Near-infrared and red light absorbing composite resin particles

Abstract

Composite resin particles comprising a dye and a resin selected from the group consisting of a polyamino acid, a polyphosphazene, a polysaccharide derivative, a polyester, a polyorthoester, a polycyanoacrylate and a copolymer thereof, characterized in that the composite resin particles have a maximum absorption in the range of 600 nm to 750 nm and an average particle size of the particles is between 0.03 [mu] m and 3 [mu] m. The composite resin particles are suitable for photomedical applications, such as phototherapy, including photothermal therapy (PTT), photodynamic therapy (PDT), chemical photodynamic nanotherapy, and fluorescent medical imaging.

Description

Technical Field

[0001] The present invention relates to a biocompatible organic nanoparticle and microparticle design for photomedical applications such as phototherapy, including photothermal therapy (PTT), photodynamic therapy (PDT), chemophotodynamic therapy, and fluorescence medical imaging. Background Art

[0002] Red and near-infrared (NIR) laser technologies are becoming increasingly important in non-invasive treatments of different diseases and medical diagnostics, including photothermal therapy, photodynamic therapy, fluorescence imaging, and photoacoustic imaging. Several of these technologies rely on NIR-absorbing nanoparticles, among which inorganic nanoparticles are commonly used, such as gold nanomaterials, carbon nanomaterials (including carbon nanotubes), metal sulfides, metal oxides, and different upconversion nanoparticles.

[0003] The use of inorganic photothermal conversion nanoparticles has been extensively reviewed (Raza et al., Journal of Materials Research and Technology, 8(1), 1497-1509 (2019); Wang et al., International Journal of Nanomedicine, 15, 1903-1914 (2020)). Although these nanoparticles have excellent NIR response, they are not biodegradable and there is a risk of bioaccumulation and long-term residence in the body, which may potentially increase the possibility of their long-term toxicity. Therefore, nanoparticles based on biocompatible organic NIR absorbers are highly favored.

[0004] Although several classical organic NIR absorbers have a good track record in photomedical applications, cyanine dyes are a particularly preferred class of NIR absorbers (hereinafter referred to as NIR-absorbers) due to their high molar extinction coefficients. The high molar extinction of NIR lasers has the advantage that the amount of NIR absorber required can be reduced and enables applications in in vitro and in vivo imaging and the treatment of deep diseases such as tumors. One of the best-known and FDA-approved cyanine dyes is indocyanine green (ICG).

[0005] NIR absorbers such as indocyanine green must be integrated into nanoparticles to extend their lifetime in vivo, disperse in body fluids, prevent photobleaching, and enhance their tumor targeting ability.

[0006] US2021154335A discloses indocyanine green in PEI resin nanoparticles.

[0007] Zeng et al. in Mol. Pharmaceutics 2012, 514-522 disclose nanoparticles of ICG, which form J-aggregates and which are encapsulated by PL-PEG-mAB or PL-PEG polymers.

[0008] Several nanoparticle designs have been proposed and recently reviewed in the prior art (Zhu et al., Biomater. Sci, 6, 746-765 (2018); Zhu et al., Current Medicinal Chemistry, 26, 1389-1405 (2019); Ng. et al., Chemical reviews, 115, 11012-11042 (2015); Shao et al., RSC Nanoscience & Nanotechnology, 40, 125-157 (2016)). The proposed nanoparticle designs generally require laborious synthesis protocols, which hinders the easy adjustment of nanoparticles for different applications. Therefore, there is a need for cyanine dye-containing nanoparticles that can be obtained via simple and scalable industrial techniques. One of these simple and scalable industrial techniques is solvent evaporation.

[0009] However, cyanine dyes such as indocyanine green are generally incompatible with industrial and easily scalable techniques such as solvent evaporation, and it is impossible to physically integrate them into biocompatible nanoparticles. Indocyanine green is insoluble or has very limited solubility in the solvents currently used in solvent evaporation techniques. Due to its partial water solubility and degradation in water, the indocyanine green dye must be fully encapsulated to avoid any contact with water. This requirement further limits the availability of technologies for making fully encapsulated indocyanine green nanoparticles and microparticles.

[0010] However, encapsulation of NIR absorbing dyes mostly induces a shift of the spectrum towards longer wavelengths. Due to the peak absorbance position of many cyanine dyes and the red shift towards higher wavelengths due to encapsulation, many known dyes (such as IR700) are insensitive to light from red lasers emitting between 600 and 750 nm.

[0011] In addition to NIR lasers, these red lasers also represent the basis of a large installed base of imaging sources in phototherapy and fluorescence medical imaging. Therefore, dyes with absorption wavelengths between 580 nm and 750 nm must be encapsulated to obtain nanoparticles suitable for phototherapy and fluorescence imaging with the aid of red and NIR lasers.

[0012] Therefore, there is a need to integrate red and near-infrared (hereinafter referred to as R / NIR) absorbing dyes having maximum absorption between 580 nm and 750 nm and having high molar extinction coefficients into biocompatible nanoparticles using industrial solvent evaporation techniques. Summary of the invention

[0013] An object of the present invention is to provide a biocompatible polymer nanocapsule or microcapsule containing a specific R / NIR absorber as defined in claim 1 .

[0014] Still another aspect of the present invention provides an aqueous dispersion of composite resin particles as defined in claim 1. The aqueous dispersion is defined in claim 11.

[0015] According to another aspect, the present invention comprises an industrially scalable method of integrating a specific class of R / NIR absorbers with biocompatible polymers, as defined in claim 16 .

[0016] Other features, elements, steps, characteristics and advantages of the present invention will become more apparent from the following detailed description of preferred embodiments of the present invention. Specific embodiments of the present invention are also defined in the dependent claims. DETAILED DESCRIPTION

[0017] The object of the present invention is achieved by composite resin particles comprising a biocompatible resin and a dye, characterized in that the resin particles have a maximum absorption at 600nm to 750nm, more preferably 600nm to 700nm, most preferably 610nm to 650nm, and the average particle size of the particles is between 0.03μm and 3μm. The maximum absorption of the composite resin particles is measured for an aqueous dispersion of the composite resin particles (concentration of 0.19mg / mL).

[0018] A. Composite resin particles

[0019] A.1. Dyes

[0020] The dye present in the composite resin particles according to the present invention can be selected from any known dye class, and when incorporated into the composite resin particles, the dye is suitable for achieving maximum absorption at 600nm to 750nm, more preferably 600nm to 700nm, and most preferably 610nm to 650nm. Typical dye classes are selected from Industrial Dyes, Chemistry, Properties, Applications (Wiley-VCH, ISBN 3-527-30426-6), edited by Klaus Hunger; Color Chemistry, Syntheses, Properties and Applications of Organic Dyes and Pigments (Heinrich Zollinger, VCH, ISBN 0-89573-421-4); and The Chemistry and Application of Dyes (Plenum Press, ISBN 0-306-43278-1), edited by David R. Waring and Geoffrey Hallas.

[0021] In practice, it appears that the dye preferably has an absorption maximum between 580 nm and 750 nm.

[0022] The dye may be selected from azo dyes, phthalocyanines, anthraquinones, indigoid dyes, cyanines, merocyanines, oxonoles, triarylmethane dyes, azomethine dyes and indoaniline dyes. Cyanine dyes are particularly preferred.

[0023] In a first particularly preferred embodiment, the dye according to the invention is a squarilium dye according to the general formula I

[0024]

[0025] in

[0026] R1 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group.

[0027] R2 and R3 are independently selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted aryl or heteroaryl groups, halogen, ester, amide, ether, nitro, nitrile, amine, ketone and aldehyde.

[0028] R2 and R3 may represent the necessary atoms to form a five-membered or six-membered ring.

[0029] X is selected from S, O, NR4 and CR5R6

[0030] R4, R5 and R6 are independently selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups and substituted or unsubstituted aryl or heteroaryl groups.

[0031] R5 and R6 may represent atoms necessary to form a five- to eight-membered ring.

[0032] In a further preferred embodiment, R2 and R3 represent the atoms necessary to form a six-membered aromatic ring, optionally also with a further aromatic ring.

[0033] In an even further preferred embodiment, R1 and R4 are selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups and substituted or unsubstituted alkynyl groups, more preferably substituted or unsubstituted alkyl groups. In an even further preferred embodiment, R5 and R6 independently represent substituted or unsubstituted alkyl groups, more preferably unsubstituted alkyl groups, most preferably methyl groups. In a particularly preferred embodiment, R1 and R4 represent substituted or unsubstituted branched alkyl groups. Branched alkyl chains are defined as alkyl chains containing at least one secondary or tertiary carbon atom.

[0034] Typical examples of the squarylium dye according to the present invention are given in Table 1, but are not limited thereto.

[0035] Table 1

[0036]

[0037]

[0038]

[0039] In a further preferred embodiment, the dye present in the composite resin particles according to the invention is a pentamethine cyanine dye according to the general formula II

[0040]

[0041] in

[0042] R1 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group.

[0043] R2 and R3 are independently selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted aryl or heteroaryl groups, halogen, ester, amide, ether, nitro, nitrile, amine, ketone and aldehyde.

[0044] R2 and R3 may represent the necessary atoms to form a five-membered or six-membered ring.

[0045] R4 is selected from hydrogen, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group.

[0046] X is selected from S, O, NR5 and CR6R7

[0047] R5, R6 and R7 are independently selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups and substituted or unsubstituted aryl or heteroaryl groups.

[0048] R6 and R7 may represent atoms necessary to form a five- to eight-membered ring.

[0049] In a further preferred embodiment, R2 and R3 represent the atoms necessary to form a six-membered aromatic ring, optionally also with a further aromatic ring.

[0050] In an even further preferred embodiment, R1 and R5 are selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups and substituted or unsubstituted alkynyl groups, more preferably substituted or unsubstituted alkyl groups. In an even further preferred embodiment, R6 and R7 independently represent substituted or unsubstituted alkyl groups, more preferably unsubstituted alkyl groups, most preferably methyl groups. In a particularly preferred embodiment, R1 and R5 represent substituted or unsubstituted branched alkyl groups. In another preferred embodiment, R4 is selected from hydrogen and substituted or unsubstituted alkyl groups, more preferably hydrogen and lower alkyl groups, most preferably hydrogen.

[0051] Typical pentamethine cyanine dyes according to formula II are given in Table 2, but are not limited thereto.

[0052] Table 2.

[0053]

[0054]

[0055] In a most preferred embodiment, the dye present in the composite resin particles according to the invention is a cyanine dye according to formula III.

[0056]

[0057]

[0058] in

[0059] R1 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group.

[0060] R2 and R3 are independently selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted aryl or heteroaryl groups, halogen, ester, amide, ether, nitro, nitrile, amine, ketone and aldehyde (check)

[0061] R2 and R3 may represent the necessary atoms to form a five-membered or six-membered ring.

[0062] R4 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group.

[0063] X is selected from S, O, NR5 and CR6R7

[0064] R5, R6 and R7 are independently selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups and substituted or unsubstituted aryl or heteroaryl groups.

[0065] R6 and R7 may represent atoms necessary to form a five- to eight-membered ring.

[0066] In a further preferred embodiment, R2 and R3 represent the atoms necessary to form a six-membered aromatic ring, optionally also with a further aromatic ring.

[0067] In an even further preferred embodiment, R1 and R5 are selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups and substituted or unsubstituted alkynyl groups, more preferably substituted or unsubstituted alkyl groups.

[0068] In an even further preferred embodiment, R6 and R7 independently represent substituted or unsubstituted alkyl groups, more preferably unsubstituted alkyl groups, most preferably methyl groups. In a most preferred embodiment, R4 is selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups and substituted or unsubstituted alkenyl groups, more preferably substituted or unsubstituted alkyl groups. In a particularly preferred embodiment, R1, R4 and R5 represent substituted or unsubstituted branched alkyl groups.

[0069] Typical examples of cyanine dyes according to the general formula III are given in Table 3, but are not limited thereto.

[0070] Table 3.

[0071]

[0072] Examples of other embodiments according to the present invention are given in Table 4, but are not limited thereto.

[0073] Table 4.

[0074]

[0075]

[0076]

[0077] In addition to the ability to convert light into heat, the dyes according to the general formulae I, II, III and the dyes of Table 4 also exhibit fluorescence, which makes them suitable also for fluorescence medical imaging.

[0078] A.2. Biocompatible resins

[0079] The biocompatible resin present in the composite resin particles according to the present invention is a resin selected from the group consisting of polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof.

[0080] The biodegradable and biocompatible resin may be selected from the resins discussed in Biodegradable Polymers, PBM Series, Vol. 2 (Citus Books 2003, ISSN 1479-1285).

[0081] The polyester is preferably polycaprolactone, polylactic acid, poly-L-lactic acid, polyglycolic acid, poly(lactic acid-co-glycolic acid), poly(L-lactic acid-co-glycolic acid), poly-D,L-lactide, and any derivatives and / or combinations thereof.

[0082] A.3. Resin particle structure

[0083] The two basic components of the composite resin particles according to the present invention, namely the dye (preferably a dye having an absorption wavelength between 580 nm and 750 nm) and the resin selected from polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof, can be arranged uniformly or non-uniformly within the resin particles.

[0084] In a preferred embodiment of the present invention, the composite resin particles are capsules comprising a core and a shell, wherein the core contains a dye, preferably a dye with an absorption wavelength between 580nm and 750nm, and the shell comprises a resin selected from polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof. The advantage of encapsulating dyes is that the life of the dye increases because the dye is better protected from the influence of the water environment, thereby avoiding the bleaching and hydrolysis of the dye, while making the dye colloidally stable in a physiological medium. In a more preferred embodiment of the present invention, the composite resin particles are capsules, wherein the core contains a dye, preferably a dye with an absorption wavelength between 580nm and 750nm, and the shell comprises polyamino acids. Capsules can be obtained by interfacial polymerization of N-carboxyl-anhydride monomers according to general structure IV.

[0085]

[0086] in

[0087] n represents 0 or 1

[0088] R1, R2 and R3 are selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted alkaryl groups and substituted or unsubstituted aryl or heteroaryl groups.

[0089] R1, R2 and R3 may represent the necessary atoms to form a five-membered to eight-membered ring

[0090] In a preferred embodiment, n represents 0. In a particularly preferred embodiment, R3 represents hydrogen or an alkyl group, most preferably hydrogen.

[0091] In another preferred embodiment, R1 and R2 are selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted alkaryl groups and substituted or unsubstituted aryl groups.

[0092] In a further preferred embodiment, the N-carboxyl-anhydride monomer according to the general structure is selected from glycine derivatives, alanine derivatives, leucine derivatives, phenylalanine derivatives, phenylglycine derivatives, valine derivatives, glutamic acid derivatives, aspartic acid derivatives, lysine derivatives, ornithine derivatives, histidine derivatives, methionine derivatives, cysteine ​​derivatives, arginine derivatives, tryptophan derivatives, cysteine ​​derivatives, isoleucine derivatives, tyrosine derivatives, proline derivatives and serine derivatives. Both D- and L-amino acid derivatives and mixtures thereof can be used.

[0093] The ratio of the weight of the dye to the weight of the resin is preferably 0.5 wt % to 10 wt %, more preferably 1 wt % to 5 wt %. When the weight of the dye is below this range, the thermal conversion or fluorescence obtained will be insufficient. When the weight of the dye is above the above range, the dye may leak from the core during the shrinkage process of the core (in a rotary evaporator), or some of the walls are thinner and break or leak the dye due to poor (uneven) spheroid formation, thereby forming free dye crystals in the dispersion over time.

[0094] The average particle size of the composite resin particles of the present invention is preferably 0.01 μm to 10 μm, more preferably 0.03 μm to 5 μm, and most preferably 0.03 μm to 3 μm. Particles with an average particle size higher than these upper limits are difficult to insert into animals or humans via a syringe. Particles with an average particle size lower than these lower limits are difficult to prepare, and due to the very large surface area, they partially lose their protective function. Other different nano effects may also occur. Such as agglomeration due to a large surface area, especially later in physiological media (osmotic pressure, blood / buffer charge and particle surface balance mechanism. In biomedicine, in order to cross the blood-brain barrier and enter cells, it is necessary for the particles to be less than 200 nm, and depending on the construction of the cell wall, some particles must even be far less than 100 nm.

[0095] A.4. Pharmaceutically active compounds

[0096] The composite resin particles according to the present invention are also suitable for on-demand drug release, where the drug is released upon heating of the particles with the aid of a suitable light source emitting light at a wavelength of 600 to 750 nm. It is therefore useful to incorporate a drug compound to achieve such on-demand drug release.

[0097] Sometimes, PTT or PDT cannot completely destroy cancer cells and may result in the survival of residual cells after photothermal treatment. Therefore, it is useful to incorporate anticancer drugs to enhance chemotherapy. When red light or NIR light is applied to the composite particles, the drug will be released due to the heat generated, thereby initiating a synergistic chemo-photothermal therapy. The anticancer drug should preferably be soluble in the water-immiscible solvent used in the preparation of the composite resin particles (see §B.).

[0098] Anticancer drugs suitable for incorporation into the particles of the present invention are cytostatics. Cytostatics for treating cancer can be selected from alkylating agents, anthracyclines, cytoskeleton disruptors, epothilones, histone deacetylase inhibitors, topoisomerase I inhibitors, topoisomerase II inhibitors, kinase inhibitors, nucleotide analogs, peptide antibiotics, platinum-based drugs, retinoids and vinca alkaloids and derivatives. Alkylating agents can be bifunctional or monofunctional. Typical bifunctional alkylating agents include cyclophosphamide, nitrogen mustard, chlorambucil and melphalan. Typical monofunctional alkylating agents include dacarbazine, nitrosoureas and temozolomide. Typical anthracyclines include daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone and valrubicin. Typical cytoskeleton disruptors include paclitaxel, docetaxel, albumin-bound paclitaxel and taxotere. Typical histone deacetylase inhibitors include vorinostat and romidepsin. Typical topoisomerase I inhibitors include irinotecan and topotecan. Typical topoisomerase II inhibitors include etoposide, teniposide and tafluposide. Typical kinase inhibitors include bortezomib, erlotinib, gefitinib, imatinib, vemurafenib and vismodegib. Typical nucleotide analogs include azathioprine, capecitabine, cytarabine, doxifluridine, fluorouracil, gemcitabine, hydroxyurea, mercaptopurine, methotrexate and thioguanine. Typical retinoids include tretinoin, alitretinoin and bexarotene. Typical vinca alkaloids include vinblastine, vincristine and vindesine.

[0099] B. Preparation of Composite Resin Particles and Composite Particle Dispersions

[0100] The composite resin particles according to the present invention are a mixture of a biocompatible resin selected from polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof, polyamino acids and polyesters and at least one dye (preferably a dye having a maximum absorption at 580 nm to 750 nm) (see §A.1).

[0101] The resin according to the invention may be functionalized with functional groups which render the resin self-dispersible in water, with the proviso that the functionalization is limited to avoid that the resin becomes completely soluble in water. Typical functional groups which render the resin self-dispersible are selected from polyethylene oxide, carboxylic acids or salts thereof, sulfonic acids or salts thereof, phosphonic acids or salts thereof, phosphoric acid monoesters or salts thereof, sulfuric acid monoesters or salts thereof, ammonium groups, sulfonium groups and phosphonium groups.

[0102] The incorporation of polyethylene glycol functional groups is particularly useful for imparting stealth properties to the composite resin particles of the present invention in the human or animal body. These stealth properties are necessary to avoid rapid absorption by the reticuloendothelial system and to remain only at the desired site.

[0103] In a preferred embodiment, the resin according to the invention is not functionalized with functional groups which render the resin self-dispersible.

[0104] The use of a dispersant in the preparation of an aqueous dispersion of composite resin particles is particularly preferred. This allows the surface properties to be adjusted without the need to design a specialized resin. Standard biocompatible resins can be used in the preparation of resin particles. The dispersant can be a surfactant or a stabilizing polymer. The surfactant can be nonionic, anionic, cationic or zwitterionic. Nonionic ethoxylated block copolymer surfactants are preferred. Even more preferred are EO-PPO-EO triblock copolymers. The block copolymers impart stealth properties to the particles of the present invention, but also result in smaller particle diameters. As stabilizing polymers, hydroxyl-functionalized polymers are particularly preferred, preferably selected from polysaccharides and polyvinyl alcohol or polyvinyl alcohol copolymers or derivatives thereof. A particularly preferred dispersant is a cationic surfactant comprising at least one primary or secondary amine group and at least one quaternary ammonium group.

[0105] A particularly preferred method for preparing the composite resin particle dispersion according to the present invention comprises the following steps:

[0106] 1. dissolving the resin according to the present invention and a dye (preferably a dye having a maximum absorption at 580 nm to 750 nm) in a water-immiscible solvent; and

[0107] 2. Dissolving the dispersant in water; and

[0108] 3. emulsifying the solution of resin and dye into the aqueous solution of dispersant; and

[0109] 4. Evaporation of the water-immiscible solvent.

[0110] If necessary, the composite particles can be separated through a separation technique such as centrifugation, freeze drying, spray drying, . . . to obtain composite resin particles in a dry state.

[0111] If additional functional compounds such as pharmaceutically active agents have to be incorporated into the composite resin particles, the compounds are preferably dissolved in the water-immiscible solvent.The solvent evaporation technique according to the present invention is particularly interesting for the incorporation of active pharmaceutical ingredients such as anticancer drugs.

[0112] A particularly preferred method of preparing a dispersion of composite resin particles comprising an additional functional compound such as an active pharmaceutical ingredient according to the present invention comprises the following steps:

[0113] 1. dissolving the resin according to the present invention, a dye (preferably a dye having a maximum absorption at 580 nm to 750 nm) and a functional compound in a substantially water-immiscible solvent; and

[0114] 2. Dissolving the dispersant in water; and

[0115] 3. Emulsifying the solution of resin, dye and functional compound into the aqueous solution of dispersant; and

[0116] 4. Evaporating the substantially water-immiscible solvent.

[0117] In a more preferred embodiment of the present invention, the composite resin particles are capsules, wherein the core contains a dye, preferably a dye having a maximum absorption at 580nm to 750nm, and the shell is a resin selected from polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof. More preferably, the resin is a polyamino acid and is prepared using a ring-opening polymerization method, an interfacial ring-opening polymerization of N-carboxyl-anhydrides (such as those according to general structure IV).

[0118] Interfacial polymerization allows the preparation of capsules in a single step process with a wide range of functionalities and particle sizes, making it particularly suitable for industrial production processes, more particularly for continuous industrial processes. The technology can be easily tuned to the functionality to be encapsulated by simply adjusting the monomer ratios, and the physical properties can be easily adjusted for different applications without major changes in the process conditions, thus constituting a robust technology with considerable latitude for industrialization.

[0119] The ring-opening polymerization of N-carboxyl-anhydrides has been reviewed by Cheng and Deming (Top. Curr. Chem., 310, 1-26 (2012)). Primary amines and optionally secondary amines are the most obvious initiators and are widely used to initiate ring-opening polymerization via nucleophilic initiation. Basic initiators can initiate ring-opening polymerization via an activated monomer mechanism, first deprotonation of the NCA, followed by ring-opening polymerization. When amine initiators are used, these two mechanisms usually proceed simultaneously. Transition metal initiation is known to provide better control of polymerization. The use of hexamethyldisilazane as an initiator to better control polymerization has also been disclosed.

[0120] In a further preferred embodiment, use the mixture of N-carboxyl-acid anhydride derived from different amino acids. In an even further embodiment, use the mixture of different chiralities, preferably the mixture of 9 / 1 to 1 / 9 ratio of D-amino acid and L-amino acid. In another preferred embodiment, use chirality and the mixture of different amino acids. Mixing D-amino acid and L-amino acid will prevent polyamino acid from forming secondary or tertiary structure, as peptide in nature. Therefore, the polymer shell obtained will be more compact and mechanical tolerance is stronger. Such polymer shell will prolong the life-span of dye in human body or animal body.

[0121] In a particularly preferred interfacial ring-opening polymerization method for preparing capsules according to the invention, N-carboxyl-anhydride monomers and dyes (preferably dyes with maximum absorption at 580 nm to 750 nm) are dissolved in a substantially water-immiscible solvent and emulsified in an aqueous solution containing a polymerization initiator. After emulsification and optionally removal of the substantially water-immiscible solvent, ring-opening polymerization is initiated at the interface. After growth, a polyamino acid shell is formed at the organic-water interface, thereby generating a core-shell structure, which encapsulates the functional component or functional formulation. The polymer shell obtained is mechanically strong and stable and allows the capsule to be separated from the liquid in which it is prepared.

[0122] Suitable N-carboxy-anhydride monomers are described in §A.1. of unpublished European patent application number EP21163396.1 and in Table 1.

[0123] The capsules according to the invention are particularly suitable for encapsulating dyes. Capsules based on micelles are less suitable for encapsulating dyes. In fact, the shell of the micellar system is in many cases too permeable relative to the polymer shell obtained by the encapsulation method of the invention.

[0124] A particularly preferred interfacial ring-opening polymerization method comprises the following steps

[0125] a) dissolving a compound according to general structure IV and a dye (preferably a dye having an absorption maximum between 580 nm and 750 nm) in a water-immiscible solvent; and

[0126] b) dissolving a polymerization initiator in an aqueous liquid; and

[0127] c) emulsifying the solution obtained in step a) into the aqueous liquid; and

[0128] d) optionally evaporating the water-immiscible solvent; and

[0129] e) polymerizing a compound according to general structure IV.

[0130] The particle size of the capsules according to the invention can be varied by changing the emulsification technique, the use of an emulsifying aid and the ratio of the emulsifying aid to the shell and the core during the emulsification process, the nature of the emulsifying aid, the viscosity of the continuous phase or dispersed phase, the ratio of the continuous phase to the dispersed phase, the nature of the core content and the nature of the shell monomers. As emulsification techniques, high shear techniques and ultrasound-based techniques are particularly preferred. The particle size of the capsules according to the invention can be adjusted by adjusting the shear in the high shear technique or by changing the power and amplitude during the ultrasound treatment.

[0131] Difunctional or polyfunctional primary or secondary amines or mixtures thereof are particularly preferred initiators for the ring-opening polymerization of NCA. The initiators are water-soluble and can be functionalized with additional hydrophilic functional groups, which are preferably selected from carboxylic acids or salts thereof, sulfonic acids or salts thereof, phosphonic acids or salts thereof, phosphates or salts thereof, sulfates or salts thereof, polyhydroxyl functional groups, polyethylene glycols, ammonium groups, sulfonium groups and phosphonium groups.

[0132] If the capsules of the present invention are used as drug delivery systems in humans or animals, the incorporation of polyethylene glycol functional groups will be particularly useful to impart stealth properties thereto. These stealth properties are necessary to avoid absorption by the reticuloendothelial system and to release the drug only at the desired site in a controlled manner.

[0133] C. Application areas

[0134] The composite resin particles according to the invention are suitable for imaging affected organs in the human and / or animal body.The strong absorption of red or NIR light makes them also suitable for diffuse optical tomography and photoacoustic imaging.

[0135] When irradiated with appropriate red light or NIR laser, the dye in the composite particles of the present invention can convert the absorbed photon energy into heat, directly ablating cancer cells with minimal invasion of surrounding healthy tissues, making the particles very suitable for tumor phototherapy treatment (PTT).

[0136] The composite resin particles of the present invention can also be used in photodynamic therapy (PDT), where the NIR absorber is excited with light of an appropriate wavelength to convert molecular oxygen into cytotoxic reactive oxygen species (ROS), such as singlet oxygen, which in turn damage cancer cells through oxidative stress and thereby induce cell death.

[0137] D. Examples

[0138] D.1. Materials:

[0139] ●D,L-phenylalanine N-carboxy anhydride, D-phenylalanine N-carboxy anhydride and L-phenylalanine N-carboxy anhydride can be prepared according to standard methods such as disclosed by Dabashvill et al. (Journal of Physical Chemistry B, 111(38), 11105–11110 (2007)) and Otake et al. (Angewandte Chemie, International Edition, 57(35), 11389–11393 (2018)).

[0140] • L-Leucine N-carboxy anhydride, D-leucine N-carboxy anhydride and D,L-leucine N-carboxy anhydride can be prepared according to standard methods as disclosed by Baars et al. (Organic Process Research and Development, 7(4), 509-513 (2003)).

[0141] ●Crosslinker-1 is a trifunctional β-ketoester according to the following structure, which can be prepared as disclosed by Speisschaert et al. (Polymer, 172, 239-246 (2019)).

[0142]

[0143] CATSURF-1 is a cationic surfactant according to the following structure, which can be prepared as disclosed as Surf 3 in WO2018137993 (Agfa NV).

[0144]

[0145] ●SQ-1 is a dye with a maximum absorption in the range of 580-750 nm and is prepared as follows: 0.11 g of squaric acid is placed together with 5 mL of pyridine and 0.21 mL (0.23 g) of acetic anhydride. The mixture is stirred for one hour. 0.69 g of 1-butyl-2,3,3-trimethyl-3H-indolium iodide is added to the mixture and stirred at 60°C for 30 minutes. After cooling to room temperature, 20 mL of deionized water is added, the crude product is precipitated and filtered. The solid product is washed with water. The washed product is boiled in water, filtered and dried in vacuo at room temperature. 0.39 g (yield: 77%) of SQ-1 is isolated.

[0146] ●SQ-5 is a dye with a maximum absorption in the range of 580-750 nm and is prepared as follows: 0.23 g of squaric acid is placed together with 10 mL of pyridine and 0.41 mL (0.45 g) of acetic anhydride. The mixture is stirred for one hour. 1.43 g of 2,3,3-trimethyl-1-(3-methylbutyl)-3H-indolium iodide is added to the mixture and stirred at 60°C for 30 minutes. After cooling to room temperature, 15 mL of deionized water is added to the reaction mixture. The mixture is stirred overnight. The product crystallizes. The crystals are filtered, washed with water and dried in vacuo at room temperature. 0.72 g (yield: 67%) of SQ-5 is isolated.

[0147] ●SQ-6 is a dye with a maximum absorption in the range of 580-750 nm and is prepared as follows: 0.23 g of squaric acid is placed together with 10 mL of pyridine and 0.41 mL (0.45 g) of acetic anhydride. The mixture is stirred for one hour. 1.43 g of 3-butyl-1,1,2-trimethyl-1H-benzo[e]indolium bromide is added to the mixture and stirred at 110°C for 30 minutes. After cooling, 15 mL of MeOH and 25 mL of deionized water are added to the reaction mixture. The product partially crystallizes. Some of the non-crystalline product is decanted and recrystallized from acetone. The combined crystals are filtered, washed with water and dried in vacuo at room temperature. 0.62 g (yield: 51%) of SQ-6 is isolated.

[0148] AC-1 is a dye with an absorption maximum in the range of 580-750 nm and is prepared as follows: 3.54 g of the starting dye (I) is added to 25 mL of MeOH. 1.24 mL (0.98 g) of 2-ethylhexane-1-amine is added and stirred at 55° C. for 45 minutes. 0.44 mL (0.56 g) of 2-ethylhexane-1-amine is added and stirred at 55° C. for another 30 minutes. 0.32 mL (0.47 g) is added to the mixture and allowed to cool to room temperature. After adding 75 mL of deionized water, the mixture is stirred at room temperature overnight. The product crystallizes. The crystals are filtered, washed with water and dried under vacuum at room temperature. 3.33 g (yield: 99%) of AC-1 are isolated.

[0149]

[0150] ●DYE-1 is a dye with a maximum absorption in the range of 580-750 nm and is prepared as follows: 7.96 g of N-(2-hydroxyphenyl)-2-methoxy-acetamide is dissolved in 70.4 mL of MeOH and 10.1 g of N4-ethyl-N4-isopropyl-2-methyl-benzene-1,4-diamine and 18.7 g of Na2CO3 in 79 mL of deionized water and 17 mL of MeOH are added. A solution of 14.6 g of KI and I2 in 12 mL of deionized water is added dropwise to the mixture over 10 minutes and stirred at room temperature for 1 hour. The crude product is filtered and dissolved in 200 mL of deionized water (stirred for 15 minutes), filtered again, washed five times with 40 mL of deionized water / MeOH (v / v=9 / 1) and dried at 50°C. The obtained product (15.5 g of 15.9 g) was recrystallized in 100 mL MeOH / deionized water (v / v=95 / 5), washed with 80 mL MeOH / H2O (v / v=7 / 3) and dried at 50° C. 11.3 g (yield: 71.5%) of DYE-1 was isolated.

[0151] ●DYE-8 is a dye with a maximum absorption in the range of 580–750 nm and is prepared as follows: 0.50 g of the starting dye (I) is added to 20 mL of MeOH. 27 mg of 2-aminoethanethiol and 0.1 mL of triethylamine are added, stirred at 65° C. for 1 hour and allowed to cool. After adding a small amount of deionized water, the mixture is stirred overnight. The product crystallizes. The crystals are filtered and dried in vacuo at room temperature. 0.27 g (yield: 59%) of Dye-8 is isolated.

[0152] D.2. Methods

[0153] D.2.1. Particle size measurement

[0154] The particle size of the capsules was measured using a Zetasizer™ Nano-S (Malvern Instruments, Goffin Meyvis).

[0155] D.2.2. Measurement of light-to-heat conversion

[0156] The ability of capsules according to the invention to convert red laser light into heat was measured as follows: the prepared aqueous capsule dispersion was diluted with water to a final dye concentration of 69 μg / mL in 2 mL and exposed to a LPC836 laser diode (650 nm) at a power of 350 mW in a polystyrene cuvette.

[0157] According to the device schematically shown in FIG. 1 , the heat generated by the capsule was measured with a vertical infrared thermal sensor (RETTIM1x90614Esf) at a distance of 2 mm from the cuvette.

[0158] After 18 minutes of laser exposure, the temperature increase (ΔT) relative to room temperature was determined.

[0159] D.2.3. UV-VIS spectra of dyes and capsules

[0160] The dye was dissolved in methanol at a concentration of 3.2 μg / mL before measuring the UV-VIS spectrum of the dye having a maximum absorption at 580 to 750 nm.

[0161] The UV-VIS spectra of the capsules were measured at a dilution of 800 times compared to the prepared dispersion (see §D.3).

[0162] UV-VIS spectra were measured on an Agilent 8433 spectrophotometer up to 1100 nm.

[0163] D.2.4. Fluorescence spectrum measurement of capsules

[0164] The fluorescence spectra were measured with a UV-Vis-NIR spectroradiometer from Ocean Optics HR 4000.

[0165] D.3. Example 1

[0166] This example illustrates the ability of a capsule according to the present invention to convert red laser light into heat.

[0167] Preparation of capsule dispersions INVCAPS-1 to INVCAPS-6 of the present invention

[0168] 1.5 g of D,L-phenylalanine N-carboxy anhydride, 0.75 g of D-leucine N-carboxy anhydride, 0.75 g of L-leucine N-carboxy anhydride, 0.34 g of crosslinker-1 and 33 mg of dye were dissolved in 25 mL of ethyl acetate. The mixture was filtered through a 1.7 micron filter.

[0169] The types of dyes dissolved are listed in Table 5.

[0170] A second solution was prepared by dissolving 0.6 g of surfactant CATSURF-1 in 30 mL of water.

[0171] The first solution was added to the second solution and mixed with Ultra Turrax T25 at 15000 RPM for 5 minutes while maintaining the temperature at 20-30° C. The solvent was removed under reduced pressure to a total weight of 30 g. The polymerization was allowed to continue at room temperature for 24 hours.

[0172] The average particle size of the resulting capsule dispersion was measured according to D.2.1. and the values ​​are reported in Table 5. The absorption maximum of the dye was measured according to §D.2.3.

[0173] Table 5.

[0174]

[0175] Preparation of comparative capsule dispersion COMPCAPS

[0176] The comparative capsule dispersion was obtained in the same manner as the capsule dispersion according to the invention, but without the presence of dye (see Table 5)

[0177] Conversion of light into heat

[0178] The laser to heat conversion capabilities of the capsule dispersions of the invention and the comparative capsule dispersions were measured as described in D.2.2. The results are listed in Table 6.

[0179] Table 6.

[0180]

[0181] *The observed fluorescence is very low and is most likely due to impurities.

[0182] From the results of Table 6, it can be concluded that capsules containing a dye having maximum absorption at 580 nm to 750 nm and a polymer shell containing polyamino acid show the ability to convert red laser light into heat, resulting in a much higher temperature increase than the same capsules without dye.

[0183] The capsules of the present invention exhibit the ability to convert laser light into heat, making them useful in photothermal therapy (PTT), photodynamic therapy (PDT), and chemophotodynamic therapy.

[0184] It can also be concluded from the results of Table 6 that some of the capsules of the present invention as described above exhibit fluorescence, making them useful for fluorescence medical imaging.

Claims

1. A composite resin particle comprising a dye and a resin selected from the group consisting of polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof, characterized in that the composite resin particle has a maximum absorption in the range of 600 nm to 750 nm, the maximum absorption being measured for an aqueous dispersion of the composite resin particle having a concentration of 0.19 mg / mL, and characterized in that the average particle size of the particles is between 0.03 μm and 3 μm. 2 . The composite resin particles according to claim 1 , wherein the dye may be selected from azo dyes, phthalocyanines, anthraquinones, indigo dyes, cyanines, merocyanines, oxonoles, triarylmethane dyes, azomethine dyes, and indoaniline dyes. 3 . The composite resin particles according to claim 1 , wherein the ratio of the weight of the dye to the weight of the resin is from 1% by weight to 5% by weight.

4. The composite resin particles according to claim 2, wherein the cyanine is a squarylium dye according to Formula I in R1 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group. R2 and R3 are independently selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted aryl or heteroaryl groups, halogen, ester, amide, ether, nitro, nitrile, amine, ketone and aldehyde. R2 and R3 may represent the necessary atoms to form a five-membered or six-membered ring. X is selected from S, O, NR4 and CR5R6 R4, R5 and R6 are independently selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups and substituted or unsubstituted aryl or heteroaryl groups. R5 and R6 may represent atoms necessary to form a five-membered to eight-membered ring.

5. The composite resin particles according to claim 2, wherein the cyanine dye is a pentamethine cyanine dye according to Formula II in R1 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group. R2 and R3 are independently selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted aryl or heteroaryl groups, halogen, ester, amide, ether, nitro, nitrile, amine, ketone and aldehyde. R2 and R3 may represent the necessary atoms to form a five-membered or six-membered ring. R4 is selected from hydrogen, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group. X is selected from S, O, NR5 and CR6R7 R5, R6 and R7 are independently selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups and substituted or unsubstituted aryl or heteroaryl groups. R6 and R7 may represent atoms necessary to form a five- to eight-membered ring.

6. The composite resin particles according to claim 2, wherein the cyanine dye is a cyanine dye according to Formula III in R1 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group. R2 and R3 are independently selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted aryl or heteroaryl groups, halogen, ester, amide, ether, nitro, nitrile, amine, ketone and aldehyde. R2 and R3 may represent the necessary atoms to form a five-membered or six-membered ring. R4 is selected from a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group, a substituted or unsubstituted alkynyl group, a substituted or unsubstituted alkaryl group, a substituted or unsubstituted aralkyl group and a substituted or unsubstituted aryl or heteroaryl group. X is selected from S, O, NR5 and CR6R7 R5, R6 and R7 are independently selected from substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted alkaryl groups, substituted or unsubstituted aralkyl groups and substituted or unsubstituted aryl or heteroaryl groups. R6 and R7 may represent atoms necessary to form a five-membered to eight-membered ring.

7. Composite resin particles according to any one of the preceding claims, wherein the particles are capsules consisting of a polymer shell surrounding a core, the core comprising the dye, the polymer shell comprising a polyamino acid and being obtainable by interfacial polymerization of N-carboxy-anhydride monomers according to general structure IV. in n represents 0 or 1 R1, R2 and R3 are selected from hydrogen, substituted or unsubstituted alkyl groups, substituted or unsubstituted alkenyl groups, substituted or unsubstituted alkynyl groups, substituted or unsubstituted aralkyl groups, substituted or unsubstituted alkaryl groups and substituted or unsubstituted aryl or heteroaryl groups. R1, R2 and R3 may represent the necessary atoms to form a five-membered to eight-membered ring. The composite resin particles according to claim 7 , wherein the polymer shell contains a crosslinking agent.

9. The composite resin particle according to any one of the preceding claims, further comprising a pharmaceutically active compound.

10. The composite resin particles according to claim 9, wherein the pharmaceutically active compound is a cytostatic selected from alkylating agents, anthracyclines, cytoskeleton disruptors, epothilones, histone deacetylase inhibitors, topoisomerase I inhibitors, topoisomerase II inhibitors, kinase inhibitors, nucleotide analogs, peptide antibiotics, platinum-based drugs, retinoids and vinca alkaloids and their derivatives.

11. An aqueous dispersion comprising composite resin particles as defined in claims 1 to 10 and a stabilizing polymer or a surfactant.

12. The aqueous dispersion according to claim 11, wherein the surfactant is a cationic surfactant comprising at least one primary or secondary amine group and at least one quaternary ammonium group.

13. A pharmaceutical composition comprising a dispersion as defined in claims 11 to 12 and a pharmaceutical carrier or excipient.

14. A pharmaceutical composition comprising a dispersion as defined in claims 11 to 12 for use in the treatment of cancer.

15. A pharmaceutical composition comprising a dispersion as defined in claims 11 to 12 for use in a medical imaging method.

16. A method for preparing an aqueous dispersion as defined in claims 11 to 12, said method comprising the steps of: a) dissolving a resin selected from polyamino acids, polyphosphazenes, polysaccharide derivatives, polyesters, polyorthoesters, polycyanoacrylates and copolymers thereof and a dye having a maximum absorption between 580 and 750 nm in a substantially water-immiscible solvent; and b) dissolving the dispersant in water; and c) emulsifying the solution of the resin and the dye obtained in step a) into the aqueous solution of the dispersant obtained in step b); and d) evaporating the substantially water-immiscible solvent.

17. A method for preparing an aqueous dispersion as defined in claims 11 to 12, said method comprising the steps of: a) dissolving an N-carboxy-anhydride monomer according to general structure IV and a dye having an absorption maximum at 580 nm to 750 nm in a water-immiscible solvent; and b) dissolving a polymerization initiator in an aqueous liquid; and c) emulsifying the solution obtained in step a) into the aqueous liquid; and d) optionally evaporating the water-immiscible solvent; and e) polymerizing said N-carboxy-anhydride monomer according to general structure IV.

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