WO2010151074A2 - Conjugués contenant des photosensibilisateurs constitués de points quantiques et de dérivés de chlore et composition pour traiter et diagnostiquer un cancer contenant ces conjugués pour mettre en oeuvre une thérapie photodynamique - Google Patents

Conjugués contenant des photosensibilisateurs constitués de points quantiques et de dérivés de chlore et composition pour traiter et diagnostiquer un cancer contenant ces conjugués pour mettre en oeuvre une thérapie photodynamique Download PDF

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WO2010151074A2
WO2010151074A2 PCT/KR2010/004134 KR2010004134W WO2010151074A2 WO 2010151074 A2 WO2010151074 A2 WO 2010151074A2 KR 2010004134 W KR2010004134 W KR 2010004134W WO 2010151074 A2 WO2010151074 A2 WO 2010151074A2
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cancer
quantum dot
chlorine
conjugate
znte
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안웅식
배수미
바토그토크흐간트므르
문란영
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주식회사 진코스
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
    • A61K41/0057—Photodynamic therapy with a photosensitizer, i.e. agent able to produce reactive oxygen species upon exposure to light or radiation, e.g. UV or visible light; photocleavage of nucleic acids with an agent
    • A61K41/0071—PDT with porphyrins having exactly 20 ring atoms, i.e. based on the non-expanded tetrapyrrolic ring system, e.g. bacteriochlorin, chlorin-e6, or phthalocyanines
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/52—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an inorganic compound, e.g. an inorganic ion that is complexed with the active ingredient
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/545—Heterocyclic compounds
    • A61K47/546—Porphyrines; Porphyrine with an expanded ring system, e.g. texaphyrine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6921—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
    • A61K47/6923—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being an inorganic particle, e.g. ceramic particles, silica particles, ferrite or synsorb
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y5/00—Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery

Definitions

  • the present invention relates to a photosensitizer containing a conjugate of a quantum dot-chlorine derivative and a composition for treating and diagnosing cancer for use in photodynamic therapy comprising the same.
  • Photodynamic therapy is a medical treatment using a combination of light and photosensitizer (PS).
  • PS photosensitizer
  • the mechanism of action is largely dependent on the molecular mechanism of tumor-selective accumulation of photosensitive agents and the interaction of light with photosensitive agents. It can be divided into tumor destruction mechanism. Each factor is not harmful by itself, but when combined with oxygen, they can produce lethal cytotoxic agents that inactivate tumor cells [Sternberg ED et al., Tetrahedron , 1998, 54 : 4151-4202; Kadish KM et al., The Porphyrin Handbook. 2000, Vol 6 : 158-161.
  • PDT exhibits dual selectivity, in which the PS is preferentially absorbed by the diseased tissue, and the PS is activated by irradiating light in specific areas. PDT kills cells through the production of singlet oxygen and other reactive oxygen species (ROS), which overwhelms numerous antioxidant defense mechanisms in the cell and causes oxidative damage to the cell's macromolecules [Weisberger KR et al., Cancer Res , 1976, 36 : 2326-2329.
  • ROS reactive oxygen species
  • the photochemical reactions that generate ROS and singlet oxygen, which are putative cytotoxic agents formed during PDT, are represented by a modified Jablonsky diagram (FIG. 1).
  • the PS is electrically excited from the bottom singlet state (S 0 ) through the excited singlet state [S 1 , ( ⁇ 10 -6 s)] with short half-life [T 1. , ( ⁇ 10 -2 s)].
  • the excited singlet state PS which has a short half-life, can perform a non-radioactive process of intersecting systems (ISC).
  • the excited triplet state PS can perform two kinds of reactions [Macdonald JI et al., J Porphyrins Phthalocyanines , 2001, 5 : 105-129.]. First, it can participate in an electron-transfer process with a biological substrate to form radicals and radical ions that can produce superoxide ions, peroxide products such as O 2 ⁇ after interaction with oxygen [Type I reaction]. Alternatively, it can carry out a photochemical process known as a type II reaction in which stable triplet oxygen ( 3 O 2 ) is converted to singlet oxygen ( 1 O 2 ) with a short half-life but high reactivity.
  • the tumor cell killing effect of PDT is related to the depth of light penetration within the cancer mass.
  • the effect of light in tissues decreases exponentially with distance [Moser JG. In Photodynamic Tumor Therapy-2 nd & 3 rd Generation Photosensitizers . Harwood Academic Publishers, London, 1997: 3-8].
  • Tissue weakness is affected by optimal absorption, scattering by endogenous molecules and drug chromophores themselves.
  • the maximum transmittance of skin tissue is in the 700-800 nm region, and the development of a photosensitizer that exhibits maximum absorption within this region remains a major challenge. Effective penetration at 630 nm was between 1 and 3 mm, while light penetration of at least 6 mm was observed at 700-850 nm.
  • the ideal PS should exhibit strong absorption in the near infrared region.
  • PS is defined as a species that induces chemical or physical modification of other species under the absorption of light.
  • Clinicians and chemists have different views on the ideal PS [Kirchner C et al., Nano Lett, 2005, 5 , 331.].
  • the chemist can place more emphasis on the high degree of extinction and the high quantum yield of singlet oxygen, while the clinician can further emphasize low toxicity and high selectivity.
  • both clinical PDTs and ideal PSs are clinically appropriate and allison et al. [Zheng H. Technology in Cancer Research & Treatment , 2005, 4 : 283-293] and Castano et al. [Anna C et al., Photochem] Photobiol , 2006, 82 : 617-625] agree that at least some of the following criteria reported by:
  • tetrapyrrole macrocycles are often used as PS. Strong absorption in the red region of the visible spectrum is a very desirable feature for effective photosensitisers because it allows for the treatment of thicker tumors (Johnson CK et al., Tetrahedron Lett , 1998, 39 : 4619-4622). . For this reason, tetrapyrroles such as porphyrin, chlorine, bacteriochlorin, porphysin, phthalocyanine, naphthalocyanine, and expanded porphyrin have been synthesized and PDT efficacy has been evaluated. PS can be classified by their chemical structure and origin.
  • porphyrin-based eg photoprine, ALA / PpIX and BPD-MA
  • chlorine-based eg perpurin and bacteriochlorine
  • dyes eg phthalocyanine, naphthalocyanine
  • PDT drug The only widely used PDT drug at present is known to be “photoprine II”, which has the ability of porphyrin to accumulate tumors and its normally low toxicity, which has, for example, a series of disadvantages:
  • Photoprint (PF) (porpyrmer sodium) is a porphyrin oligomer complex and was first approved by a health care agency in 1993 for PDT of bladder cancer in Canada [Tian YY et al., Laser Phys. Lett, 2008, 5 , 764-767], several countries in Europe, the United States, and Asia, are currently approved for specific clinical applications and are being studied for use in other malignant and nonmalignant diseases [Menezes PFC et al. , Laser Phys , 2007, 17 , 461-467. Photoacid (PS) and photogram (PG) is the corresponding photosensitizer produced in Germany and Russia, respectively.
  • PS photoacid
  • PG photogram
  • Photogem is the most used photosensitizer in Brazil [Menezes PFC et al., Laser Phys , 2007, 17 , 461-467; Menezes PFC et al., Laser Phys, 2005, 15 , 435-442. These three HpDs are known to exhibit the same chemical and photophysical, diagnostic, and therapeutic characteristics as PF [Sokolov VV et al., Proc. SPIE , 1995, 2325 , 367-374; Mironov AF et al., J. Photochem. Photobiol, B, 1990, 4 , 297-306.
  • Quantum dots have emerged as an important class of materials that offer great potential for other types of applications ranging from energy conversion to biomedical science.
  • Quantum dots are usually fluorescent semiconductor nanocrystals with diameters on the order of 2-10 nanometers, or approximately 200-10,000 atoms [Daniele Gerion et al., J. Phys. Chem. B. 2001, 105, 8861-8871.
  • General structures of QDs include inorganic cores, inorganic shells, and aqueous organic coatings in which biomolecules are bound.
  • the size of the QDs provides them with unique optimal properties that can be adjusted from the UV region to the infrared region by changing their size, shape and composition [Burda C.
  • the present inventors have found that through the preparation of the quantum dot-chlorine derivative conjugates, the PDT efficiency of the photosensitizer can be significantly improved and the present invention has been completed.
  • quantum dots which are potential candidates for fluorescence imaging.
  • the present invention provides a photosensitizer containing a conjugate of quantum dot-chlorine derivatives.
  • the quantum dot is CdS, CdSe, CdTe, ZnS, ZnSe, ZnTe, HgS, HgSe, HgTe, AlP, AlAs, AlSb, GaP, GaAs, GaSb, InP, InAs, InSb, PbS, PbSe, PbTe, GeS , GeSe, GeTe, SnS, SnSe, SnTe, PbS, PbSe and PbTe.
  • the quantum dots include CdSe / ZnS, ZnTe / ZnSe, ZnSe / ZnTe, ZnTe / ZnO, ZnO / ZnTe, ZnSe / ZnO, ZnO / ZnSe, ZnS / ZnO, ZnO / ZnS, ZnTe / ZnS, ZnS / ZnS , InP / ZnTe and ZnTe / InP is characterized in that the quantum dot of the core / shell structure selected from the group consisting of.
  • the chlorine derivative is photodiazine, ladchlorin (Radachlorin), 2- (1-hexylethyl) -2- divinyl pyrophorovide- ⁇ (HPPH) [2- (1-hexylethyl) -2 It can be selected from -devinylpyropheophorbide- ⁇ (HPPH)], or mono -L- aspartyl chlorin e 6 (NPe 6) [mono -L-aspartylchlorin e 6 (NPe 6)].
  • the photosensitizers containing the conjugates of the quantum dot-chlorine derivatives are characterized by exhibiting photosensitizing activity against light in the range of 650 nm to 800 nm.
  • the present invention provides a method for preparing a conjugate of a quantum dot-chlorine derivative comprising the following steps:
  • the quantum dots are fluorescent nanoparticles.
  • the quantum dots most preferably include a CdSe core and a ZnS shell.
  • chlorine derivatives are compounds that can be activated by light in order to exhibit a photodynamic therapeutic effect.
  • the linker functional group is preferably ethylenediamine.
  • the chlorine derivative is extracted from Spirulina maxima algae.
  • the ligand is preferably 3-mercaptopropionic acid.
  • the hydrophilic organic amine is preferably N-methyl-D-glucosamine.
  • the method for preparing a chlorine derivative having a linker functional group comprises the following steps:
  • the methyl pheophoride-a is a chlorine derivative having the structure of formula (1).
  • R 1 and R 2 are CH 3 .
  • the mono-Boc-protected ethylenediamine is tert -butyl-N- (2-aminoethyl) carbamate having a structure of formula (2).
  • the Boc is a residue of di- tert -butyl-dicarbonate.
  • the protected chlorine derivative is a compound having the structure of formula (3).
  • R 1 and R 2 are CH 3 and R 3 is COO (CH 3 ) 3 .
  • the chlorine derivative from which the Boc functional group is removed is a compound having the structure of Formula 4 below.
  • R is H.
  • the method for preparing a quantum dot with a ligand attached comprises the following steps:
  • a method for preparing a quantum dot-chlorine derivative conjugate by conjugating a quantum dot with a ligand attached to a chlorine derivative having a linker functional group comprises the following steps:
  • the coupling agent is 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDCHCI) and N -hydroxysulfosuccinimide (Sulfo-NHS).
  • EDCHCI 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride
  • Sulfo-NHS N -hydroxysulfosuccinimide
  • the method for converting a covalently linked quantum dot-chlorine derivative conjugate to a water soluble conjugate by treating a hydrophilic organic amine comprises the following steps:
  • the present invention provides a composition for treating or diagnosing cancer for use in photodynamic therapy comprising a photosensitive agent containing a conjugate of a quantum dot-chlorine derivative as an active ingredient.
  • the photosensitive agent is characterized in that the light is activated in vitro or in vivo for the light beam in the range of 650 nm to 800 nm.
  • the cancer may be selected from the group consisting of skin, digestive, urinary, genital, respiratory, circulatory, brain and nervous system cancers.
  • the cancer is lung cancer, non-small cell lung cancer, colon cancer, bone cancer, pancreatic cancer, skin cancer, head or neck cancer, uterine cancer, ovarian cancer, rectal cancer, stomach cancer, anal muscle cancer, colon cancer, breast cancer, fallopian tube carcinoma, endometrial carcinoma Cervical carcinoma, vaginal carcinoma, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, endocrine gland cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute Among the group consisting of leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system (CNS) tumor, primary central nervous system lymphoma, spinal cord tumor, brain stem glioma and pituitary adenoma It may be selected, but is not necessarily limited thereto.
  • CNS central nervous system
  • the QD-photosensitizer conjugates of the invention are soluble in water and thus can be administered intravenously in the form of a solution.
  • composition for treating or diagnosing cancer of the present invention is intravenous injection, intraperitoneal injection, intramuscular injection, intracranial injection, intratumoral injection, intraepithelial injection, dermal penetration, esophageal administration, abdominal administration, arterial injection, intraarticular injection , And oral administration.
  • the present invention is a composition
  • a photosensitive agent containing a conjugate of a quantum dot-chlorine derivative as an active ingredient
  • kits for cancer treatment for use in photodynamic therapy comprising a light source for irradiating light with a wavelength in the range of 650 nm to 800 nm.
  • Chlorine is a compound having a structure of Formula 5, and is a large heterocyclic aromatic ring composed of pyrrole and pyrroline connected by four methine linkages at the center.
  • Magnesium-containing chlorine is called chlorophyll and is the central photosensitive pigment in the chloroplast.
  • the chlorine derivative means a compound having the chlorine as a basic skeleton structure. Chlorine and chlorine derivatives are effectively used as photosensitizers in photodynamic therapy because of their photosensitization.
  • Standards for optimal photosensitizers in PDT include strong absorption of light, low dark toxicity, high light toxicity, high quantum yield of singlet oxygen, high selectivity, fast excretion, ease of synthesis, pure composition , Excellent solubility in water, stability to photobleaching, and the like.
  • Chlorine and chlorine derivatives are selective for tumor tissue and in the absence of light, they are low in toxicity, rapidly excreted in the body and can be manufactured in large quantities from inexpensive raw materials.
  • one of the chlorine derivatives that may be used as a preferred embodiment may be photoditazine, but is not limited thereto.
  • Photodiazine is an N-methyl-D-glucosamine derivative of chlorine e 6 and has the structure of Formula 6 (FIG. 2).
  • the photodithazine of the present invention can be produced from the pheophorbide a based on chlorophyll-a derivatives by a known method, and commercially available products can also be used.
  • Photodiazine (PZ) exhibits strong absorption in the 650-680 nm range, which is soluble in water and more passes through biological tissue than at 630 nm where porphyrin is excited.
  • Photodiazine has the following bioactivity:
  • the chlorine derivative has a disadvantage in that it is difficult to penetrate into a tumor deeply located as a photosensitizer and thus does not cure completely and has a high possibility of recurrence.
  • Quantum dots are fluorescent semiconductor nanocrystals that have a unique glow and distribute them to many dye molecules, which store excitation energy and exist on the surface of quantum dots and produce singlet oxygen. In addition, they are effective photon collectors because of their broad absorption spectrum. Quantum dots have a narrow emission spectrum, and the photon absorption of the quantum dots can be controlled by the spectral transparent window of the human skin. Furthermore, quantum dots are carriers for drug delivery with high photostability.
  • Quantum dots are fluorescent nanocrystals approximately 2-10 nm in size and are biocompatible.
  • the quantum dots can be surface coated to make them water-soluble, biocompatible, target-specific, and functionalized.
  • a conjugate of the quantum dot and a chlorine derivative is provided. Since the conjugates of the quantum dot-chlorine derivatives of the present invention have minimal light scattering and absorption in the near infrared region of the spectrum by quantum dots, relatively low intensity light can be used to penetrate the tissue to a depth of several centimeters, More deeply located tumors may be accessed. In the conjugates of the quantum dot-chlorine derivatives of the present invention, due to the large transition dipole moment of the quantum dots, the quantum dots are strong absorbers, making them suitable agents for PDT applications.
  • CdSe QDs can be used to sensitize PDT agents by fluorescence resonance energy transfer (FRET) mechanisms.
  • FRET fluorescence resonance energy transfer
  • the production of singlet oxygen in CdSe QDs involves two steps. First, photoexcitation moves from the QD to the photosensitive agent. Secondly, the dye transfers energy to oxygen present in the enclosed solvent.
  • photoexcitation moves from the QD to the photosensitive agent.
  • the dye transfers energy to oxygen present in the enclosed solvent.
  • it can be used as an imaging, energy collector ( 1 O 2 ) and drug carrier through chemical coupling of a photosensitizer for PDT, a fluorescent (quantum dot) for enriched material for fluorescence diagnostics and MRI (FIG. 3).
  • conjugate of the quantum dot and the chlorine derivative of the present invention is excellent in water solubility and light stability, and has less side effects such as dark toxicity, exotherm, isomers, and impurities.
  • the term "inhibition of cell proliferation", “inhibition of cell growth” or similar form is used to suppress an increase in cell number, to any extent to any inhibition, for example, about 20% or more, about 50% or more, about That means containing over 90%, over 99% and complete inhibition ie 100% inhibition.
  • photodynamic therapy refers to a treatment by a combination of a photosensitizer (drug), light and oxygen.
  • a photosensitizer drug
  • oxygen oxygen
  • a chlorine derivative such as photoditagen having a linker functional group is prepared, and a ligand is attached to a quantum dot including a CdSe core and a ZnS shell, and then the chlorine derivative is attached to the quantum dot through the linker functional group and the ligand.
  • Conjugation synthesizes a conjugate of quantum dot-chlorine derivatives.
  • the linker functional group and the ligand should be selected to enable fluorescence resonance energy transfer (FRET) of the quantum dots after the quantum dots and the chlorine derivative are conjugated.
  • FRET fluorescence resonance energy transfer
  • ethylenediamine is used as the linker functional group and 3-mercaptopropionic acid is used as the ligand, but any one can be used as long as the two compounds can be conjugated to enable energy transfer of quantum dots.
  • the conjugate of the quantum dot and the chlorine derivative was found to have a markedly marked cell growth inhibitory effect compared to the chlorine derivative alone when irradiated with a laser.
  • CaSki cell line which is a cervical cancer cell line
  • MTT assay MTT assay
  • the method for preparing the conjugates of the quantum dot-chlorine derivatives of the present invention comprises the following steps:
  • the first is to modify chlorine compounds isolated from natural materials, and the second is to synthesize from pyrrole in a multi-step synthesis process. Any of these may be used, but in the present invention, a method of separating chlorophyll-a from natural materials and using it as a substrate is used to undergo further modification.
  • the starting material, methyl pheophorbid-a is obtained from Spirulina maxima algae. This process is briefly shown in Scheme 1 below.
  • the photosensitizer to be synthesized must have high purity, which is one of the characteristics required for the ideal photosensitizer, this is one of the important parts for producing a conjugate composed of the photosensitizer and quantum dots.
  • one of the main purposes is to synthesize a water-soluble photosensitizer. Therefore, the two methyl ester groups in the chlorine-e 6 derivatives are hydrolyzed under alkaline conditions and converted to carboxylic acid groups to make them water soluble. Such modifications must be performed before conjugation to the nanoparticles. After hydrolysis, the protecting group must be removed from the photosensitizer to bond to the nanoparticles. This process is briefly shown in Scheme 2 below.
  • Quantum dots must first react with 3-mercaptopropionic acid to functionalize the surface with carboxyl groups. Carboxylation of the quantum dots makes it possible to covalently bond the quantum dots to a photosensitiser having amino linker functionality. This process is briefly shown in Scheme 3 below.
  • the conjugation is carried out using the carbodiimide coupling technique.
  • This process involves a zero-length crosslinker, 1-ethyl-3- [3-dimethylaminopropyl] carbodiimide, in the presence of a hydrophilic activating functional group, N-hydroxysulfosuccinimide (Sulfo-NHS).
  • EDCHCI hydrochloride
  • the final step is to make the conjugate water soluble by complexing between N-methyl-D-glucosamine and two carboxyl groups in the conjugate. This process is briefly shown in Scheme 5 below.
  • the absorption maximum point of the QD-chlorine derivative conjugate was reduced to 656 nm as compared to the maximum point of the chlorine derivative alone (663.8 nm).
  • cancer cells can be diagnosed by utilizing the fluorescence of quantum dots in the QD-photosensitizer conjugate.
  • the present invention can be usefully used as a single agent for the treatment and diagnosis of cancer by chemically conjugating a photosensitizer for photodynamic therapy and a quantum dot, which is a phosphor for enhancing substances for fluorescence diagnosis and MRI.
  • 1 shows a modified Jablonsky diagram. Where 1 is absorption, 2 is non-radioactive decay, 3 is fluorescence, 4 is cross-over, 5 is phosphorescence, and 6 is energy transfer.
  • Figure 2 shows the structure of the photoditazine.
  • Figure 3 illustrates the fluorescence resonance energy transfer between quantum dots and chlorine derivatives.
  • FIG. 8 compares the fluorescence emission spectra of quantum dots (QD), carboxylated quantum dots (QDCOOH) and pre-bonded photosensitizers (PS).
  • QD quantum dots
  • QDCOOH carboxylated quantum dots
  • PS pre-bonded photosensitizers
  • FIG. 11 shows the results of cell growth inhibition after treatment of CaSki cell line, which is a cervical cancer cell line, with quantum dot-chlorine derivative conjugates and chlorine derivatives of 0.03125 and 0.0625 uM, respectively, without laser irradiation.
  • CaSki cell line which is a cervical cancer cell line with 0.03125 and 0.0625 uM of quantum dot-chlorine derivative conjugate and chlorine derivative, respectively.
  • the dried Spirulina maxima (Spirulina maxima) birds 500 g for 2 hours and refluxed to 2 L of acetone under nitrogen. The supernatant was then filtered over Whatman filter paper on a Buener funnel when hot and extra acetone was added to the remaining solid above. The extraction and filtration were repeated three times according to the same procedure. The green filtrate was evaporated and the residue was redissolved in 300 ml of acetone, cooled in a refrigerator and filtered to remove red solid impurities. The filtrate containing pheophytin-a was evaporated and treated with methanol solution of 5% sulfuric acid (500 ml) for 12.5 hours at room temperature under dark, nitrogen.
  • the solution was diluted with dichloromethane ( ⁇ 500 ml) and rinsed with water ( ⁇ 500 ml), rinsed with 10% aqueous sodium bicarbonate solution ( ⁇ 500 ml) and then rinsed three times with water.
  • the organic layer was separated, dried over anhydrous sodium sulfate and evaporated to dryness.
  • the residue was purified by column chromatography eluting with 2% acetone in dichloromethane on silica gel 60 (230-400 mesh). The product was recrystallized from dichloromethane / methanol.
  • Example 3 Preparation chlorin -e 6 -13 1 -N- (2- Boc-NH- ethyl) -amide -15 2 chlorine from -e 6 -13 1 -N- (2- amino-ethyl) -amide -15 2 (17 3 -dicarboxylic acid, PS, compound 4) synthesis
  • UV-vis (CH 3 OH + CH 2 Cl 2 ): ⁇ max , nm (Abs) 663.8 (0.71), 607.2 (0.22), 531.3 (0.23), 502.0 (0.30), 405.1 (2.08) and 330.2 (0.95).
  • MES buffer solution (2 ml, 0.1M, pH 5.0) is added to a mixture of carboxylated quantum dots (10 mg) dissolved in deionized water (1 ml) to which 1 M NaOH solution (3 drops) is added and the mixture was stirred at room temperature.
  • a solution of EDCHCI (10 mg) and sulfo-NHS (27.6 mg) in MES buffer solution (1 ml, 0.1M, pH 5.0) was added and the pH of the reaction mixture was adjusted to 1M NaOH solution (about 3 drops). Immediately adjusted to 6.3. The reaction mixture was stirred at rt for 30 min.
  • the remaining unreacted EDC was then quenched by the addition of 2-mercaptoethanol (3.5 ml, 14.3 mol / L) and the mixture was left for 10 minutes.
  • the pH of the reaction mixture was raised to 7.4 using concentrated PBS.
  • a solution of chlorine-e6 derivative in PBS buffer (2 ml, pH 7.4) was added to the solution containing the activated quantum dots and stirred slowly at room temperature for 2 hours.
  • the reaction was quenched for 10 minutes by adding glycine solution (1 ml, 14 mg) to hydrolyze the unreacted sulfo-NHS groups on the quantum dot surface.
  • the reaction solution was filtered through a hydrophilic cellulose acetate syringe filter (0.45 mm).
  • the filtered conjugate solution was purified by dialysis using a 12-14 kDa MWCO microcentrifuge tube with 4 buffer exchanges for 48 hours to remove byproducts and unreacted compounds.
  • the fluorescence spectra of the quantum dot alone, the photosensitizer alone and the conjugate were compared.
  • Figure 8 is a three-17 prepared in Example 3, when excited at 330 nm - a comparison of the fluorescence spectra of the dicarboxylic acid (PS), a quantum dot and a carboxylated quantum dots prepared in Example 4.
  • PS dicarboxylic acid
  • Quantum dots prepared in Example 4.
  • both quantum dots (611-612 nm) and photosensitizer (666 nm) exhibit effective strong emission in the 600-700 nm range.
  • the fluorescence emission spectrum of the quantum dot-photosensitizer conjugate showed two peaks overlapping each other at 611.6 nm and 640 nm when excited at 330 nm. Among these peaks the emission peak at 611.6 nm is indicative of the presence of quantum dots in the conjugate.
  • the emission peak at 640 nm indicates the presence of a photosensitizer in the conjugate, but the wavelength of this peak was reduced compared to the photosensitiser alone (666 nm, FIG. 8).
  • CaSki cell lines in DMEM (Dulbecco's modified eagle's medium) medium (Gibco, Rockville, MD, USA), 5% fetal bovine serum (Gibco, Rockville, MD, USA), 0.37% sodium bicarbonate, 20 mM HEPES and streptoto Mycin / penicillin (Gibco, Rockville, MD, USA) was used to add and incubated in 37 °C, 5% CO 2 incubator.
  • CaSki cell lines were dispensed into 3 wells in 96 well plates at 3 ⁇ 10 3 , and medium was changed after 12 hours after treatment with quantum dot-chlorine derivative conjugates and chlorine derivatives at concentrations of 0.03125 and 0.0625 uM, respectively. After medium replacement, the cell growth inhibition effect of the group irradiated with 662 ⁇ 3 nm laser and the group irradiated with 6.25J / Cm 2 was observed after 24 hours by MTT assay.
  • FIGS. 11 and 12 show the results for the group not irradiated with laser. As can be seen from FIG. 11, when the laser was not irradiated, that is, when there was no light, the chlorine derivative alone and the conjugate of the present invention did not show cell growth inhibitory effects. 12 shows the results for the group irradiated with the laser. As can be seen from Figure 12, compared with the chlorine derivative alone, the cell growth inhibitory effect was significantly superior in the conjugate of the present invention.

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Abstract

La présente invention concerne des conjugués contenant des photosensibilisateurs constitués de points quantiques et de dérivés de chlore et une composition pour traiter et diagnostiquer un cancer contenant ces conjugués pour mettre en oeuvre une thérapie photodynamique. Les conjugués de la présente invention peuvent être utilisés en tant que médicament unique pour traiter et diagnostiquer un cancer.
PCT/KR2010/004134 2009-06-26 2010-06-25 Conjugués contenant des photosensibilisateurs constitués de points quantiques et de dérivés de chlore et composition pour traiter et diagnostiquer un cancer contenant ces conjugués pour mettre en oeuvre une thérapie photodynamique WO2010151074A2 (fr)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2717920A2 (fr) * 2011-06-11 2014-04-16 University Of Central Florida Research Foundation, Inc. Nanosondes activables pour une administration intracellulaire de médicament
RU2638446C1 (ru) * 2016-12-14 2017-12-13 федеральное государственное автономное образовательное учреждение высшего образования "Национальный исследовательский ядерный университет МИФИ" (НИЯУ МИФИ) Способ направленного разрушения раковых клеток
CN107722075A (zh) * 2017-10-09 2018-02-23 大连理工大学 一类二氢卟吩葡萄糖苷类化合物及其制备方法与应用

Families Citing this family (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101668561B1 (ko) 2015-07-13 2016-10-21 원광대학교산학협력단 녹색형광단백질을 발현하는 유전자 및 로즈 벵갈을 이용한 암 질환의 광역학 치료용 조성물 및 이를 이용한 광역학 치료방법
US20170049891A1 (en) * 2015-08-17 2017-02-23 Nanoco Technologies Ltd. 5-aminolevulinic acid conjugated quantum dot nanoparticle
KR101911725B1 (ko) 2017-01-16 2018-12-28 주식회사 코어파마 적색형광단백질을 발현하는 유전자 및 틴 에틸 에티오푸르푸린을 이용한 광역학 치료용 조성물 및 이를 이용한 광역학 치료방법
KR102277244B1 (ko) * 2019-09-03 2021-07-13 인제대학교 산학협력단 생체 영상화 및 광열치료용 탄소양자점-폴리피롤 나노 복합체 및 이의 제조방법
KR102294374B1 (ko) 2019-09-09 2021-08-26 한국교통대학교산학협력단 전기화학적 무선 진단이 가능한 탄소 양자점의 제조방법, 및 이를 활용한 전기화학적 무선진단방법
KR102256817B1 (ko) 2019-09-09 2021-05-26 한국교통대학교산학협력단 자가 치유 전기화학적 무선진단이 가능한 탄소양자점 기반 수화젤의 제조방법 및 이를 활용한 자기 치유 전기화학적 무선진단방법
CN114933904B (zh) * 2022-06-14 2024-02-06 中国医科大学 一种用于光学诊疗的超薄壳层手性硒化镉/硫化镉材料及其制备方法与应用

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20080104495A (ko) * 2007-05-28 2008-12-03 전남대학교산학협력단 양자점을 이용한 생물학적 검출키트 및 방법

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20080104495A (ko) * 2007-05-28 2008-12-03 전남대학교산학협력단 양자점을 이용한 생물학적 검출키트 및 방법

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
BECHET, DENISE ET AL.: 'Nanoparticles as vehicles for delivery of photodynamic therapy agents' TRENDS IN BIOTECHNOLOGY vol. 26, no. 11, 2008, pages 612 - 621 *
CHO, SUNG JU ET AL.: 'Long-Term Exposure to CdTe Quantum Dots Causes Functional Impairments in Live Cells' LANGMUIR vol. 23, 2007, pages 1974 - 1980 *
SHAN, YAMING ET AL.: 'NHS-mediated QDs-peptide/protein conjugation and its application for cell labeling' TALANTA vol. 75, 2008, pages 1008 - 1014 *
TSAY, M. JAMES ET AL.: 'Singlet Oxygen Production by Peptide-Coated Quantum Dot- Photosensitizer Conjugates.' J. AM. CHEM. SOC. vol. 129, 2007, pages 6865 - 6871 *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2717920A2 (fr) * 2011-06-11 2014-04-16 University Of Central Florida Research Foundation, Inc. Nanosondes activables pour une administration intracellulaire de médicament
EP2717920A4 (fr) * 2011-06-11 2015-04-29 Univ Central Florida Res Found Nanosondes activables pour une administration intracellulaire de médicament
US9474809B2 (en) 2011-06-11 2016-10-25 University Of Central Florida Research Foundation, Inc. Activatable nanoprobes for intracellular drug delivery
RU2638446C1 (ru) * 2016-12-14 2017-12-13 федеральное государственное автономное образовательное учреждение высшего образования "Национальный исследовательский ядерный университет МИФИ" (НИЯУ МИФИ) Способ направленного разрушения раковых клеток
CN107722075A (zh) * 2017-10-09 2018-02-23 大连理工大学 一类二氢卟吩葡萄糖苷类化合物及其制备方法与应用

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