell buffer, the liver was mixed with 15 ml of ice-cold cell buffer, and the organs were then homogenized. The total volume of the homogenate was determined and 3 aliquots of 200 μl each were transferred to 20 ml liquid scintillation vials. In each of the 20 ml liquid scintillation vials 1 ml of Biolute S ^® (tin) was added and the samples were fed to the liquid scintillation measurement as described.

Cell buffer:

NaCl 118 mM

KC1 4.74 mM

KH ₂ P0 ₄ 0.59 mM

Na ₂ HP0 ₄ 0.59 mM

HEPES 10 mM

NaHCO 4.15 mM

MgCl ₂ 1.18 mM

D-glucose 5.5 mM

CaCl ₂ 125 mM

The result of organ distribution after 90 minutes of infusion is shown in FIG. 4. The long-lived, bile salt-bearing liposomes (hatched columns) are much more specifically present in the liver than the control liposomes (black column). As in the control, the spleen and kidney are practically not affected.

(3) Enrichment of the liposomes according to the invention on the hepatocyte surface

Liposomes containing 0.3 mol% of the fluorescent dye Ph ₂ -DiOC18 (available from Molecular Probes Europe, Leiden, Holland) were used to investigate the accumulation of long-lived liposomes on liver hepatocytes.

The fluorescence intensities of the liposome preparations were determined by fluorescence spectroscopy, and suspensions with control liposomes and bile salt-bearing liposomes with the same fluorescence intensity were always used for the studies using confocal laser scanning microscopy. The excitation of the Ph ₂ -DiOC18-carrying liposomes was carried out using an argon laser at a wavelength of 488 nm The fluorescence signal was detected at a wavelength of 505 nm.

The hepatocytes were cultivated with collagen type

1 coated slides with a diameter of 30 mm, which were placed in 35 mm culture dishes. The slides supporting the cell lawn were coated on the edge with silicone and a temperature-controlled flow chamber was placed on the slides, which had an inlet and an outlet.

The cell culture medium located above the cells was then removed as completely as possible, and 150 μl of a liposome suspension according to the invention prepared as described in Example 2 and containing 2 mM liposomal PC were added to the cells. After incubating the cells at 37 ° C. for 5 minutes, the liposome suspension was aspirated as completely as possible, and the cells were carefully washed 3 times with 150 μl of cell buffer each. Then 150 ul cell culture medium was added to the cells and the flow chamber was inserted into the confocal laser microscope. The image was taken with a confocal laser scanning microscope (LSM 510 UV; ZEISS, Jena, Germany; recording speed: 2.24 μs / pixel; pixel size in the x and y direction: 0.2 - 0.4 μm; distance the cutting planes in the z direction: 0.05 μm to 0.15 μ; setting the pinhole so that the half-maximum z resolution was between 0.5 and 1 μm at full width)

For longer incubation, the hepatocytes were left in the culture dish for cultivation and carefully 3 times with each

2 ml of cell buffer rinsed. 2 ml of a liposome suspension according to the invention containing 2 mM liposomal PC were then added, and the hepatocytes were cultured in the incubator under the usual conditions for a further 6 hours. The liposome suspensions were then removed and the hepatocytes were washed very carefully 3 times with 2 ml of cell buffer. The slides were inserted into the slide as described Flow chamber introduced and viewed under the confocal laser microscope as described above.

5 shows the results of the binding of liposome aggregates to the hepatocyte surface, FIG. 5A showing the binding of bile salt-bearing liposomes after 5 minutes, FIG. 5B showing the binding of bile salt-bearing liposomes after 6 hours, FIG. 5C an enlarged section from FIG. 5B and 5D shows the comparison with control liposomes after 6 h. Fluorescent liposome aggregates appear in the form of brightly fluorescent dots (see arrows in FIGS. 5B and 5C). The specific binding to hepatocytes is clearly visible (cf. FIGS. 5B and C compared to FIG. 5D).

(4) Localization of the liposomes according to the invention in the liver

The liposomes were infused analogously to that described in Example 4. After perfusion of the liver, small pieces of the individual liver lobes were removed with a scalpel. The removed pieces of liver were placed on a thin cork disc and immediately frozen in liquid N ₂ . Ultrathin sections of the liver tissue were made from the frozen pieces of liver using a microtome. These were viewed under the fluorescence microscope.

In contrast to control liposomes, long-lasting bile salt-bearing liposomes showed a clear fluorescence intensity in large parts of the liver tissue 90 minutes after injection. Long-lived control liposomes, on the other hand, showed sporadic fluorescence intensity in only a few places. The strong accumulation of the liposomes constructed according to the invention along the sinusoids of the liver shows that the liposomes constructed according to the invention also mediate a bile salt-mediated interaction with hepatocytes in vivo.

Example 5 Fusion of liposomes loaded with a model substance. - hepatocytes in primary culture

In order to facilitate or accelerate the fusion of liposomes with hepatocytes, liposomes were used which contained a suitable fusion peptide (SBP25) (WO 99/39742 A). The SBP25 peptide was chemically synthesized, purified by high-performance liquid chromatography and has the following amino acid sequence: Ac-Phe-Pro-Ile-Pro-Leu-Pro-Tyr-Cys-Trp-Leu-Ala-Arg-Ala- Leu-Ile-Lys-Arg-Ile-Gln-Ala-Met-Ile-Pro-Lys-Gly-NH ₂ (HPLC purity 87%).

The liposomes were prepared as described in Example 2 by extrusion, using the lipid composition given in Table 4. SBP25 was dissolved in methanol, and 0.04 μmole SBP25 per μmol phospholipid was added to the respective mixture of lipids dissolved in organic solvents to prepare SBP25-bearing liposomes. The organic solvent was then removed. The following buffer (pH 7.6) was used for the resuspension of the lipids:

NaCl 110 mM

KC1 4.74 mM

KH ₂ P0 ₄ 0.59 mM

Na ₂ HP0 ₄ 0.59 mM

HEPES 20 mM

NaHCO 4.15 mM

MgCl ₂ 1.18 mM

In this buffer, 25 mg / ml of dextran, which had been derivatized with fluorescein (available from Molecular Probes, Inc .; the average molecular weight of the dextran was 10,000 g / mol, so that the concentration of the fluorescein molecules bound to dextran was: 2.5-5 mM fluorescein).

Table 4 Composition of the liposomes used for the studies on the fusion of liposomes with hepatocytes in mol%

The non-encapsulated lipid was separated using gel permeation chromatography. A chromatography column with a diameter of 15 mm and a length of 55 mm, which was filled with Sephadex S-500 (Pharmacia), was used for gel permeation chromatography. The liposome suspensions were brought to a volume of 1 ml, and the subsequent separation of the liposomes and the non-encapsulated fluorescent dextran was carried out under hydrostatic pressure.

For the fusion of the modified liposomes with the hepatocytes in primary culture, the liposome suspensions contained 2 mM liposomal PC. After 2 h, 5 h or 20 h, the liposome suspensions were removed, the cells were washed and then viewed under a fluorescence microscope.

After 2, 5 and 20 h the liposomes were removed from the cells and the hepatocytes were observed under the fluorescence microscope. After 2 h no fluorescent staining of the hepatocytes was observed. After 5 h, SBP25-bearing liposomes and SBP25 and bile salt-bearing liposomes showed a slight fluorescence staining of the hepatocytes. After 20 h, SBP25-bearing liposomes and SBP25 and bile salt-bearing liposomes, in contrast to control liposomes, showed a clear fluorescent staining of the cytosol of the hepatocytes. The presence of SBP25 and bile salt-bearing liposomes resulted in a significantly more intense staining of the cytoplasm of the hepatocytes than the presence of SBP25-bearing liposomes. Bile salt mediated binding of the bile salt and SBP25-bearing liposomes is thus able to accelerate the fusion process mediated by SBP25.

Claims

Expectations

1. Conjugate with the following components:

(a) a lipid structural unit,

(b) a bile acid or bile salt structural unit, the structural units (a) and (b) being covalent, optionally via an intermediate component

(c) a linker structural unit.

2. The conjugate according to claim 1, wherein the lipid structural unit (a) is derived from fat or fat-like substances occurring in cells.

3. The conjugate according to claim 1 or 2, wherein the lipid structural unit (a) is a phosphoglyceride which is attached to (b) or

(c) is covalently linked to a phosphodiester linkage,

4. Conjugate according to one of the preceding claims, wherein the lipid structural unit (a) contains a C12 to C18 fatty acid residue.

5. The conjugate according to claim 3, wherein the phosphoglyceride has a diacyl radical, the acyl radicals independently of one another having a C12 to C18 chain.

6. Conjugate according to one of the preceding claims, wherein the bile acid or bile salt structural unit (b) is a 5β-cholanic acid derivative.

7. The conjugate according to claim 1 or 6, wherein the structural unit (b) is derivatized with a tauryl group.

8. Conjugate according to claim 1, 6 or 7, wherein the linkage of the structural unit (b) to (a) or (c) via the 3- Position of the steroid structure of the bile acid or bile salt is done.

9. The conjugate according to claim 8, wherein the link to the

3-position of the steroid framework is in the ß configuration.

10. Conjugate according to one of the preceding claims, wherein the linker structural unit (c) is a polyalkylene glycol and / or an alkyl diacarboxylate group.

11. Conjugate according to one of the preceding claims for use in the medical field.

12. Particle comprising a conjugate according to any one of claims 1 to 11.

13. Particles according to claim 12 with the property of being able to bind proteins which transport bile acid or bile salt.

14. Particles according to claim 12 or 13, wherein the amount of conjugate is 0.5-5% by weight, based on the total amount of particles.

15. Particles according to one of claims 12 to 14, constructed in the form of a liposome which carries the conjugate.

16. Particles according to claim 15, wherein the liposome contains, in addition to the conjugate, phospholipids which are selected from the following group with respective proportions:

20 to 40 mol% of phospholipids with long-chain saturated fatty acids of C16 and above,

10 to 30 mol% of phospholipids with long-chain unsaturated fatty acids from C16 and above, and 10 to 40 mol% of phospholipids with medium-chain saturated fatty acids from C8 to C14, and 40 to 90 mole% phospholipids with medium chain unsaturated fatty acids from C8 to C14.

17. Particles according to claim 15 or 16, wherein the liposome further contains cholesterol.

18. Particles according to claim 17, wherein the content of cholesterol is 20 to 60 mol%.

19. Particles according to any one of claims 15 to 17, wherein the liposome further contains a negatively charged lipid component.

20. Particles according to claim 19, wherein the content of the negatively charged lipid component is 5 to 15 mol%.

21. Particle according to one of claims 12 to 20, wherein the average particle size of the particle is in the range from 10n to 500nm.

22. Particle according to one of claims 12 to 21, wherein it additionally comprises a fusion peptide.

23. Particle according to one of claims 12 to 22, wherein it additionally comprises a stealth component.

24. Particle according to one of claims 12 to 23, wherein it additionally comprises a therapeutic or a diagnostic agent.

25. Use of a conjugate according to one of claims 1 to 11 or a particle according to one of claims 12 to 24 in medical therapy or diagnostics.

26. Kit with the following components:

- a particle according to any one of claims 12 to 23 and

- a therapeutic or diagnostic agent.