Nanovectors and uses

Abstract

The present invention relates to the field of nanovectors for the delivery of active substances in the body, in particular for the treatment of tumours. In particular, the use of these nanovectors makes it possible to improve the pharmacokinetics of the active substances with a more selective delivery, for example in the tumour tissues.

Claims

1. A nanovector for the delivery of an active substance in a human being, wherein said nanovector comprises a nanoparticle on the surface of which the active substance is bonded by physisorption, said nanoparticle is a polysiloxane-based nanoparticle with a mean diameter of less than 10 nm, the active substance is chosen from organic molecules which have a molar mass of between 2% and 40% of the total molar mass of said nanoparticle, and said nanoparticle comprises: a. polysiloxanes, with a weight ratio of silicon of at least 8% of the total weight of the nanoparticle, b. a chelating agent grafted onto the nanoparticle, wherein the chelating agent which is grafted onto the nanoparticle is selected from DOTA, DTPA, EDTA, EGTA, BAPTA, NOTA, DOTAGA and DTPABA or a mixture thereof, and c. optionally, a metal element, said metal element being complexed to the chelating agent.

2. The nanovector according to claim 1, wherein the load content, expressed in milligrams of active substance per gram of nanovector, is greater than 0.5 mg/g.

3. The nanovector according to claim 1, wherein the nanoparticle is of formula (I) below:
Si.sub.n[O].sub.m[OH].sub.o[Ch.sub.1].sub.a[Ch.sub.2].sub.b[Ch.sub.3].sub.c[M.sup.y+].sub.d[D.sup.z+].sub.e[Gf].sub.f(I) in which: n is between 20 and 5000, m is greater than n and less than 4 n, o is between 0 and 2 n, Ch.sub.1, Ch.sub.2 and Ch.sub.3 are chelating agents, which may be identical or different, bonded to the Si atoms of the polysiloxanes by an SiC covalent bond; a, b and c are integers between 0 and n and a+b+c is less than or equal to n, M.sup.y+ and D.sup.z+ are metal cations, which may be identical to or different from one another, with y and z=1 to 6; d and e are integers between 0 and a+b+c, and d+e is less than or equal to a+b+c, Gf is a targeting graft, which may be identical to or different from one another, each bonded to the Si by an SiC bond and resulting from the grafting of a targeting molecule allowing the targeting of the nanoparticle to biological tissues of interest, and f is an integer between 0 and n.

4. The nanovector according to claim 1, wherein the nanovector comprises a metal element chosen from elements with a high atomic number Z.

5. The nanovector according to claim 1, wherein said nanoparticle is a polysiloxane-based nanoparticle with a mean diameter of between 1 and 5 nm, comprising gadolinium complexed to the chelating agent obtained by grafting of DOTAGA onto the nanoparticle.

6. The nanovector according to claim 1, wherein the active substance is chosen from small organic molecules having a molar mass of less than 5000.

7. The nanovector according to claim 1, wherein said nanoparticle comprises a targeting agent covalently grafted to the polysiloxanes and allowing active targeting of biological zones of interest.

8. The nanovector according to claim 1, wherein said active substance is chosen from anti-cancer substances.

9. The nanovector according to claim 8, wherein said anti-cancer substance is chosen from the following substances: actinomycin, all-trans retinoic acid, azacitidine, azathioprine, bleomycin, bortezomib, carboplatin, capecitabine, cisplatin, chlorambucil, cyclophosphamide, cytarabin, daunorubicin, docetaxel, doxifluridine, doxorubicin, epirubicin, epothilone, etoposide, fluorouracil, gemcitabine, hydroxyurea, idarubicin, imatinib, irinotecan, mechlorethamine, mercaptopurine, methotrexate, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, teniposide, tioguanine, topotecan, valrubicin, vemurafenib, vinblastine, vincristine, vindesine, lenalidomide, ibrutinib, abiraterone, erlotinib, everolimus, nilotinib, sunitinib, sorafenib, goserelin, nedaplatin, laboplatin, TATE peptide and heptaplatin, or a mixture thereof.

10. The nanovector according to claim 1, wherein the nanoparticle comprises a chelate of an element with an atomic number greater than 40, having a radiosensitizing effect.

11. The nanovector according to claim 1, wherein the nanoparticle comprises a metal element chelate, said metal element being chosen for use in imaging by magnetic resonance imaging, scans or scintigraphy.

12. The nanovector according to claim 1 wherein said nanoparticle comprises: a. polysiloxanes, with a weight ratio of silicon between 8% and 50% of the total weight of the nanoparticle, b. the chelating agent in a proportion of between 5 and 100 per nanoparticle, and c. optionally, the metal element in a proportion of between 5 and 100 per nanoparticle, said metal element being complexed to the chelating agent.

13. The nanovector according to claim 1, wherein the load content, expressed in milligrams of active substance per gram of nanovector, is between 1 mg/g and 100 mg/g.

14. The nanovector according to claim 1, wherein the nanovector comprises a metal element chosen from elements with a high atomic number Z chosen from gadolinium, bismuth or a mixture thereof.

15. The nanovector according to claim 3, wherein the nanovector comprises a metal element chosen from elements with a high atomic number Z chosen from gadolinium, bismuth or a mixture thereof.

16. An injectable pharmaceutical solution comprising a nanovector according to claim 1, and at least one pharmaceutically acceptable excipient.

17. The injectable pharmaceutical solution according to claim 16, wherein the nanovector comprises an element with a high atomic number Z greater than 40, and wherein said high-Z element is at a concentration between 10 and 200 mM in said solution.

18. The injectable pharmaceutical solution according to claim 16, wherein the active substance of the nanovector is chosen from doxorubicin, cisplatin and the TATE peptide.

19. The injectable pharmaceutical solution according to claim 16, wherein the injectable pharmaceutical solution is directly obtained by means of a method for preparing a nanovector for the delivery of an active substance in a human being, said method comprising mixing two solutions that can be administered in a human being: a. a first solution comprising a nanoparticle, said nanoparticle being a polysiloxane-based nanoparticle having a mean diameter of less than 10 nm, and b. a second solution comprising an active substance or a mixture of active substances chosen from organic molecules, under concentration ratio, pH and temperature conditions which allow an interaction by physisorption of the active substance or mixture of active substances at the surface of said nanoparticle.

20. A method for preparing a nanovector according to claim 1, said method comprising mixing two solutions: a. a first solution comprising a nanoparticle, said nanoparticle being a polysiloxane-based nanoparticle having a mean diameter of less than 10 nm, wherein said nanoparticle comprises polysiloxanes with a weight ratio of silicon of at least 8% of the total weight of the nanoparticle and a chelating agent grafted onto the nanoparticle, wherein the chelating agent which is grafted onto the nanoparticle is selected from DOTA, DTPA, EDTA, EGTA, BAPTA, NOTA, DOTAGA and DTPABA or a mixture thereof, and b. a second solution comprising an active substance chosen from organic molecules having a molecular weight of between 2% and 40% of the molecular weight of said nanoparticle, under concentration ratio, pH and temperature conditions which allow an interaction by physisorption of the active substance at the surface of said nanoparticle.

21. The method according to claim 20, wherein the [active substance in the second solution]:[nanoparticle in the first solution] concentration ratio by weight is greater than 0.5 mg/g.

22. The method according to claim 20 wherein the active substance is chosen from small organic molecules with a molar mass of less than 5000 g.Math.mol.sup.1.

23. The method according to claim 20, wherein the active substances are substance is chosen from one of the following anti-cancer substances: actinomycin, all-trans retinoic acid, azacitidine, azathioprine, bleomycin, bortezomib, carboplatin, capecitabine, cisplatin, chlorambucil, cyclophosphamide, cytarabin, daunorubicin, docetaxel, doxifluridine, doxorubicin, epirubicin, epothilone, etoposide, fluorouracil, gemcitabine, hydroxyurea, idarubicin, imatinib, irinotecan, mechlorethamine, mercaptopurine, methotrexate, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, teniposide, tioguanine, topotecan, valrubicin, vemurafenib, vinblastine, vincristine, vindesine, lenalidomide, ibrutinib, abiraterone, erlotinib, everolimus, nilotinib, sunitinib, sorafenib, goserelin, nedaplatin, laboplatin and heptaplatin, or a mixture thereof.

24. The method according to claim 20, comprising a step of purifying the nanovector obtained after mixing of the solutions, so as to remove the possible active substance that has remained free in solution, and recovering the nanovector comprising the nanoparticle, at the surface of which the active substance is bonded by physisorption.

Description

DESCRIPTION OF THE FIGURES

(1) FIG. 1: General principle of the physisorption on the nanovectors. The active substance is added to the nanoparticles. Step 1: the active substance interacts by physisorption at the surface of the nanoparticles. Step 2: the active substance saturates the sphere of interaction of the nanoparticles. Step 3: the active substance can no longer interact with the surface of the nanoparticles and remains free in solution.

(2) FIG. 2: Curve representing the change in concentration of active species of the subnatant as a function of the concentration of drug placed in the presence of the polysiloxane-based nanoparticles.

(3) FIG. 3: Absorption spectra of the subnatants of Examples 1 and 2.

(4) FIG. 4: Absorption spectra of the subnatants of Examples 3 and 4.

(5) FIG. 5: Absorption values at 497 nm of the subnatants of the solutions of doxorubicin in the presence of AGuIX at 100 mM, as a function of the amount of doxorubicin introduced.

(6) FIG. 6: Absorption spectra of the subnatants of Examples 6 and 7, diluted 20-fold.

(7) FIG. 7: Absorption spectra of the subnatants of Examples 9 and 10, diluted 20-fold.

(8) FIG. 8: Absorption spectrum of the subnatant of Example 8, diluted 20-fold.

(9) FIG. 9: Fluorescence spectrum of the subnatants of Examples 6 and 7, diluted 20-fold.

(10) FIG. 10: Fluorescence spectrum of the subnatants of Examples 9 and 10, diluted 20-fold.

(11) FIG. 11: Fluorescence spectrum of the subnatants of Example 8, diluted 20-fold.

(12) FIG. 12: Absorption values at 280 nm of the subnatants of the solutions of TATE peptide (diluted 20-fold) in the presence of AGuIX at 100 mM, as a function of the amount of TATE peptide introduced.

(13) FIG. 13: Absorption spectrum of the subnatants of Examples 12, 13 and 14 after treatment as described in the examples.

(14) FIG. 14: Absorption spectrum of the subnatants of Examples 15 and 16 after treatment as described in the examples.

(15) FIG. 15: Absorption spectrum of the subnatants of Examples 17 and 18 after treatment as described in the examples.

(16) FIG. 16: Absorption values at 706 nm of the subnatants of the solutions of cisplatin in the presence of AGuIX at 100 mM as a function of the amount of cisplatin introduced. The subnatants were subjected beforehand to a treatment as described in the examples in order to allow the detection of the cisplatin by absorption.

(17) FIG. 17: Absorbance spectra of the subnatants of Examples 18 and 19 after treatment as described in the preceding examples.

(18) FIG. 18: Absorbance spectrum of the subnatants of Examples 18 and 20 after treatment as described in the examples.

EXAMPLES

(19) The examples below make it possible to illustrate the invention but are in no way limiting in nature.

(20) The aim of the various examples presented below is to illustrate the possibility of the polysiloxane nanoparticles acting as transporter of molecules used in chemotherapy. The intended molecules adsorb at the surface of the nanoparticles according to the mechanism proposed in FIG. 1. The examples below also make it possible to determine the drug concentration limit above which the drugs are no longer retained at the surface of the nanoparticle.

(21) During the various examples, the maximum drug load content on polysiloxane nanoparticles was determined, when said particles are present at a concentration corresponding to a concentration used during clinical trials for the nanoparticle in question. Increasing concentrations of active substances were thus brought into contact with the nanoparticles. Solutions are purified by tangential filtration. The molecules which have not be able to adsorb to the surface of the nanoparticles pass through the membrane and are found in the subnatant, where they can be detected by spectroscopic techniques such as UV/Visible absorption or else fluorescence spectroscopy (FIG. 2).

(22) Preparation of a Solution of Polysiloxane-Based Ultrafine Nanoparticles

(23) The solution of polysiloxane-based ultrafine nanoparticles (AGuIX) was synthesized according to the procedure described in the publication G. Le Duc et al., Cancer Nanotechnology, 2014.

(24) A solution of AGuIX at a gadolinium concentration of 10 mM is analysed by DLS with a laser at 633 nm. A number-average hydrodynamic diameter of 3.2 nm is obtained.

Nanovectors for Doxorubicin Delivery

Example 1

(25) 50 mol (Gd.sup.3+) of AGuIX nanoparticles were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM ([Gd.sup.3+]). 2.85 mg of doxorubicin are placed in a 2.5 ml flask. 1.1 ml of ultrapure water are added to the flask, which is stirred until the doxorubicin has completely dissolved. A solution at 2.6 g/l of doxorubicin is then obtained, and is protected from the light with aluminium. 215 l of this solution are then added to the solution of AGuIX, as are 160 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 112 mg/l of doxorubicin is thus obtained.

(26) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out in order to obtain a supernatant of 200 l. The subnatant is analysed by UV-visible analysis. The supernatant is diluted 50-fold and is analysed by UV-visible analysis.

Example 2 (Comparative)

(27) A solution of doxorubicin at 112 mg/l is prepared according to the procedure described in Example 1, the solution of AGuIX being replaced with ultrapure water.

Example 3

(28) 50 mol (Gd.sup.3+) of AGuIX nanoparticles were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM ([Gd.sup.3+]). 2.85 mg of doxorubicin are placed in a 2.5 ml flask. 1.1 ml of ultrapure water are added to the flask, which is stirred until the doxorubicin has completely dissolved. A solution at 2.6 g/l of doxorubicin is then obtained, and is protected from the light with aluminium. 327 l of this solution are then added to the solution of AGuIX, as are 48 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 170 mg/l of doxorubicin is thus obtained.

(29) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out in order to obtain a supernatant of 200 l. The subnatant is analysed by UV-visible analysis. The supernatant is diluted 50-fold and is analysed by UV-visible analysis.

Example 4 (Comparative)

(30) A solution of doxorubicin at 170 mg/l is prepared according to the procedure described in Example 3, the solution of AGuIX being replaced with ultrapure water.

Comparative Results Examples 1/2 and 3/4

(31) FIGS. 3 and 4 represent, respectively, the absorption spectra of the subnatants of the solutions of Examples 1 and 2, and of the solutions of Examples 3 and 4. These data show that the doxorubicin interacts with the AGuIX nanoparticles. Indeed, for the solution containing 100 mM ([Gd.sup.3+]) of AGuIX and 112 mg/l, the doxorubicin is adsorbed at the surface of the nanoparticle and is not detected in the subnatant after tangential filtration, contrary to a solution of doxorubicin alone. For the solution containing 100 mM ([Gd.sup.3+]) of AGuIX and 170 mg/l, a very weak signal is detected by UV/VIS spectrophotometry, indicating that the majority of the doxorubicin is adsorbed at the surface of the nanoparticles and that a small amount passes through the membrane.

(32) A solution obtained by the procedure of Example 3 (doxorubicin at 170 mg/l and AGuIX at 100 mM [Gd.sup.3+]) is diluted 50-fold and analysed by DLS with a laser at 633 nm. A number-average hydrodynamic diameter of 3.7 nm is obtained, which is greater than the diameter of 3.2 nm obtained for the AGuIX nanoparticles, indicating a surface interaction of the nanoparticles with the doxorubicin.

(33) For a solution of AGuIX nanoparticles at 100 mM ([Gd.sup.3+]) corresponding to 100 g/l of nanoparticles, a retention of doxorubicin is observed up to a minimum concentration of 112 mg/l, which corresponds to a load content by weight greater than 1.12 mg/g of nanoparticles (FIG. 5).

(34) Nanovectors for Delivery of TATE Peptide

Example 6

(35) 50 mol (Gd.sup.3+) of AGuIX were dispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM ([Gd.sup.3+]). 14.94 mg of tyr3-octreotate (TATE) peptide are placed in a 2.5 ml flask. 498 l of ultrapure water are added to the flask, which is stirred until the peptide has completely dissolved. A solution containing 30 g/l of peptide is then obtained. 48 l of this solution are then added to the solution of AGuIX, as are 328 l of ultrapure water. The flask is stirred for 30 minutes. A solution containing 100 mM of gadolinium and 2.90 g/l of peptide is thus obtained.

(36) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 200 l. The subnatant is analysed by UV-visible analysis and fluorometry after 20-fold dilution.

Example 7 (Comparative)

(37) A solution of TATE peptide at 2.90 g/l is prepared according to the procedure described in Example 6, the solution of AGuIX being replaced with ultrapure water.

Example 8

(38) 50 mol (Gd.sup.3+) of the AGuIX nanoparticles were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM ([Gd.sup.3+]). 6.1 mg of tyr3-octreotate (TATE) peptide are placed in a 2.5 ml flask. 203.3 l of ultrapure water are added to the flask, which is stirred until the peptide has completely dissolved. A solution containing 30 g/l of peptide is then obtained. 97 l of this solution are then added to the solution of AGuIX, as are 279 l of ultrapure water. The flask is stirred for 30 minutes. A solution containing 100 mM of gadolinium and 5.80 g/l of peptide is thus obtained.

(39) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 320 l. The subnatant is analysed by UV-visible analysis (20-fold dilution) and fluorometry (40-fold dilution).

Example 9

(40) 50 mol (Gd.sup.3+) of the AGuIX nanoparticles were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM ([Gd.sup.3+]). 14.94 mg of tyr3-octreotate (TATE) peptide are placed in a 2.5 ml flask. 498 l of ultrapure water are added to the flask, which is stirred until the peptide has completely dissolved. A solution containing 30 g/l of peptide is then obtained. 193 l of this solution are then added to the solution of AGuIX, as are 182 l of ultrapure water. The flask is stirred for 30 minutes. A solution containing 100 mM of gadolinium and 11.60 g/l of peptide is thus obtained.

(41) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 200 l. The subnatant is analysed by UV-visible analysis and fluorometry after 20-fold dilution.

Example 10 (Comparative)

(42) A solution of TATE peptide at 11.60 g/l is prepared according to the procedure described in Example 5, the solution of AGuIX being replaced with ultrapure water.

Example 11

(43) 50 mol (Gd.sup.3+) of the AGuIX nanoparticles were redispersed in 250 l of ultrapure water in order to obtain a solution at 200 mM ([Gd.sup.3+]). 0.6 mg of tyr3-octreotate (TATE) peptide are placed in a 2.5 ml flask. 20 l of ultrapure water are added to the flask, which is stirred until the peptide has completely dissolved. A solution containing 30 g/l of peptide is then obtained. 20 l of this solution are then added to the solution of AGuIX, as are 230 l of ultrapure water. The flask is stirred for 30 minutes. A solution containing 100 mM of gadolinium and 1.20 g/l of peptide is thus obtained.

(44) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 320 l. The subnatant is analysed by UV-visible analysis (20-fold dilution) or fluorometry (40-fold dilution).

Results of Examples 6 to 10

(45) FIGS. 6, 7 and 8 represent, respectively, the absorption spectra of the 20-fold diluted subnatants of the solutions of Examples 6 and 7, of the solutions of Examples 9 and 10 and of the solution of Example 8. These data show that the TATE peptide adsorbs at the surface of the nanoparticles up to a concentration of approximately 2 g.Math.L.sup.1 (FIG. 12). Indeed, before this limiting concentration, the TATE peptide is not detected by UV/VIS spectrophotometry in the subnatants of the solutions purified by tangential filtration.

(46) FIGS. 9, 10 and 11 represent, respectively, the fluorescence spectra of the 20-fold diluted subnatants of Examples 6 and 7, of the Examples 9 and 10 and of Example 8. In the same way as for the detection by UV/VIS spectrophotometry, these data show that the TATE peptide interacts with the AGuIX nanoparticles.

(47) A solution obtained by the procedure of Example 8 (TATE at 2.90 g/l) is diluted 10-fold and analysed by DLS with a laser at 633 nm.

(48) A number-average hydrodynamic diameter of 3.4 nm is obtained, which is greater than the diameter of 3.2 nm obtained for the AGuIX nanoparticles, indicating a surface interaction of the nanoparticles with the TATE peptide.

(49) For a solution of AGuIX nanoparticles at 100 mM ([Gd.sup.3+]) corresponding to 100 g/l of nanoparticles, a retention of the TATE peptide is observed up to a minimum concentration of 2 g/l, which corresponds to a load content by weight of greater than 20 mg/g of nanoparticles (FIG. 12).

Nanovectors for Delivery of Cisplatin

Example 12

(50) 50 mol (Gd.sup.3+) of AGuIX were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM [Gd.sup.3+]. 3.1 mg of cisplatin are placed in a 2.5 ml flask. 1.2 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat to 40 C. until it is completely dissolved. A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 24 l of this solution are then added to the solution of AGuIX, as are 351 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 120 mg/l of cisplatin is thus obtained.

(51) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 160 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this solution are added to 200 l of buffer and 100 l of ODPA. The resulting solution is heated at 100 C. for 15 min. Once the reaction is finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV-visible analysis.

Example 13

(52) 50 mol (Gd.sup.3+) of AGuIX were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM [Gd.sup.3+]. 3.1 mg of cisplatin are placed in a 2.5 ml flask. 1.2 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat to 40 C. until it is completely dissolved.

(53) A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 36 l of this solution are then added to the solution of AGuIX, as are 339 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 180 mg/l of cisplatin is thus obtained.

(54) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 160 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this solution are added to 200 l of buffer and 100 l of ODPA. The resulting solution is heated at 100 C. for 15 min. Once the reaction is finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV/VIS spectrophotometry.

Example 14

(55) 50 mol (Gd.sup.3+) of AGuIX were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM [Gd.sup.3+]. 3.1 mg of cisplatin are placed in a 2.5 ml flask. 1.2 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat to 40 C. until it is completely dissolved. A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 72 l of this solution are then added to the solution of AGuIX, as are 303 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 360 mg/l of cisplatin is thus obtained.

(56) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 160 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this solution are added to 200 l of buffer and 100 l of ODPA. The resulting solution is heated at 100 C. for 15 min. Once the reaction is finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV-visible analysis.

Example 15

(57) 50 mol (Gd.sup.3+) of AGuIX were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM [Gd.sup.3+]. 2.8 mg of cisplatin are placed in a 2.5 ml flask. 1.1 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat to 40 C. until it is completely dissolved. A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 142 l of this solution are then added to the solution of AGuIX, as are 233 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 720 mg/l of cisplatin is thus obtained.

(58) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 140 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this solution are added to 200 l of buffer and 100 l of ODPA. The resulting solution is heated at 100 C. for 15 min. Once the reaction is finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV-visible analysis.

Example 16 (Comparative)

(59) A solution of cisplatin at 720 mg/l is prepared according to the procedure described in Example 15, the solution of AGuIX being replaced with ultrapure water.

Example 17

(60) 50 mol (Gd.sup.3+) of AGuIX were redispersed in 125 l of ultrapure water in order to obtain a solution at 400 mM [Gd.sup.3+]. 2.8 mg of cisplatin are placed in a 2.5 ml flask. 1.1 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat to 40 C. until it is completely dissolved. A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 229 l of this solution are then added to the solution of AGuIX, as are 146 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of gadolinium and 1160 mg/l of cisplatin is thus obtained.

(61) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 160 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this solution are added to 200 l of buffer and 100 l of ODPA. The resulting solution is heated at 100 C. for 15 min.

(62) Once the reaction is finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV-visible analysis.

Results (Examples 12, 13, 14, 15 and 17)

(63) Examples 12, 13, 14, 15 and 17 (cisplatin at 120-180-360-720-1160 mg/l) are analysed by DLS (samples diluted 10-fold) with a laser at 633 nm. The respective number-average hydrodynamic diameters are: 3.8, 3.7, 3.8, 3.4, 3.7 nm. They are greater than the diameter of 3.2 nm obtained for the AGuIX nanoparticles, indicating a surface interaction of the nanoparticles with the cisplatin.

Example 18 (Comparative)

(64) A solution of cisplatin at 1160 mg/l is prepared according to the procedure described in Example 15, the solution of AGuIX being replaced with ultrapure water.

Results of Examples 15/16 and 17/18

(65) FIG. 13 represents the absorbance of the subnatants of Examples 12, 13 and 14 after treatments as described in the examples.

(66) FIGS. 14 and 15 represent, respectively, the absorption spectra of the subnatants of the solutions of Examples 15 and 16, and of the solutions of Examples 17 and 18. These data show that the cisplatin adsorbs at the surface of the nanoparticles up to a concentration of approximately 240 mg.Math.L.sup.1 (FIG. 16). Indeed, before this limiting concentration, the signal detected in the ODPA-treated subnatants, by UV/VIS spectrophotometry at 706 nm, is not modified.

(67) For a solution of AGuIX nanoparticles at 100 mM ([Gd.sup.3+]) corresponding to 100 g/l of nanoparticles, a retention of the cisplatin is observed up to a minimum concentration of 240 mg/l, which corresponds to a load content by weight of greater than 2.4 mg/g of nanoparticles (FIG. 16).

Example 19

(68) Nanoparticles based on polysiloxane and on free chelates for cisplatin delivery.

(69) For the synthesis of these nanoparticles, 6.187 ml (26.17 mmol) of APTES are added to 90 ml of diethylene glycol. The solution is stirred for 1 h at ambient temperature before 10 g (17.45 mmol) of DOTAGA anhydride are added. The solution is left to stir for 5 days. At the end of this, 7.952 ml of TEOS (34.90 mmol) are added to the solution, which is stirred for 1 hour. 900 ml of ultrapure water are then added, before heating at 50 C. with stirring for 18 h. The solution is then concentrated to 200 ml on a Vivaflow cassette with membranes having a cut-off threshold of 5 kDa. The pH is adjusted to 2 by adding hydrochloric acid. The solution is purified by a factor of 50 by Vivaflow, before being neutralized to pH 7.4 by controlled addition of 1 M sodium hydroxide. The solution is filtered and then lyophilised. After redispersion in water, the nanoparticles have a hydrodynamic diameter of 5.2 nm.

(70) 50 mol (DOTAGA) of silica nanoparticles (62.5 mg) were redispersed in 141 l of ultrapure water in order to obtain a solution at 354 mM of DOTAGA and 443 mg/l. 3 mg of cisplatin are placed in a 2.5 ml flask. 1.2 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat at 40 C. until it has completely dissolved. A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 229 l of this solution are then added to the solution of silica nanoparticles, as are 130 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of free chelate (125 g/l of nanoparticles) and 1160 mg/l of cisplatin is thus obtained.

(71) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 200 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this solution are added to 200 l of buffer and 100 l of ODPA. This new solution is heated at 100 C. for 15 min. Once the reaction has finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV-visible analysis.

(72) FIG. 17 represents the absorption spectra of the subnatants of the solutions Examples 18 and 19. A weaker signal can be observed in the subnatants for the cisplatin added to the solution of silica nanoparticles. The signal at 706 nm of the two UV spectra makes it possible to estimate a retention of a cisplatin concentration of 260 mg/l for a solution of nanoparticles at 125 g/l, corresponding to a load content by weight of 2.1 mg/g of nanoparticles.

Example 20

(73) Possibility of varying the load content by modifying the surface of the nanoparticles by chelation of bismuth ions.

(74) The nanoparticles described in Example 19 are dispersed in water (283 mg, 227 mol of DOTAGA) in order to have a DOTAGA concentration of approximately 200 mM. The pH of the solution is adjusted to 5.5 by adding NaOH. 817 l of a solution of BiCl.sub.3 at 250 mM in 6 M HCl are slowly added in 3 additions with stirring at a temperature of 70 C. to accelerate the complexation. Between each addition, the pH is readjusted to 5.5 by slowly adding a 10 M sodium hydroxide solution. The solution is then heated to 80 C. for 1 hour after the final addition. At the end of this, ultrapure water is added to reach a chelate concentration of 100 mM at a pH of 5.5. The solution is then heated at 80 C. for 18 h. The excess Bi.sup.3+ is removed by tangential filtration, then the solution is neutralized to reach a pH of 7 by adding sodium hydroxide, before filtration on a 0.2 m membrane and lyophilisation. After redispersion in the water, the nanoparticles have a hydrodynamic diameter of 6.0 nm.

(75) 30 mol of AGuIX@DOTA@Bi (Bi.sup.3+) (67.8 mg) were redispersed in 75 l of ultrapure water in order to obtain a solution at 400 mM (904 g/L of nanoparticles). 3 mg of cisplatin are placed in a 2.5 ml flask. 1.2 ml of ultrapure water are added to the flask, which is stirred. Since cisplatin is not very soluble at ambient temperature, it is necessary to heat at 40 C. until it is completely dissolved. A solution containing 2.5 g/l of cisplatin is then obtained, and is protected from the light with aluminium. 118 l of this solution are then added to the solution of AGuIX@DOTA@Bi, as are 107 l of ultrapure water. The flask is stirred for 30 minutes in the dark. A solution containing 100 mM of bismuth (226 g/l of nanoparticles) and 1000 mg/l of cisplatin is thus obtained.

(76) This solution is placed in a 3 kDa Vivaspin, and a tangential filtration cycle is carried out so as to obtain a supernatant of 80 l. The subnatant is analysed by UV-visible analysis. The cisplatin is detectable by UV/VIS absorption at a wavelength of 706 nm after reaction with ODPA. For the reaction with cisplatin, a solution of ODPA at 1.4 mg/ml and a phosphate buffer (pH 6.8) are prepared. The subnatant is diluted 5-fold. 140 l of this concentration are added to 200 l of buffer and 100 l of ODPA. This new solution is heated at 100 C. for 15 min. Once the reaction has finished and the temperature has returned to ambient temperature, 560 l of DMF are added. The final solution is filtered and then analysed by UV-visible analysis.

Results of Examples 18 and 20

(77) FIG. 18 represents the absorption spectra of the subnatants of the solutions of Examples 18 and 20.

(78) As shown in FIG. 18, the chelation of bismuth at the surface of the nanoparticles leads to a non-retention of the cisplatin at the surface of the nanoparticles, proving that changes regarding the number of metal elements chelated at the surface of the nanoparticles make it possible to vary the load content of the nanoparticles.