WO2017184085A1

WO2017184085A1 - Core-shell particles

Info

Publication number: WO2017184085A1
Application number: PCT/SG2017/050226
Authority: WO
Inventors: Gen Yong; Ye Liu
Original assignee: Agency For Science, Technology And Research
Priority date: 2016-04-21
Filing date: 2017-04-21
Publication date: 2017-10-26

Abstract

There is provided a core-shell particle comprising a core comprising at least one active agent and a method of forming said core-shell particle, wherein said core having ions of an additive surrounding a surface of said core; and an encapsulating shell comprising (i) a polymer and/or (ii) a metal or a semi-metal oxide, phosphate or carbonate. In a preferred embodiment, chondroitin sulphate nanocrystals is first prepared by using pH recrystallization method. Sodium citrate is then added to form citrate ions-capped chondroitin sulphate particles. Poly L-lysine (PLL) and tetramethoxysilane (TMOS) are subsequently added to form a silica shell surrounding the citrate-ions capped chondroitin sulphate particles, and thus forming the claimed core-shell particle. The particles may be used as sustained release formulations in therapy.

Description

Title of Invention: Core-Shell Particles

Technical Field

The present invention generally relates to core-shell particles and methods for their preparation. The core- shell particles may be used for sustained and controlled drug delivery.

Background Art

Conventional methods for preparing core-shell particles treat the core as a sacrificial template to be removed after forming the shell. The void space left is then available for loading of active agents via a diffusion process. Other methods use carrier materials to aid in the condensation and entrapment of active agents into a core, followed by a secondary coating that forms a shell over the core. Yet other methods start from charged micronized drug crystals and use layer by layer encapsulation to encapsulate these crystals. These methods suffer from poor size control of the core and may also require the core to have specific surface charges in order for the shell to encapsulate the core. The resulting core-shell particles may also be too large and be unsuitable for use in sustained and controlled drug delivery. In other methods, a concentration gradient is used to drive the uptake of actives into a drug carrier, however such methods are reliant on the movement of actives through a barrier, and the size of the actives that can be loaded via this method is limited (for example, the active loading method). In yet other methods, drug carriers are suspended in solutions containing a high concentration of active molecules where the drug molecules are allowed to passively diffuse into the drug carrier, however in such methods, only a fraction of the active molecules are loaded into the drug carrier and hence are inefficient.

Maintaining a therapeutic concentration of actives at a target location is a challenging problem in many industries. Factors that affect therapeutic concentration comprise incompatible chemical environment, physical constraints on dose, size of dose to achieve efficacy, consumer/patient compliance and compound chemistry.

There is therefore a need to provide core- shell particles and methods for forming the same that overcomes, or at least ameliorates, one or more of the disadvantages described above.

Summary

According to one aspect of the present disclosure, there is provided a core-shell particle comprising: a core comprising at least one active agent, said core having ions of an additive surrounding the surface of said core; and an encapsulating shell comprising (i) a polymer and/or (ii) a metal or a semi- metal oxide, phosphate or carbonate.

Advantageously, the core-shell particle may be a core-shell nanoparticle. The core shell nanoparticle may exhibit sustained release profiles of the active agent which allows for controlled drug delivery.

Further advantageously, due to the core-shell nature of the particles, the active agent encapsulated in the core by the shell may protect the core contents from early and undesired release in the body.

According to another aspect of the present disclosure, there is provided a core-shell particle comprising: a core comprising at least one chondroitin salt, said core having ions of an additive comprising at least one citrate ion surrounding the surface of said core; and an encapsulating shell comprising Si0₂.

According to another aspect of the present disclosure, there is provided a method of forming a core-shell particle, comprising the steps of:

(a) providing a core comprising at least one active agent, said core having ions of an additive surrounding a surface of said core; and (b) encapsulating a shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, phosphate or carbonate, onto said core to thereby form said core-shell particle.

Advantageously, the size and/or density of the core may be controlled by the addition of the additive, thereby allowing size control of the final core. By being able to achieve good size control, enhanced pharmaceutical properties of the encapsulated active agent may be obtained. Further advantageously, sustained release profiles of the active agent may be obtained which allows for controlled drug delivery. Further advantageously, direct encapsulation of the active agent may allow for ultra-high efficiency loading and encapsulation of the active agent. Thus, a high loading of active agent may be achieved in the core.

Further advantageously, the method of the present disclosure may produce core-shell particles with even particle morphology and size distribution in the core and improved formulation stability and bio-availability of the particles.

Further advantageously, this method may produce core-shell particles that are capable of both sustained and burst release.

Also advantageously, this method may produce core-shell particles that are bio-compatible and bio-degradable. Further advantageously, this method may allow for cost-efficient production of the core- shell particles.

Further advantageously, this method may allow for layer-by-layer encapsulation of an active agent.

(a) providing a core comprising at least one chondroitin salt, said core having citrate ions disposed on a surface of said core;

(b) encapsulating a shell comprising Si0₂ onto said core to thereby form said core-shell particle. According to a further aspect of the present disclosure, there is provided a core-shell nanoparticle produced by the method disclosed herein.

According to another aspect of the present disclosure, there is provided a core-shell nanoparticle disclosed herein for use in therapy. According to a further aspect of the present disclosure, there is provided a core-shell particle for use in sustained release formulations.

Definitions

The following words and terms used herein shall have the meaning indicated: The term "core-shell" as used herein refers to particles comprising a core material with at least one shell disposed thereabout (including less than 100% coverage). The term "core-shell" encompasses particles with one shell, and also encompasses particles with multiple shells.

The term "active agent" as used herein refers to an organic or inorganic molecule that may be synthetic or natural. The active agent may be a therapeutic agent or a cosmetic agent.

The term "therapeutic agent" as used herein refers to a biological or chemical compound/agent that provides a desired biological or pharmacological effect upon administration to a subject. The term "cosmetic agent" as used herein refers to a biological or chemical compound/agent that is suitable for administration to the skin and is capable of improving the appearance of skin.

The term "nanocrystalline" as used herein refers to crystals in the range of nanometers. The term "additive" as used herein refers to an agent suitable for controlling the size of the core. The size of the core may be controlled by stabilization of the surface chemistry of the core particles, thereby preventing aggregation. The term "encapsulate" or "encapsulating" as used herein refers to particles that are wholly or partially confined within a shell.

The term "saturated" as used herein refers to a point of maximum concentration, in which no more solute may be dissolved. The term "conjugate" as used herein refers to two or more molecules that are covalently linked.

The term "metal oxide" as used herein refers to a compound having at least one metal atom and at least one oxygen atom (where the oxygen atom is bonded either to another metal atom or to another oxygen atom) of varying stoichiometry that is covalently bonded to the metal atom. The term "metal oxide" encompasses within its meaning "mixed-metal oxides" which refers to compounds having at least two different metal atoms. The "metal" may be selected from the group consisting of alkali metals, alkaline earth metals, metals of the third, fourth, or fifth main group of the periodic table of elements, metals of the transition metal groups of the Periodic Table of Elements, the lanthanides, and the actinides.

The term "semi-metal oxide" as used herein refers to a compound having at least one semi-metal and at least one oxygen atom (where the oxygen atom is bonded either to another metal atom or to another oxygen atom) of varying stoichiometry that is covalently bonded to the metal atom. The "semi-metal" or "metalloids" in general are known to the skilled artisan. The term "semi-metal" may include boron, silicon, germanium, arsenic, antimony, tellurium, carbon, aluminium, selenium, polonium and astatine.

The term "surrounding" refers to ions of an additive that are disposed around a surface of a core and encompasses within its meaning ions of the additive which may be disposed on the surface of the core, or disposed on one or more intermediate shells or layers around the core.

As used herein, "intermediate polymer shell" refers to a polymer shell surrounding the core of the core-shell particle, which is further encapsulated by an encapsulating shell. The word "substantially" does not exclude "completely" e.g. a composition which is "substantially free" from Y may be completely free from Y. Where necessary, the word "substantially" may be omitted from the definition of the invention. Unless specified otherwise, the terms "comprising" and "comprise", and grammatical variants thereof, are intended to represent "open" or "inclusive" language such that they include recited elements but also permit inclusion of additional, unrecited elements.

As used herein, the term "about", in the context of concentrations of components of the formulations, typically means +/- 5% of the stated value, more typically +/- 4% of the stated value, more typically +/- 3% of the stated value, more typically, +/- 2% of the stated value, even more typically +/- 1% of the stated value, and even more typically +/- 0.5% of the stated value.

Throughout this disclosure, certain embodiments may be disclosed in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosed ranges. Accordingly, the description of a range should be considered to have specifically disclosed all the possible sub-ranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

Certain embodiments may also be described broadly and generically herein. Each of the narrower species and subgeneric groupings falling within the generic disclosure also form part of the disclosure. This includes the generic description of the embodiments with a proviso or negative limitation removing any subject matter from the genus, regardless of whether or not the excised material is specifically recited herein. Detailed Disclosure of Embodiments

Exemplary, non-limiting embodiments of the core-shell particles and methods for preparing the same will now be disclosed.

The present disclosure provides a core-shell particle comprising: a core comprising at least one active agent, said core having ions of an additive surrounding a surface of said core; and an encapsulating shell comprising (i) a polymer and/or (ii) a metal or a semi- metal oxide, phosphate or carbonate.

The present disclosure also provides a method of forming a core-shell particle, comprising the steps of:

(a) providing a core comprising at least one active agent, said core having ions of an additive surrounding a surface of said core; and

(b) encapsulating a shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, phosphate or carbonate, onto said core to thereby form said core-shell particle.

The particle may comprise an intermediate polymer shell disposed between the core and additive ions.

Advantageously, the addition of the additive to the core may be done to control the size of the core, thereby allowing the production of core-shell particles in a desired size range. The additive may control the aggregation and dispersity of the active core particles by stabilizing their surface chemistry and preventing unwanted aggregation. Ions of the additive may adhere to the surface of the core, either directly or through an intermediate polymer shell, forming a negatively charged layer/hydration shell that would repel additional negatively charged core particles from adhering to the surface of the core, hence restricting the growth of the core. This advantageously allows for the tailored production of core-shell particles that are able to maintain a therapeutic concentration of active agent at a target location. The presently disclosed core-shell particles are therefore useful in sustained and controlled drug delivery and may be an improvement over other core-shell particles produced through conventional methods, for example, the passive loading method, or the active loading method.

The ions of the additive may adhere to the surface of the core or an intermediate polymer shell through charge-charge interactions or intermolecular bonding forces such as van der Waals forces or hydrophobic interactions.

The methods disclosed herein may also advantageously produce core-shell particles with controllable release characteristics. Release characteristics may be controlled by the thickness and degree of crosslinking of the polymer(s) and/or metal or semi-metal oxide(s) in the shell. A thicker and/or more cross-linked shell may lead to core-shell particles that exhibit a longer release. Burst release may be obtained via surface treatment of the core with additive and immediate recovery through ethanol wash without a prolonged reaction.

The additive may be reacted with the core from 2 to 48 hours, or 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48 hours, or any range or value therebetween.

The additive may stabilize the surface of the core. The ions of the additive may have antagonistic charge and/or affinity interactions with a core particle. The ions of the additive may be disposed on the surface of the core, or disposed on the intermediate polymer shell. The ions of the additive may be conjugated to the surface of the core or conjugated to the intermediate polymer shell. The ions of the additive and the intermediate polymer shell may form a negatively charged shell. The negatively charged shell may repel additional negatively charged core particles from adhering to the surface of the core, hence restricting the growth of the core. The additive may comprise one or more citrates and/or polyethylene glycol (PEG). The polymer of the intermediate polymer shell may be selected from the group consisting of at least one poly-L-lysine (PLL), polyethyleneimine (PEI), dextran, polyelectrolytes, polycaprolactames (PCL), alginates, and chitosan.

The encapsulating shell may be selected from the group consisting of:

(a) shell comprising or consisting of polymer; (b) shell comprising or consisting of metal oxide, phosphate or carbonate;

(c) shell comprising or consisting of semi-metal oxide, phosphate or carbonate;

(d) shell comprising or consisting of polymer and metal oxide, phosphate or carbonate; and

(e) shell comprising or consisting of polymer and semi-metal oxide, phosphate or carbonate.

The metal oxide may be AI₂O₃, ZnO, Ti0₂ and iron oxides, preferably Fe₂0₃ or FeO. The semi-metal oxide may be silica (SiO₂). The metal carbonate may be calcium carbonate. The metal phosphate may be calcium phosphate. The encapsulating shell may comprise or consist of SiO₂. The SiO₂ may be cross-linked SiO₂. The shell may comprise or consist of at least one silicon comprising polymer.

The polymer of the encapsulating shell may be selected from the group consisting of poly-L-lysine (PLL), polyethyleneimine (PEI), dextran, polyelectrolytes, polycaprolactames (PCL), alginates, chitosanand polymerized silicon alkoxide.

The shell may comprise or consist of SiO₂, such as cross-linked SiO₂.

The active agent may be a therapeutic agent or a cosmetic agent. The active agent may be selected from the group consisting of chondroitin salts, heparin salts, glycosaminoglycans, glycoproteins, biomolecules, chondroitin sulfate, and heparin sulfate. The active agent may be chondroitin sulfate or heparin sulfate.

The active agent may be in amorphous form, partially crystalline form, or crystalline form. The core-shell particle may be a core-shell nanoparticle. The present disclosure also provides a core-shell particle comprising a core of at least one chondroitin salt having at least one citrate ion on the surface of said core and an encapsulating shell comprising SiO₂.

The present disclose also provides a core-shell particle comprising a core of at least one chondroitin salt, an intermediate polymer shell comprising PLL having at least one citrate ion disposed on the intermediate polymer shell, and an encapsulating shell comprising S1O₂.

The core may have an average diameter of about 10 nm to about 1000 nm. The average diameter may be about 10 nm to about 1000 nm, about 50 nm to about 1000 nm, about 100 nm to about 1000 nm, about 150 nm to about 1000 nm, about 200 nm to about 1000 nm, about 250 nm to about 1000 nm, about 300 nm to about 1000 nm, about 350 nm to about 1000 nm, about 400 nm to about 1000 nm, about 450 nm to about 1000 nm, about 500 nm to about 1000 nm, about 500 nm to about 1000 nm, about 600 nm to about 1000 nm, about 650 nm to about 1000 nm, about 700 nm to about 1000 nm, about 750 nm to about 1000 nm, about 800 nm to about 1000 nm, about 850 nm to about 1000 nm, about 900 nm to about 1000 nm, about 950 nm to about 1000 nm, about 50 nm to about 950 nm, about 50 nm to about 900 nm, about 50 nm to about 850 nm, about 50 nm to about 800 nm, about 50 nm to about 750 nm, about 50 nm to about 700 nm, about 50 nm to about 650 nm, about 50 nm to about 600 nm, about 50 nm to about 550 nm, about 50 nm to about 500 nm, about 50 nm to about 450 nm, about 50 nm to about 400 nm, about 50 nm to about 350 nm, about 50 nm to about 300 nm, about 50 nm to about 250 nm, about 50 nm to about 200 nm, about 50 nm to about 150 nm, about 50 nm to about 100 nm, or about 50 nm, about 100 nm, about 150 nm, about 200 nm, about 250 nm, about 300 nm, about 350 nm, about 400 nm, about 450 nm, about 50 nm, about 500 nm, about 550 nm, about 600 nm, about 650 nm, about 700 nm, about 750 nm, about 800 nm, about 850 nm, about 900 nm, about 950 nm, or about 1000 nm, or any value or range therebetween. The core may have an average diameter of about 130 nm to about 270 nm (i.e. 200±70 nm). The present disclosure also provides a core-shell particle comprising: a core comprising at least one active agent, said core having ions of an additive on the surface of said core; and

an encapsulating shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, phosphate or carbonate. The present disclosure also provides a core-shell particle comprising: a core comprising at least one active agent, said core having ions of an additive disposed around the surface of said core; and an encapsulating shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, phosphate or carbonate. The present disclosure also provides a core-shell particle comprising: a core comprising at least one active agent,

an intermediate polymer shell encapsulating said core; ions of an additive disposed on the surface of said intermediate polymer shell; and

an encapsulating shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, phosphate or carbonate.

The present disclosure also relates to a method of forming a core-shell particle, comprising the steps of:

(b) encapsulating a shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, carbonate or phosphate, onto said core to thereby form said core-shell particle.

Step (a) may comprise adding an additive to the core for controlling the size and/or density of the core particles.

Step (a) may comprise adding an additive to the core, wherein ions of the additive have antagonistic charge and/or affinity interactions with the core particles for controlling the size and/or density of the core. Ions of the additive may repel additional core particles from adhering to the surface of the core, thereby controlling the size and/or density of the core.

Step (a) may comprise adding a polymer, thereby forming an intermediate polymer shell disposed between the core and the additive ions. The ions of the additive may adhere to the surface of the intermediate polymer shell, thereby forming a negatively charged shell. Step (a) may comprise conjugating ions of the additive to the surface of the core, or conjugating ions of the additive to the intermediate polymer shell.

Step (a) may comprise adhering ions of the additive to the surface of the core, or conjug adhering ions of the additive to the intermediate polymer shell, wherein the additive ions are adhered via charge-charge interactions or intermolecular bonding forces such as van der Waals forces or hydrophobic interactions.

In one embodiment, citrate ions adhere to the surface of precipitated chondroitin sulfate particles through an intermediate PLL layer, forming a negatively charged layer/hydration shell that may repel additional negatively charged chondroitin sulfate molecules from adhering to the surface of the chondroitin sulfate particle, hence restricting the growth of the chondroitin sulfate particle.

The final size of the core may be determined by the ratio of additives to actives in solution. The ratio of additive to active agent may be in the range of about 100: 1 to about 1: 100, or about 90: 1 to about 1: 100, or about 80: 1 to about 1: 100, or about 70: 1 to about 1 : 100, or about 60: 1 to about 1: 100, or about 50: 1 to about 1: 100, or about 40: 1 to about 1: 100, or about 30: 1 to about 1: 100, or about 20: 1 to about 1: 100, or about 10: 1 to about 1: 100, or about 100: 1 to about 1:90, or about 100: 1 to about 1:80, or about 100: 1 to about 1:70, or about 100: 1 to about 1 :60, or about 100: 1 to about 1:50, or about 100: 1 to about 1:40, or about 100: 1 to about 1:30, or about 100: 1 to about 1:20, or about 100: 1 to about 1: 10, about 90: 1 to about 1:90, about 80: 1 to about 1:80, about 70: 1 to about 1:70, about 60: 1 to about 1:60, about 50: 1 to about 1:50, about 40: 1 to about 1:40, about 30: 1 to about 1:30, about 20: 1 to about 1:20, about 10: 1 to about 1: 10, about 5: 1 to about 1: 10, or any value or range therebetween. The polydispersity index (PDI) of the core may be in the range of about 0.10 to about 0.20, or about 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20.

Step (a) may comprises adding at least one active agent to a solution, forming a saturated solution of active agent, thereby driving precipitation of the active agent from said solution. The solution may be an acidic solution. The acidic solution may be HC1. The pKa of the solution may be about 2.0 to about 2.54, or about 2.0 to about 2.5, about 2.0 to about 2.4, about 2.0 to about 2.3, about 2.0 to about 2.2, about 2.0 to about 2.1, about 2.1 to about 2.54, about 2.2 to about 2.54, about 2.3 to about 2.54, about 2.4 to about 2.54, about 2.5 to about 2.54, about 2.1 to about 2.5, about 2.2 to about 2.4, or about 2.0, about 2.1, about 2.2, about 2.3, about 2.4, about 2.5, about 2.54, or any range or value therebetween. The additive of step (a) may control the size of the precipitation product. The additive may control the size of the crystallization product with antagonistic charge/affinity interactions with the surface of the crystal that shields and inverts the surface properties of the crystalline particles (for example, by charge inversion and phase inversion). The additive may prevent further crystal growth with the final size of the product determined by the ratio of additives to actives in solution.

The precipitation product may be a crystallization product,

Step (a) may comprise adding an additive, wherein ions of the additive have antagonistic charge and/or affinity interactions with the core particles for controlling the size and/or density of the core. The additive may be a surface chemistry antagonist. The additive may be a citrate salt. The citrate salt may be sodium citrate. The ions of the additive may be citrate ions.

Step (b) may comprise a polycondensation reaction, a gelation reaction or an in-situ polymerization reaction. Step (b) may comprise a sol gel reaction in a solution comprising silicon alkoxide. The silicon alkoxide may be selected from the group consisting of tetramethoxy silane, tetraethoxy silane, and tetraethyl orthosilicate. The resulting encapsulation layer may be a cross-linked encapsulation layer. The shell may comprise an encapsulation matrix. Advantageously, the shell may provide for robust and tunable release kinetics, superior mechanical properties from crosslinking, and be biodegradable and resorb-able. The silanes may be selected from the group consisting of tetramethoxy silane, tetraethoxy silane, tetraethyl orthosilicate, trie thoxy (ethyl) silane, aminopropyltrimethoxy silane, methyltrimethoxy silane, phenyltriethoxy silane.

The method of the present disclosure may further comprise a step (c) functionalizing the shell with a secondary functionalization, wherein step (c) occurs after step (b). The secondary functionalization may be provided by adding a compound selected from the group consisting of polyethylene glycol (PEG), pH sensitive PEG, affibodies, antibodies, IgG, IgM, targeting ligands like VEGF-C, cRGD and folate, DNA and aptamers, proteins, lipoproteins, apolipoproteins, glycoproteins, glycans, carbohydrates, saccharides and oligosaccharides, polymers and oligomers, lipids.

The secondary functionalization may be via a bioconjugation. Broadly, bioconjugation can be achieved via any of the commonly used bioconjugation techniques such as EDC/sNHS, maleimide/thiol, biotin/avidin etc to attach the biomolecule onto the encapsulation matrix.

In the method of the present disclosure, the polymer of the intermediate polymer shell and additive may be added to the active agent consecutively or simultaneously.

In the method of the present disclosure, step (b) may be performed at least twice. This may allow for layer-by-layer encapsulation.

(a) providing a core comprising at least one chondroitin salt, said core having citrate ions surrounding a surface of said core;

(b) encapsulating a shell comprising Si0₂ onto said core to thereby form said core-shell particle. The present disclosure also provides a method of forming a core-shell particle, comprising the steps of:

(b) encapsulating a shell comprising Si0₂ onto said core to thereby form said core-shell particle.

There is also provided a method of forming a core-shell particle, comprising the steps of:

(a) providing a core comprising at least one active agent, said core having ions of an additive disposed on a surface of said core; and (b) encapsulating a shell comprising (i) a polymer; and/or (ii) a metal or a semi-metal oxide, carbonate or phosphate, onto said core to thereby form said core-shell particle.

(a) providing a core comprising at least one active agent,

(ai) encapsulating said core with an intermediate polymer shell having ions of an additive disposed on a surface of said intermediate polymer shell; and

There is also provided a method of forming a core-shell particle, comprising the steps of: (a) providing a core comprising at least one chondroitin salt,

(ai) encapsulating said core with an intermediate polymer shell comprising PLL having citrate ions disposed on a surface of said intermediate polymer shell; and

(b) encapsulating a shell comprising Si0₂, onto said core to thereby form said core-shell particle.

The present disclosure also provides a method of forming a core-shell nanoparticle, wherein said core comprises at least one active agent, and said shell comprises one or more polymers and/or metal oxides, comprising the steps of:

(a) forming nanoparticles of the at least one active agent;

(b) adding an additive for controlling the size and/or density of the nanoparticles;

(c) adding one or more polymers and/or metal or semi-metal alkoxides; and

(d) encapsulating the nanoparticles with a shell of polymer and/or metal or semi-metal oxides. The present disclosure also provides a method of forming a core-shell nanoparticle, wherein said core comprises chondroitin salt, and said shell comprises at least one polymer, comprising the steps of:

(a) forming nanoparticles of chondroitin salt;

(b) adding a citrate salt for controlling the size and/or density of the nanoparticles;

(c) adding at least one semi-metal alkoxides; and

(d) encapsulating the nanocrystalline particles with a shell comprising semi-metal oxide. The present disclosure further provides a core-shell particle produced by the method disclosed herein.

The present disclosure also provides a core-shell particle disclosed herein for use in therapy. The core-shell particle may be used for drug delivery.

The present disclosure also provides core-shell particles for use in sustained release formulations.

The core-shell particles may be useful in self-healing materials, self-cleaning surfaces and/or an ti -bacterial surfaces.

Brief Description of Drawings

The accompanying drawings illustrate a disclosed embodiment and serves to explain the principles of the disclosed embodiment. It is to be understood, however, that the drawings are designed for purposes of illustration only, and not as a definition of the limits of the invention.

Fig. l

[Fig. 1] is a general reaction scheme for the method of preparing core-shell particles according to the invention.

Fig. 2

[Fig. 2] is an exemplary reaction scheme for the synthesis of hybrid crystalline chondroitin sulfate particles of the present invention. Fig. 3

[Fig. 3] is a particle characterization of hybrid crystalline chondroitin sulfate (HNCS) particles. A) SEM of HNCS. B) TEM of HNCS showing uniform spherical morphology. C) Particle size measurements via TEM show average particle diameter of 240 ± 70 nm. D) Release profile of HNCS tagged with fluorescein isothiocyanate (FITC). Scale bars represent Ιμηι.

Fig. 4

[Fig. 4] is a graph comparing the release profiles of FITC conjugated HNCS prepared with single layer of encapsulation via silica sol-gel, and FITC conjugated HNCS prepared with double layer of encapsulation via silica sol-gel .

Fig. 5

[Fig. 5] is a graph showing the drug loading efficiency and particle size of core-shell particles of the present invention with varying concentrations of acid used in the precipitation of the drug particle.

Fig. 6

[Fig. 6] is a transmission electron microscopy (TEM) of single layer encapsulated HNCS particles after 2 hours of drug release.

Fig. 7

[Fig. 7] shows FITC labeled DNA particles prepared in the same fashion as HNCS, conjugated with PEG and aE-cadherin and incubated with LNCAP cells for 15 mins on ice. Cells were recovered and washed and analysed via fluorescence microscopy. Arrows indicate DNA nanoparticles that have attached to the LNCAP cells. No nanoparticles were observed in control cell populations without aE-cadherin.

Fig. 8

[Fig. 8] shows heparin sulphate particles prepared in the same fashion as HNCS. Nanocrystalline HS is able to increase BMP-2-mediated ALP expression in a dose- dependent manner. At the lower BMP-2 dose, soluble HS has a greater effect on BMP-2 at higher doses (5, 10, 20μg/mL). At the higher BMP-2 dose, both soluble and encapsulated HSpm display similar levels of enhancement in ALP activity. Detailed Description of Drawings

Referring to Fig. 1, crystalline particles of at least one active agent (1) are first prepared, thereby forming a crystalline active core (2). An additive (3) for controlling the size and/or density of the crystalline particles is then added. One or more polymers and/or metal oxides (4) is added, followed by the encapsulation of the crystalline active core with the one or more polymers and/or metal oxides, thereby forming a core-shell particle (5).

Referring to Fig. 2, crystalline chondroitin sulphate was first prepared from chondroitin sulphate (1 ') through pH crystallisation, thereby forming the active core (2'). Sodium citrate (3 ') was then added to form citrate capped chondroitin sulphate (3 "). Poly-L-lysine (PLL) (4') and tetramethoxy silane (6') were then added to the mixture, followed by encapsulation of the active core via sol-gel condensation of tetramethoxy silane, thereby forming a core-shell particle (5').

Referring to Fig. 4, FITC conjugated HNCS was prepared with either a single or double layer of encapsulation via a silica sol-gel reaction, and the release characteristics were studied over 40 days. Single layer encapsulated HNCS showed complete release over 4 hours, while double layer encapsulated HNCS showed release over 40 days.

Referring to Fig. 5, 0.1 to 0.8M HC1 was used in the preparation of chondroitin sulfate particles, and the resulting particle size and drug loading efficiency into the particles was tested. Particle size was observed to increase with increasing HC1 concentration, up to a maximum of 530nm in 0.8M HC1. Drug loading efficiency was observed to be greater than 99.5% efficiency for all conditions tested.

Referring to Fig. 6, prepared HNCS particles were dialysed with a 300kDa MWCO membrane and particles were recovered after 2 hours and analysed via TEM and compared to control HNCS particles without staining. Release particles showed much lower contrast than control HNCS particles which may be due to the leaching of chondroitin sulfate from the particles. Examples

Non-limiting examples of the invention and a comparative example will be further described in greater detail by reference to specific Examples, which should not be construed as in any way limiting the scope of the invention. Example 1

A method for the synthesis of hybrid crystalline particles with sustained release of actives from the nanoparticles is disclosed. The hybrid particle comprises two components; a crystalline core comprising of actives that have been condensed out of solution in a controlled manner with good size control and dispersity, and an encapsulating shell of either metal oxides or polymers with varying degrees of crosslinking and molecular weight allowing for sustained release of the crystalline active.

1 wt% of chondroitin sulfate was first prepared, then diluted to 0.2 wt% in 100 mM HC1 in 1ml reaction volumes in 2ml Eppendorf tubes and mixed vigorously for 20 seconds then left to stand for 30 minutes at room temperature. 100 μΐ of 0.01 wt% poly-L-lysine (PLL) and 10 μΐ of 10 wt% sodium citrate were then added to the solution and mixed vigorously for 20 seconds and left to stand for 60 minutes at room temperature. 5 μΐ of tetramethoxy silane was then added to the mixture and mixed vigorously for 120 minutes. 800ul of ethanol was then added to the reaction mixture and the solid product was recovered as a pellet that was then resuspended in 1 ml ethanol. A further 100 μΐ of 0.01 wt% PLL and 5 μΐ of tetramethoxy silane was added to the mixture and mixed vigorously for 120 minutes. The solid product was recovered as a pellet and washed and resuspended in ethanol.

The pellet was dried and characterized via SEM and TEM (Fig. 2). The SEM and TEM measurements show uniform particle size and morphology with particles taking on a spherical morphology with an average diameter of 240 + 70 nm.

Using a 200 nm diameter for the crystalline core and 2 mg/ml initial solution concentration of chondroitin sulfate and 1.5g/ml density of condensed chondroitin sulfate, this method potentially gives a 750 times increase in loading efficiency over traditional passive loading techniques. Example 2a - Secondary functionalization lOOx excess of tris-carboxyethylphosphine (TCEP) was added to a protein/peptide/enzyme/antibody of interest in degassed buffer. This was then reacted with a lOx molar excess of a silica maleimide crosslinker on ice to crosslink the biomolecule to the silica matrix.

Example 2b aE-cadherin was first reacted with N-hydroxysulfosuccinimide(NHS)-PEG-Silane for 5 minutes on ice at 1: 1 molar ratio of antibody to silane, then transferred to a lmg/ml solution of silica coated nanoparticles and allowed to react for a further 45 minutes on ice. Particles were washed twice then resuspended in PBS.

Industrial Applicability

The core-shell particles and methods of the present disclosure may be useful in the medical and cosmetic fields.

The size and/or density of the crystalline core of the core-shell particles may be controlled by the addition of the additive, thereby allowing size control of the final product. By being able to achieve good size control, enhanced pharmaceutical properties of the encapsulated active may be obtained. Further advantageously, sustained release profiles of the active may be obtained which allows for controlled drug delivery. Direct encapsulation of the crystalline active may also allow for ultra-high efficiency loading and encapsulation of the active agent. Thus, a high loading of active agent may be achieved in the core. The method of the invention may produce core-shell particles where there is even particle morphology and size distribution in the core and improved formulation stability and bio-availability. The method of the invention may also produce core-shell particles that are bio- compatible and bio-degradable. The method may allow for cost-efficient production of the core- shell particles.

The core-shell particles may exhibit sustained release profiles of the active which allows for controlled drug delivery. Due to the core-shell nature of the particles, the active agent encapsulated in the core by the shell may protect the core contents from early and undesired release in the body. The core-shell particles may be used in drug delivery.

It will be apparent that various other modifications and adaptations of the invention will be apparent to the person skilled in the art after reading the foregoing disclosure without departing from the spirit and scope of the invention and it is intended that all such modifications and adaptations come within the scope of the appended claims.

Claims

A core-shell particle comprising:

a core comprising at least one active agent, said core having ions of an additive surrounding the surface of said core; and

The particle according to claim 1, wherein ions of the additive have antagonistic charge and/or affinity interactions with a core particle.

The particle according to claim 1 or 2, wherein the particle comprises an intermediate polymer shell disposed between the core and additive ions.

The particle according to claim 3, wherein ions of the additive adhere to the surface of the intermediate polymer shell.

The particle according to claim 3 or 4, wherein the ions of the additive and the intermediate polymer shell form a negatively charged shell.

The particle according to any one of claims 3 to 5, wherein the polymer of the intermediate polymer shell is selected from the group consisting of at least one poly-L-lysine (PLL), polyethyleneimine (PEI), dextran, polyelectrolytes, polycaprolactames (PCL), alginates, and chitosan.

7. The particle according to any one of claims 1 to 6, wherein the additive comprises one or more citrates, and/or polyethylene glycol (PEG).

8. The particle according to any one of claims 1 to 7, wherein the encapsulating shell is selected from the group consisting of:

(c) shell comprising or consisting of semi-metal oxide, phosphate or carbonate;

9. The particle according to any one of claims 1 to 8, wherein the metal or semi-metal oxide, phosphate or carbonate is selected from the group consisting of Si0₂, calcium carbonate, calcium phosphate, Ti0₂ and iron oxides, preferably Fe₂0₃ or FeO.

10. The particle according to any one of claims 1 to 9, wherein the polymer of the encapsulating shell is selected from the group consisting of at least one poly-L-lysine (PLL), polyethyleneimine (PEI), dextran, polyelectrolytes, polycaprolactames (PCL), alginates, chondroitin and polymerized silicon alkoxide.

11. The particle according to any one of claims 1 to 10, wherein the encapsulating shell comprises or consists of Si0₂.

12. The particle according to claim 11, wherein the Si0₂ may be cross-linked Si0₂.

13. The particle according to any one of claims 1 to 12, wherein the active agent is a therapeutic agent or a cosmetic agent.

14. The particle according to any one of claims 1 to 13, wherein the active agent is selected from the group consisting of chondroitin salts, heparin salts, glycosaminoglycans, glycoproteins, biomolecules, chondroitin sulfate, and heparin sulfate.

15. The particle according to any one of claims 1 to 14, wherein the core has an average diameter of 10 nm to 1000 nm.

16. The particle according to claim 15, wherein the core has an average diameter of 130 nm to 270 nm.

17. A core-shell particle comprising:

a core comprising at least one chondroitin salt, said core having ions of an additive comprising at least one citrate ion surrounding the surface of said core; and

an encapsulating shell comprising Si0₂.

18. A method of forming a core-shell particle, comprising the steps of:

19. The method of claim 18, wherein step (a) comprises adding an additive to the core for controlling the size and/or density of the core.

20. The method of claim 18 or 19, wherein ions of the additive have antagonistic charge and/or affinity interactions with a core particle for controlling the size and/or density of the core.

21. The method of claim 20, wherein ions of the additive repel additional core particles from adhering to the surface of the core, thereby controlling the size and/or density of the core.

22. The particle of any one of claims 18-21, wherein step (a) comprises adding a polymer, thereby forming an intermediate polymer shell disposed between the core and the additive ions. The particle of claim 22, wherein ions of the additive adhere to the surface of the intermediate polymer shell, thereby forming a negatively charged shell.

24. The method of any one of claims 19 to 23, wherein the ratio of additive to active agent is in the range of 100: 1 to 1: 100.

25. The method of claim 24, wherein the ratio of additive to active agent is in the range of 5: 1 to 1 : 10.

26. The method of any one of claims 18 to 26, wherein the core has an average diameter of 10 nm to 1000 nm.

The method of claim 26, wherein the core has an average diameter of 130 nm to 270nm.

The method of any one of claims 18 to 27, wherein step (a) comprises adding at least one active agent to a solution, forming a saturated solution of active agent, thereby driving precipitation of the active agent from said solution.

29. The method of claim 28, wherein the solution is selected from the group consisting of hydrochloric acid (HC1), ethanol, isopropanol and sodium hydroxide.

30. The method of any one of claims 18 to 29, wherein step (b) comprises a polycondensation reaction, a gelation reaction or an in-situ polymerization reaction.

31. The method of claim 30, wherein step (b) comprises a sol gel reaction in a solution comprising silicon alkoxide.

32. The method of any one of claims 18 to 31, wherein the polymer of step (b) is selected from the group consisting of poly-L-lysine (PLL), polyethyleneimine (PEI), dextran, polyelectrolytes, polycaprolactames (PCL), alginates, chondroitin and polymerized silicon alkoxide.

The method of claim 31 or 32, wherein the silicon alkoxide is selected from the group consisting of tetramethoxy silane, tetraethoxy silane, tetraethyl orthosilicate, triethoxy (ethyl) silane, aminopropyltrimethoxy silane, methyltrimethoxy silane, and phenyltriethoxy silane.

The method of any one of claims 18 to 33, further comprising a step (c) functionalizing the shell with a secondary functionalization, wherein step (c) occurs after step (b).

The method of claim 34, wherein the secondary functionalization is provided by adding a compound selected from the group consisting of polyethylene glycol (PEG), pH sensitive PEG, affibodies, antibodies,

IgG, IgM, targeting ligands like VEGF-C, cRGD and folate, DNA and aptamers, proteins, lipoproteins, apolipoproteins, glycoproteins, glycans, carbohydrates, saccharides and oligosaccharides, polymers and oligomers, lipids.

36. The method of any one of claims 18 to 35, wherein the active agent is a therapeutic agent, or a cosmetic agent.

37. The method of any one of claims 18 to 36, wherein the active agent is selected from the group consisting of chondroitin salts, heparin salts, glycosaminoglycans, glycoproteins, biomolecules, chondroitin sulfate, and heparin sulfate.

38. The method of any one of claims 19 to 37, wherein the additive is selected from the group consisting of citrates, and/or polyethylene glycol.

39. The method of claim 38, wherein the citrate is sodium citrate.

40. The method of any one of claims 18 to 39, wherein step (b) is performed at least twice, thereby forming at least two shell layers.

41. A method of forming a core-shell particle, comprising the steps of:

42. A core-shell particle produced by the method of any one of claims 18 to 41.

43. A core-shell particle according to any one of claims 1-17 and 42 for use in therapy.

44. A core-shell particle according to any one of claims 1-17 and 42 for use in sustained release formulations.