Method for synthesizing zeolite nanoparticles in a saline phosphate buffer and associated nanoparticles, suspensions and pharmaceutical composition

Abstract

The present invention relates to a method for synthesizing nanoparticles consisting of or comprising at least one zeolite nanocrystal according to which: - a first composition / solution 1 containing an aluminum source and a source of an ion of an alkali metal M, in particular K, is prepared; a second composition / solution 2 comprising a silicon source and a source of an ion of an alkali metal M, in particular K, is prepared, said compositions / solutions 1 and 2 being free of any organic structuring agent; - the two compositions / solutions 1 and 2 are mixed; - the mixture is crystallized; and - said nanoparticles thus formed are optionally separated. According to the invention, said first composition / solution 1 and said second composition / solution 2 are both constituted of said source and of the aqueous saline phosphate buffer. The present invention also relates to colloidal suspensions of the nanoparticles obtained and to a pharmaceutical composition containing said nanoparticles.

Description

[0001] PROCESS FOR THE SYNTHESIS OF ZEOLITE NANOPARTICLES IN A SALINE PHOSPHATE BUFFER - ASSOCIATED NANOPARTICLES, SUSPENSIONS AND PHARMACEUTICAL COMPOSITION

[0002] Technical field

[0003] The present invention relates to a process for synthesizing zeolite nanoparticles, suspensions of these nanoparticles, a pharmaceutical composition containing these nanoparticles and the nanoparticles obtained according to the process of the invention.

[0004] These suspensions and nanoparticles can be used in particular in the medical field, in particular for the treatment of hypoxic solid tumors.

[0005] State of the art

[0006] Zeolites are a class of crystalline microporous materials. They are formed by a three-dimensional arrangement of TO4 (T= Si, Al or other heteroatoms) bonded to each other by sharing an oxygen atom. In aluminosilicate zeolites, the presence of AIG ions 5 ' gives the framework negative charges which must be compensated by so-called compensating cations (alkaline or alkaline-earth), such as Na + , K + , That 2+ , Sr 2+in order to maintain electrical neutrality. The vacant space is defined by the aluminosilicate framework formed by these three-dimensional chains. The vacant spaces in the tetrahedral crystal lattice are connected to each other and occupied by alkali ions and water molecules. Dehydration of zeolites removes the water molecules present. Zeolites also have a microporous network formed by cavities, channels connecting the cavities and pores connecting the channels or cavities with the outside. Zeolites can be natural or synthetic. There are currently more than 250 types of zeolites.

[0007] WO 2019 / 030200 A1 describes FAU and EMT type zeolite nanocrystals for which sodium ions have been replaced by Fe or Gd ions and which can be charged with gaseous dioxygen. These nanocrystals can be used for the diagnosis and treatment of brain tumors, including glioblastoma. The zeolite nanocrystals in these two documents are synthesized in water, which limits their medical uses. Indeed, over the long term and at high concentrations, these nanocrystals are likely to cause allergy-type problems. The publication by Kharchenko et al entitled "Formation of copper nanoparticles in LTL nanosized zeolite: kinetics study" and published in The journal of physical chemistry C, volume 120, no. 46 on November 5, 2016 describes a method for synthesizing LTL type nanoparticles. The composition of the mixture used for crystallization is as follows: 5K2O: 10SiO2: 0.5 AI2O3: 200 H2O.This solution is an aqueous solution and does not contain PBS.

[0008] The publication by F. Mayer et al entitled “nanozeolite-LTL with Gd IH deposited in the large and EU 111 in the small cavities as a magnetic resonance optical imaging probe" and published in the journal Chemistry - A European Journal, John Wiley & Sons, Inc, DE volume 20, n°12 on February 12, 2014 does not describe the synthesis of LTL-type nanozeolites. This document indicates that it is possible to migrate ions present in the nanozeolite to the outside and replace them.

[0009] An aim of the present invention is therefore to propose a new pharmaceutical composition containing zeolite nanoparticles which can be used in the diagnosis and / or treatment of certain pathologies, in particular hypoxic solid tumors and in particular brain tumors.

[0010] Another aim of the invention is to propose a new method for synthesizing such nanoparticles.

[0011] Another aim of the invention is to provide colloidal suspensions of these nanoparticles.

[0012] Another aim of the invention is to provide zeolite nanoparticles which have reduced toxicity and / or have an improved oxygen adsorption and / or release capacity.

[0013] Summary of the invention

[0014] A first aspect of the invention relates to a method for synthesizing nanoparticles consisting of or comprising at least one zeolite nanocrystal according to which:

[0015] - a first composition / solution 1 is prepared containing a source of aluminum and a source of an ion of an alkali metal M, in particular K;

[0016] - a second composition / composition / solution 2 is prepared comprising a source of silicon and a source of an ion of an alkali metal M, in particular K, said compositions / suspensions 1 and 2 being free of any organic structuring agent; - the compositions / composition / solutions 1 and 2 are mixed and the mixture is left stirring,

[0017] - the mixture is crystallized at a temperature greater than or equal to 50C; and

[0018] - the said nanoparticles thus formed are possibly separated,

[0019] Characteristically, according to the invention, said first composition / solution 1 and said second composition / solution 2 are each constituted by said source and an aqueous buffer (PBS) constituted by water and 137 mM / L of NaCl, 2.7 mM / L of KCl, 10 mM / L of Na2HPO4, 1.76 mM / L of KH2PO4 and having a pH equal to 7.2 and said mixture of compositions / solutions 1 and 2 has the following molar composition (I): x1 M20: yAI2O3: 10SiO2: z PBS (I) in which 2.8 <x1 < 8,8,

[0020] 0.3 <y< 1 ,5 et 40 <z< 300.

[0021] The method of the invention is simple to implement and makes it possible to obtain a stable colloidal suspension of LTL-type nanoparticles in PBS.

[0022] Preferably, the compositions are suspensions.

[0023] The above-mentioned compositions / solutions 1 and 2 consist of PBS, the silicon or aluminum source and the alkali metal source.

[0024] Depending on the method of implementation, it is sometimes necessary to heat the mixture of PBS buffer and other reagents to obtain a clear suspension.

[0025] Regarding temperatures, the first composition / solution 1 can be prepared at room temperature. The second composition / solution 2 is prepared, for example, at a temperature below 30°C and in particular equal to 25°C. The mixing of the two compositions / solutions can be carried out at a temperature greater than or equal to 0°C ambient or less than 30°C and in particular 25°C. Then, the mixture must be left stirring at a temperature equal to or greater than 0°C and equal to or less than 30°C and in particular equal to 25°C for at least 4 hours and in particular 16 hours. The mixture can be crystallized at a temperature greater than or equal to 90°C and less than or equal to 200°C and in particular equal to 170°C.

[0026] Detailed description

[0027] The aluminum source may be chosen from any aluminum source capable of providing monomeric oxyhydroxide species. For the purposes of the present invention, "any aluminum source capable of providing monomeric oxyhydroxide species" means that this aluminum source does not provide polymeric aluminum oxyhydroxide species and makes it possible to solubilize all the aluminum in the clear aqueous solution 1. In particular, the aluminum source may be chosen from alumina, hydrated alumina, aluminum powder, Al, Al2(SO4)3, aluminum hydroxide AI(OH)s, sodium aluminate (NaAlC ), potassium aluminate (KAIO2) and kaolin clays.

[0028] The silicon source can be chosen from any silicon source capable of providing monomeric or Si2-Sie oxyhydroxide species. Thus, the quantity of [SiOOH] units n larger is limited in composition / solution 2.

[0029] In particular, the silicon source may be selected from silica hydrogel, silicic acid, colloidal silica, fumed silica, tetraalkyl orthosilicates, silica hydroxides, precipitated silica, silica fume and clays. Colloidal silica is preferred in all embodiments of the method of the invention.

[0030] The alkali metal source M may be selected from a Na source and an alkali metal source K. Hydroxides of these two alkali metals are preferred as alkali metal sources.

[0031] According to a particular embodiment, M = K, said source of aluminum is aluminum hydroxide, said mixture of compositions / solutions 1 and 2 is crystallized at a temperature greater than or equal to 90°C and less than or equal to 200°C and in particular equal to 170°C for a duration equal to or greater than 4 hours and equal to or less than 16 hours.

[0032] According to a particular variant, 2.8 <x1 < 8, 0,3<y< 1.5 et 100<z<300 et selon une variante plus particulière x1 = 5, y = 0,5 et z = 200.

[0033] Advantageously, the purification / separation of said nanoparticles is carried out using a membrane, in particular a dialysis membrane. The operation of this method is simple, inexpensive, generally carried out using commercially available membranes (dialysis membrane), and it also saves water. It also allows the PBS ions to be retained in the porous network of the nano zeolites.

[0034] According to a second aspect of the invention, it relates to a method for manufacturing a colloidal suspension of nanoparticles, according to which nanoparticles are synthesized according to the method of the invention and said synthesized nanoparticles are dispersed in optionally sterile water or in an optionally sterile PBS buffer, consisting of water and 137 mM / L of NaCl, 2.7 mM / L of KCl, 10 mM / L of Na2HPO4 and 1.76 mM / L of KH2PO4 and having a pH equal to 7.2. A suspension of zeolite nanoparticles in water or in PBS is thus obtained.

[0035] It should be noted that the synthesis method of the invention also directly allows the obtaining of suspensions of LTL type nanoparticles in PBS.

[0036] According to another aspect, the present invention relates to a nanoparticle consisting of or comprising at least one LTL type zeolite nanocrystal which comprises PBS ions, in particular Na ions + 'K + , HPO42 ; HsO + , H2PO4; CT and in particular Na ions + and K + . Said nanoparticle may advantageously contain at least two species of the aforementioned ions. Depending on the pH, it may contain different quantities of HPO4 ions. 2 ; HsO + , H2PO . These ions, whatever they may be, are located in the porous network of the nano zeolite and / or in the interstices of its crystal network (as compensation ions) and / or on the surface of its crystal network, which can merge with the surface of the nanoparticle. Preferably the Na cations + and K + are found in the interstices of the crystal lattice as compensating ions.

[0037] Advantageously, said nanoparticle further contains in its crystal lattice cations of a metal chosen from Fe, Gd, Cu and Ce and in particular in an amount greater than or equal to 0.2% by mass and less than or equal to 5% by mass and / or a gas contained in its pore volume and chosen from oxygen, nitrogen, carbon dioxide and mixtures thereof. When it contains an ion as mentioned above, it can serve as a contrast product in diagnosis, in particular of solid tumors, in particular of the brain. When it contains a gas as mentioned above, it can have a therapeutic action by releasing oxygen; this therapeutic effect is particularly marked in the presence of an anticancer agent. This anticancer agent can be chosen from the anticancer agents subsequently described with reference to the combination product of the invention.

[0038] According to a particular embodiment which can be combined with each of the aforementioned embodiments, said zeolite has the following chemical formula (II):

[0039] |K a N / A b (H2O) C | (Alx Si y O 72 ) (II) in which x is greater than 9 and / or y is less than 27 and / or a is greater than 6. Advantageously, a is greater than 6 and y is less than 27. Advantageously, a is less than or equal to 10 and y is less than or equal to 26. The formula between the bars shows the compensation ions (K + and Na + ) and the water molecules present in the nano zeolite.

[0040] According to a particular embodiment of the invention, combinable with the aforementioned embodiments, said zeolite has the following chemical formula (III):

[0041] IK9.04 Na2.57 (H2O)24.46 | (Al .12 Si25.5lO?2) (III)

[0042] Advantageously, said nanoparticles each have a total pore volume greater than or equal to 0.4 cm 3 / g and less than or equal to 1.5 cm 3 / g and / or a specific surface area SBET greater than or equal to 200 m 2 / g and less than or equal to 1000 m 2 / g. In particular, said nanoparticles may each have a total pore volume greater than or equal to 0.5 cm 3 / g and less than or equal to 1.4 cm 3 / g and in particular equal to 0.73 or 0.85 and / or a specific surface area SBET greater than or equal to 300 m 2 / g and less than or equal to 900 m 2 / g and in particular equal to 538 m 2 / g or 490 m 2 / g.

[0043] According to another aspect, the present invention relates to a colloidal suspension of nanoparticles consisting of or comprising at least one LTL type zeolite nanocrystal as defined above in a dispersant chosen in particular from alcohols, water and saline phosphate buffer and mixtures thereof. Preferably, PBS is chosen.

[0044] According to another aspect, the present invention relates to a pharmaceutical composition which contains as a pharmaceutically active agent or as a contrast agent, a plurality of nanoparticles consisting of or comprising at least one LTL type zeolite nanocrystal or the suspension according to the invention.

[0045] According to another aspect, the present invention relates to a combination product comprising or consisting of an anticancer agent and the pharmaceutical composition according to the invention for their simultaneous or delayed administration. The anticancer agent is advantageously administered first. The anticancer agent may be chosen from temozolomide® (4-methyl-5-oxo-2,3,4,6,8-pentazabicyclo

[0046] [4.3.0]nona-2,7,9-triene-9-carboxamide methazolastone), doxorubicin (8S, 10S)-10-(4-amino-5-hydroxy-6-methyl-tetrahydro-2H-pyran-2-yloxy)-6,8, 11 - trihydroxy-8-(2-hydroxyacetyl)-1-methoxy-7,8,9,10-tetrahydrotetracene-5,12-dione), cisplatin (cis-diamminedichloroplatinum(ll)).

[0047] According to another aspect of the invention, it relates to a nano zeolite according to the invention or a suspension according to the invention for its use as a medicament, in particular for the treatment of solid tumors, in particular of the brain, in particular glioblastoma, or for its use as a contrast agent, in particular in magnetic resonance imaging (MRI).

[0048] The inventors have demonstrated that the LTL-type nanoparticles according to the invention, which are therefore synthesized in PBS saline buffer, are less cytotoxic and therefore likely to be better accepted by the body. PBS has the same salt concentration as the human body; it also contains the dihydrogen phosphate / hydrogen phosphate pair, which allows the maintenance of blood pH in the body. Without the inventors being bound to this explanation, it would seem that certain ions of the PBS contained in the porous network of the nanozeolites stabilize the nanoparticles and prevent them from degrading in the biological environment, which reduces their toxicity potential. In addition, the ions of the PBS can also regulate the charge on the surface of the nanoparticles, which can influence their behavior and interaction with biological cells and tissues.The nanoparticles of the invention thus avoid or reduce the allergy phenomena described above.

[0049] Regardless of the embodiment and aspect of the invention, the LTL zeolite nanocrystal may have the shape of a flake (flat parallelepiped). This nanocrystal may have an average length greater than or equal to 5nm and less than or equal to 20nm and an average width greater than or equal to 2 and less than or equal to 15nm. In particular, the average length may be 30nm and the average width 10nm. Throughout the application, the nanoparticle is preferably made of a single crystal of LTL type zeolite.

[0050] The porous network of the zeolite can extend in only one direction.

[0051] The pharmaceutical composition of the invention may be formulated for oral, mucosal (intranasal) or parenteral administration.

[0052] Definitions

[0053] The terms "average size" designate, within the meaning of the invention, the average size (in number or in intensity) measured by dynamic light scattering. The average size corresponds to the diameter of the sphere which scatters at the same speed as that of the particle being measured. The fundamental size distribution obtained by DLS is an intensity distribution. The operating conditions are as defined in the remainder of the application. Within the meaning of the invention, the term "nanoparticle" designates any solid particle with a maximum dimension of less than 10 Onm. It may be a single crystal or an aggregate of crystals.

[0054] The term "composition" refers to a homogeneous or inhomogeneous mixture of at least two compounds that may be in different physical states. This term therefore includes suspensions, solutions and dispersions. This term does not limit a particular physical state of matter.

[0055] The term PBS refers to phosphate buffered saline which is an aqueous solution of NaCl (0.137 M), KCl (0.0027 M), sodium phosphate dibasic (0.01 M) and potassium phosphate monobasic (0.0018 M). Its pH is 7.2.

[0056] The term "LTL zeolite nanoparticle" refers to nanoparticles comprising or consisting of a single crystal of LTL zeolite. LTL zeolite crystallizes in a hexagonal system. The chemical formula of an LTL zeolite nanocrystal is: IK a Nab(H2O) c l [Al x If y O72] with x+y< 36, the Na ions + and K + being present as compensating ions, so as to ensure the electrical neutrality of the nanocrystal. Water molecules may be absent from the nanocrystal (c=0).

[0057] The term "solution" means an aqueous solution having substantially the same refractive index as water, i.e., a refractive index of approximately 1.333 ± 0.030, measured under standard conditions.

[0058] The terms "pore volume" or "pore network" refer to the set formed by the cavities, the channels connecting these cavities to each other and the pores which connect the channels and cavities to the exterior of the nanoparticle.

[0059] The term "pharmaceutically active" means an agent which confers curative or preventive therapeutic properties to the composition which contains it and which, if it has harmful side effects, does not outweigh the curative and / or preventive effects.

[0060] The term "pharmaceutically acceptable" means any agent that can be ingested, injected or brought into contact with the skin and mucous membranes without generating an adverse effect or without generating an adverse effect of such magnitude that it disrupts the patient's state of health. Brief description of the figures

[0061] The characteristics and advantages of the invention will appear on reading the following description based on the appended figures, among which:

[0062] - Fig. 1 represents the size distribution of the K-LTL-PBS nanoparticles of the invention; The nanoparticles are dispersed in a PBS solution;

[0063] - Fig. 2A represents images obtained by scanning electron microscope of nanoparticles (nanocrystals) of type K-LTL-PBS and Gd-LTL-PBS with a scale of 1 pm / 500 nm and Fig. 2B represents images obtained by transmission electron microscope of nanoparticles (nanocrystals) of type K-LTL-PBS with a scale of 100 nm and 40 nm;

[0064] - Fig. 3 represents the quantity of adsorbed nitrogen as a function of the relative pressure obtained under the operating conditions described below for K-LTL-PBS type nanoparticles;

[0065] - Fig. 4 represents a comparison of the viability of U87-MG cells exposed to two colloidal suspensions of nanoparticles containing respectively 10 pg / mL and 100 pg / mL of K-LTL-Water or Gd-LTL-Water nanoparticles with that obtained for the same concentrations of K-LTL-PBS and Gd-LTL-PBS nanoparticles;

[0066] - Fig. 5 represents on the left the percentage of hemolysis as a function of the dose of nanoparticles (in pg / mL) of type K-LTL and Gd-LTL synthesized with water or PBS;

[0067] - Fig. 6A represents the change in oxygen concentration in water obtained by contacting with K-LTL-Water and Gd-LTL-Water particles, previously loaded with O2, and Fig. 6B represents the change in oxygen concentration in PBS obtained by contacting with K-LTL-PBS and Gd-LTL-PBS particles of the invention, previously loaded with O2.

[0068] EXAMPLES

[0069] SYNTHESIS AND CHARACTERIZATION OF NANOPARTICLES

[0070] The raw materials used in the following examples are listed below:

[0071] - Sodium hydroxide (Sigma Aldrich, 98%)

[0072] - Potassium hydroxide (Sigma Aldrich, 90%)

[0073] - Aluminum hydroxide (type A2100, Alfa Aesar, 76.5% by mass AI(OH)s, 23.5% by mass H2O) - Aluminum powder (325 mesh, Alfa Aesar 99.5%)

[0074] - Colloidal silica (SiÛ2 (Ludox HS-30, 30 wt.% SiC>2, pH=9.5-10.2 Sigma Aldrich)

[0075] - Colloidal silica (SiÛ2 (Ludox SM-30, 30 wt.% SiC>2, pH=9.7-10.3 Sigma Aldrich)

[0076] - Gadolinium nitrate Gd(NOs)3, 6H2O (Aldrich, 99.9%)

[0077] - Phosphate buffered saline (PBS) (Sigma Aldrich, pH=7.15-7.25)

[0078] These starting reagents were used as received from the manufacturers, without further purification.

[0079] Analysis methods used and operating conditions

[0080] The different zeolite materials obtained in the examples were characterized on different size scales.

[0081] Scanning and transmission electron microscopy (SEM / TEM) imaging:

[0082] The morphology of the nanoparticles was examined by scanning electron microscopy (SEM) using a Tescan Mira I LMH device at a voltage of 20 kV.

[0083] Transmission electron microscopy (TEM): Colloidal suspensions were sonicated for 15 minutes, then 2 to 3 drops of fine particle suspensions were dried on 300 mesh copper electron microscope grids coated with carbon film. Images were obtained using a FEI Tecnai G2 30 microscope (Vacc = 300 kV, LaB6).

[0084] Dynamic Light Scattering (DLS) Analysis:

[0085] The hydrodynamic diameters of the zeolite material in the different suspensions were determined with a Malvern Zetasizer Nano instrument. The surface charge of the samples was determined by measuring the zeta potential of the suspensions. The analyses were carried out on samples after purification with a solid concentration of 1 wt% dispersed in a PBS solution at pH = 7.2. The backscattering geometry (scattering angle 173°, HeNe laser with an output power of 3 mW at a wavelength of 632.8 nm) allows measurements at a high sample concentration, since complete penetration of the incident light through the sample is not required.

[0086] Chemical analysis by ICP-MS (inductively coupled plasma mass spectrometry)

[0087] The chemical composition of the zeolite samples was characterized by inductively coupled plasma mass spectrometry using Agilent Technologies 7900 equipment. The samples were prepared according to the following procedure: (i) 50 mg of zeolite sample was dissolved in 3 mL of hydrofluoric acid (HF) (Sigma-Aldrich 40-45%), (ii) a 0.5 mL solution (HNO3 / HCl equals 1:3 v / v) was added and then the mixture was heated at 110 °C for 1 h in a polytetrafluoroethylene (PTFE) bottle (100 mL), and subsequently (iii) 96.5 mL of distilled water and 2 g of boric acid (H3BO3) were added. The obtained solution was stirred overnight to facilitate the dissolution of boric acid. Finally, 10 mL of the solution was diluted 10 times with distilled water before analysis.

[0088] Example 1: Synthesis of K-LTL type zeolite nanocrystals in a PBS solution (K-LTL-PBS)

[0089] Alumina suspension C is prepared by dissolving 2 g of KOH in 8 g of PBS followed by slow addition of 0.496 g of aluminum hydroxide.

[0090] A silicate suspension D is prepared by mixing 10 g of colloidal silica (LUDOX® SM-30) with 1.1 g of KOH and 2.88 g of PBS.

[0091] The two suspensions C and D are mixed and then left stirring at room temperature for 24 hours.

[0092] The initial molar composition of the mixture is as follows:

[0093] 5K2O: 0.5AI2O3: 10SiO2: 200 (PBS)

[0094] The mixture obtained is crystallized for 16 hours in an oven at 170°C.

[0095] The obtained nanoparticle suspension is purified by dialysis membrane (Visking Dialysis Tubing, Code No. - DTV.12000.08 Size No.8 (25.4mm, 30m), Molecular Weight 12-14000 Daltons), until a pH of 7.2 is reached. 80 to 100 mL (2.0 to 2.5 g) of a colloidal suspension containing 2.5% by mass of K-LTL type zeolite nanocrystals are obtained. The mass yield of crystals ranges from 66 to 76%. The final suspensions were sonicated for 2 hours. Formula of the K-LTL-PBS zeolite nanoparticle obtained by ICP:

[0096] |Na2.57K9.04(H2O)24.46 | (Si25.5lAI-10.12O72)

[0097] The formula in parentheses corresponds to the crystal lattice, between the bars are the number of moles of compensation ions and the number of moles of water.

[0098] Example 2: Synthesis of colloidal suspensions of nanoparticles according to the invention A given quantity of nanoparticles of example 1 was dispersed in 10 mL of PBS under sonication for 2 hours. Colloidal suspensions containing respectively 1%, 2%, 3% and 4% by mass of nanoparticles of type K-LTL-PBS dispersed in PBS are obtained. These nanoparticles are stable in colloidal suspension at room temperature for at least one week.

[0099] Example 3: Synthesis of zeolite nanoparticles charged with metal ions

[0100] 5 mL of each of the two colloidal suspensions obtained in Example 2, each containing 2.5% by mass of nanoparticles, were added separately to 25 mL of a solution of gadolinium (III) nitrate hexahydrate (Gd(NO3)3.6H2O) with a concentration of 0.03 mM). The suspension was then kept stirring at room temperature for 1 h. This ion exchange process was carried out twice. The final suspension was then washed 3 times with PBS to finally obtain a suspension of Gd-LTL-PBS zeolite in PBS and therefore at pH=7.2.

[0101] Example 4: Incorporation of dioxygen and / or CO2 into the porous network of nanoparticles

[0102] Each of the colloidal aqueous suspensions obtained in Example 2 or Example 3 was then bubbled with C>2, CO2 or carbogen at a flow rate of 800 mL / min for 30 minutes.

[0103] This produces nanoparticles whose porous network is at least partially filled with O2 and / or CO2.

[0104] Example 5: Morphological characterization of the nanoparticles obtained in examples 1, 3, and 4.

[0105] Scanning / transmission electron microscopy (SEM) / (TEM) were used to determine the morphology and size of the synthesized K-LTL-PBS and Gd-LTL-PBS nanoparticles.

[0106] The curve giving the size distribution of K-LTL-PBS nanoparticles dispersed in PBS is shown in Fig. 1. ;

[0107] Scanning electron microscopy images of K-LTL and Gd-LTL zeolite nanoparticles (see Fig. 2A) show small clusters of agglomerated nanoparticles; however, images obtained by transmission microscopy (see Fig. 2B) show that these nanoparticles are small individual rectangular-shaped nanocrystals with a width of 10 nm and a length of 30 nm.

[0108] In addition, high-resolution transmission electron microscopy images show that the nanoparticles are highly crystalline. These are nanoparticles each formed from a single nanocrystal. Example 6: Chemical analysis of K-LTL nanoparticles: method and results

[0109] The chemical formula of the zeolite material of K-LTL and Gd-LTL type nanoparticles is as follows (determined by ICP):

[0110] | Na2.57K9.04(H2O)24.46 | (Si25.5lAll0.12O72)

[0111] |Nai.1oK9.0lGdo.4(H20)24.32 | (Si25.42All O.34O72)

[0112] The formula in parentheses corresponds to the crystal tetrahedral lattice, between the bars are the number of moles of compensation ions and the number of moles of water.

[0113] Example 7: Determination of porosity by adsorption / desorption of dinitrogen

[0114] The porosity of the K-LTL-PBS and Gd-LTL-PBS-PBS zeolites synthesized in the above examples was characterized by nitrogen gas adsorption / desorption measurements.

[0115] The textural properties of the nanoparticles obtained in the above examples were determined by the nitrogen adsorption / desorption isotherms measured at a temperature of -196°C using a “Micromeritics 3Flex” surface characterization device. All samples were degassed at 250°C under vacuum overnight before analysis. The external surface area of ​​the zeolite nanoparticles and the micropore volume of the zeolite nanoparticles were evaluated by the t-plot method.

[0116] Nitrogen adsorption / desorption measurements for both zeolites resulted in hybrid isotherms of type I and IV according to the IUPAC classification (see Fig. 3). The measurement of the SBET specific surface area, the total pore volume and the micropore volume of the zeolite nanoparticles are grouped in Table 1 below. A micropore is defined as a pore smaller than 0.08 mm.

[0117] Table 1

[0118] Table 2 below lists the chemical composition of the K-LTL-PBS and Gd-LTL-PBS nanoparticles of the invention. Table 2

[0119] The pharmacological properties of the nanoparticles of the invention were studied. The results are shown in the following examples.

[0120] Example 8: Toxicity test on human cell line.

[0121] The cells used were derived from a human glioblastoma cell line, U87-MG, purchased from American Type Culture Collection (ATCC). The cells were cultured in DMEM 1 g / L glucose (Sigma-Aldrich) supplemented with 10% fetal bovine serum (Eurobio), 2 mM glutamine (Sigma-Aldrich), and penicillin (100 U / mL) / streptomycin (100 pg / mL) (Sigma-Aldrich). The cells were maintained in culture at 37°C with 5% CO2 and 95% humidity.

[0122] Cell viability was assessed 72 hours after exposure to zeolite nanoparticles using the WST-1 test (Roche) according to the manufacturer's instructions. The results are shown in Fig. 4. This figure also shows the percentage of cell viability with exposure to nanoparticles synthesized in water instead of PBS. It can be seen that for the K-LT-PBS and Gd-LTL-PBS nanoparticles of the invention, the percentage of viable cells is higher than for nanoparticles synthesized from the same mixture but with water instead of PBS (Na-LTL-Water and Gd-LTL-Water).

[0123] Example 9: Red blood cell viability test.

[0124] To confirm the absence of toxicity of the nanoparticles of the invention, red blood cells were exposed to increasing concentrations of nanoparticles according to the invention (from the aforementioned examples).

[0125] The results are visible in Fig. 5. It can be seen that for nanoparticles synthesized in water, the maximum percentage of hemolysis is 1.12%; 0.649%; 0.00% and 0.00% respectively for K-LTL-Water, Gd-LTL-Water, K-LTL-PBS and Gd-LTL-PBS nanoparticles.

[0126] Furthermore, in the presence of a large amount of nanoparticles (500 pg / mL), the highest hemolysis percentage obtained for nanoparticles is 0.0% for Gd-LTL-PBS particles. These results confirm that the use of PBS during the synthesis of nanoparticles decreases their toxicity.

[0127] Example 10: Study of oxygen release by zeolitic nanoparticles in aqueous and hypoxic conditions

[0128] A hypoxia chamber (IN VIV02 500™, 3M) was used to achieve a stable and precisely controlled gas composition of the atmosphere with an accuracy of 0.1% O2 by adjusting the amount of N2.

[0129] The PBS solution (Sigma-Aldrich) was equilibrated with the gas mixture contained in the hypoxia chamber for 1 h before the experiment.

[0130] A closed tube containing 12 mL of PBS equilibrated at 37°C and a dissolved oxygen sensor (SevenGo (Duo) pro™ / OptiOx™, Mettler Toledo) were used inside the hypoxia chamber. Before the experiment, the baseline was established by measuring the oxygen content of the PBS for 30 min.

[0131] The nanoparticles of the above-mentioned examples, previously loaded with oxygen, were then added to the system and the dissolved oxygen in the PBS solution was continuously measured for 1 h. The results are shown in Fig. 6. It can be seen that the K-LTL-Water and / or Gd-LTL-Water nanoparticles exhibit excellent release capacity (4.07 and 5.12 times higher than the baseline, respectively) due to their chemical composition. In the case of the K-LTL-PBS and / or Gd-LTL-PBS nanoparticles, the O2 loading in the nanoparticles was monitored. In the case of the network extending in one direction, the use of PBS rather than water during particle synthesis may result in differences in the physicochemical properties of the zeolite such as zeolite surface area, pore volume, and charge distribution, which in turn may affect the interactions between the zeolite and oxygen and influence oxygen release.A longer release kinetics is beneficial for more sustained reoxygenation of the target tissue throughout the therapeutic protocol. The slower release rate allows cells to adapt sustainably to reoxygenation and thus maintain the molecular processes linked to reoxygenation. This also allows for more time between injection and radiotherapy session. Longer reoxygenation also reduces the number of injections required.

Claims

Claims 1. Process for the synthesis of nanoparticles consisting of or comprising at least one zeolite nanocrystal according to which: - a first composition / solution 1 is prepared containing a source of aluminum and a source of an ion of an alkali metal M, in particular K; - a second composition / solution 2 is prepared comprising a source of silicon and a source of an ion of an alkali metal M, in particular K, said compositions / solutions 1 and 2 being free of any organic structuring agent; - mix compositions / solutions 1 and 2 and leave the mixture stirring, - the mixture is crystallized at a temperature greater than or equal to 50°C; and possibly - said nanoparticles thus formed are separated, characterized in that said first composition / solution 1 and said second composition / solution 2 are both constituted by said source and a buffer constituted by water and 137 mM / L of NaCl, 2.7 mM / L of KCl, 10 mM / L of Na2HPO4, 1.76 mM / L of KH2PO4 and having a pH equal to 7.2 (PBS) and in that said mixture of compositions / suspensions 1 and 2 has the following molar composition (I): XI M2O : YAI2O3 : 10SiO2: Z PBS (I) in which 2.8 <X1 < 8,8 0.3 <Y< 1 ,5, 40 <Z<300.

2. Method according to claim 1, characterized in that said nanoparticles are separated using a membrane.

3. Method according to claim 1 or 2, characterized in that M = K, in that said source of aluminum is aluminum hydroxide and in that said mixture is crystallized at a temperature greater than or equal to 90°C and less than or equal to 200°C and in particular equal to 170°C for a duration equal to or greater than 4 hours and equal to or less than 16 hours.

4. Nano zeolite comprising or consisting of a zeolite nanocrystal of LTL type, characterized in that it comprises PBS ions, in particular Na ions + , K + , HPO4 2 ; H2PO4; H3O + and CI'.

5. Nano zeolite according to claim 4, characterized in that it further contains in its crystal lattice cations of a metal chosen from Fe, Gd, Cu and Ce and in particular in an amount greater than or equal to 0.2% by mass and less than or equal to 5% by mass and / or a gas contained in its pore volume and chosen from oxygen, nitrogen, carbon dioxide and their mixtures.

6. Nano zeolite according to claim 4 or 5, characterized in that said LTL type zeolite has the following chemical formula (II): |K a Nab (H2O) C | (Al x If y O 72 ) (II) in which x is greater than 9 and / or y is less than 27 and / or a is greater than 6.

7. Nano zeolite according to claim 6, characterized in that said zeolite has the following chemical formula (III) IK9.04 Na2.57 (H2O)24.46 | (AI10.12 Si25.5iO?2) (III) or the following chemical formula: |Nai.ioK9.oiGdo.4(H20)24.321 (Si25.42Alio.34072).

8. Nano zeolite according to any one of claims 4 to 7, characterized in that it has a total pore volume greater than or equal to 0.5 cm 3 / g and less than or equal to 1.4 cm 3 / g and in particular equal to 0.73 or 0.85 and / or a specific surface area SBET greater than or equal to 300 m 2 / g and less than or equal to 900 m 2 / g and in particular equal to 538 m 2 / g or 490 m 2 / g.

9. Nano zeolite according to any one of claims 4 to 8, characterized in that it has the shape of a flake or a flat parallelepiped.

10. Nano zeolite according to any one of claims 4 to 9, characterized in that it has an average length greater than or equal to 5nm and less than or equal to 20nm and an average width greater than or equal to 2 and less than or equal to 15nm.

11. Nano zeolite according to any one of claims 4 to 10, characterized in that its porous network extends in a single direction.

12. Colloidal suspension of nanoparticles characterized in that it contains the nanoparticles according to any one of claims 4 to 11 in a dispersant chosen from alcohols, water, phosphate buffered saline (PBS) and mixtures thereof.

13. Pharmaceutical composition characterized in that it contains as pharmaceutically active product or as contrast agent, a plurality of nanoparticles consisting of or comprising at least one nano zeolite according to any one of claims 4 to 11 or the suspension according to claim 12.

14. Combination product comprising or consisting of an anticancer agent and the pharmaceutical composition according to claim 13 for their simultaneous or delayed administration.

15. Nanoparticle according to any one of claims 4 to 11 for its use as a medicament, in particular for the treatment of solid tumors, in particular of the brain, in particular glioblastoma.

16. Nanoparticle according to any one of claims 4 to 11 for its use as a contrast agent, in particular in magnetic resonance imaging (MRI).

17. Suspension according to claim 12, in particular in PBS, for its use as a medicament, in particular for the treatment of solid tumors, in particular of the brain, in particular glioblastoma.

18. Suspension according to claim 12, in particular in PBS, for its use as a contrast agent, in particular in magnetic resonance imaging (MRI).

Images