TW201718021A - 包含金屬奈米粒子、連接物及抗體之複合物 - Google Patents

包含金屬奈米粒子、連接物及抗體之複合物 Download PDF

Info

Publication number
TW201718021A
TW201718021A TW105123349A TW105123349A TW201718021A TW 201718021 A TW201718021 A TW 201718021A TW 105123349 A TW105123349 A TW 105123349A TW 105123349 A TW105123349 A TW 105123349A TW 201718021 A TW201718021 A TW 201718021A
Authority
TW
Taiwan
Prior art keywords
noble metal
metal nanoparticles
linker
antibody
complex
Prior art date
Application number
TW105123349A
Other languages
English (en)
Inventor
陳嘉南
陳嘉雄
趙月秀
吳奕宏
紀俐伶
Original Assignee
華上生技醫藥股份有限公司
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 華上生技醫藥股份有限公司 filed Critical 華上生技醫藥股份有限公司
Publication of TW201718021A publication Critical patent/TW201718021A/zh

Links

Classifications

    • 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
    • A61K47/62—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 a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/24—Heavy metals; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/24—Heavy metals; Compounds thereof
    • A61K33/242—Gold; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/24—Heavy metals; Compounds thereof
    • A61K33/243—Platinum; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/24—Heavy metals; Compounds thereof
    • A61K33/38—Silver; Compounds thereof
    • 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
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/20—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing sulfur, e.g. dimethyl sulfoxide [DMSO], docusate, sodium lauryl sulfate or aminosulfonic acids
    • 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
    • A61K47/56—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 macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/59—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 macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
    • A61K47/60—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 macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
    • 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
    • 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
    • A61K47/6927—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 a solid microparticle having no hollow or gas-filled cores
    • A61K47/6929—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 a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
    • A61K9/143—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with inorganic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C243/00—Compounds containing chains of nitrogen atoms singly-bound to each other, e.g. hydrazines, triazanes
    • C07C243/24—Hydrazines having nitrogen atoms of hydrazine groups acylated by carboxylic acids
    • C07C243/26—Hydrazines having nitrogen atoms of hydrazine groups acylated by carboxylic acids with acylating carboxyl groups bound to hydrogen atoms or to acyclic carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C271/00—Derivatives of carbamic acids, i.e. compounds containing any of the groups, the nitrogen atom not being part of nitro or nitroso groups
    • C07C271/06—Esters of carbamic acids
    • C07C271/08—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms
    • C07C271/10—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms
    • C07C271/22—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms to carbon atoms of hydrocarbon radicals substituted by carboxyl groups
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/50—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton
    • C07C323/51—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton
    • C07C323/52—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton the carbon skeleton being acyclic and saturated
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/50—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton
    • C07C323/51—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton
    • C07C323/60—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton with the carbon atom of at least one of the carboxyl groups bound to nitrogen atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D339/00—Heterocyclic compounds containing rings having two sulfur atoms as the only ring hetero atoms
    • C07D339/02—Five-membered rings
    • C07D339/04—Five-membered rings having the hetero atoms in positions 1 and 2, e.g. lipoic acid
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
    • C07K5/06026—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atom, i.e. Gly or Ala
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C07K5/1008—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2300/00—Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
    • A61K31/351—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom not condensed with another ring
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/437—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a five-membered ring having nitrogen as a ring hetero atom, e.g. indolizine, beta-carboline
    • 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
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/22—Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Epidemiology (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Inorganic Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Molecular Biology (AREA)
  • Genetics & Genomics (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Ceramic Engineering (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Immunology (AREA)
  • Nanotechnology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicinal Preparation (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Peptides Or Proteins (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

本發明係基於至少發現一種金屬奈米粒子，其可同時結合pH敏感性連接物、抗體及PEG以形成複合物。本發明之複合物可將該抗體傳遞至疾病過程中所涉及之靶細胞或接近細胞，以使該抗體可選擇性地靶向及作用於該等細胞。

Description

包含金屬奈米粒子、連接物及抗體之複合物
本發明係關於一種金屬奈米粒子、連接物及抗體或融合蛋白質之複合物。特定言之，本發明提供一種包含金屬奈米粒子、連接物、聚乙二醇(PEG)及抗體或融合蛋白質之複合物。
將治療及診斷化合物特定、有效地傳遞至細胞(尤其至細胞溶質)為許多醫藥公司之主要目標。已使用多種不同方法以提昇特定性及吸收。舉例而言，奈米技術已廣泛地用於藥物傳遞及癌症療法之新策略的研發中。已研發出pH敏感性奈米系統，其中具體藉由酸性腫瘤環境來觸發藥物釋放，且此等系統可改善癌症治療之功效。Feng Wang等人藉由經由酸不穩定鍵將小紅莓(doxorubicin)繫鏈於具有聚(乙二醇)位置之AuNP的表面上來揭示藥物傳遞系統(American Chemical Society,2011，第5卷，第5期，第3679-3692頁)。Tian-Meng Sun等人揭示使用經由腙鍵結合至金奈米粒子之小紅莓的癌症幹細胞療法(Biomaterials 35,2014，第836-845頁)。US 2013/0331764係關於用於將抗癌藥物傳遞至癌細胞之方法，其藉由使抗癌藥物與pH敏感性金屬奈米粒子結合，以使其與癌細胞分開。WO 2013/139942提供包含金屬奈米粒子及至少一種連接物之奈米粒子。
依那西普(Etanercept，Enbrel®)為TNF-α之有效抑制劑，已顯示其 在類風濕性關節炎(RA)及幼年型類風濕性關節炎(JRA)中提供快速及持久改善(Ann Rheum Dis 2000,59(增刊I)：146-149)。存在改善向靶細胞傳遞依那西普且由此提昇依那西普之治療作用的需要。依那西普為由重組DNA產生之融合蛋白質。其將腫瘤壞死因子(TNF)受體融合至IgG1抗體之定端。其由將兩種天然存在之可溶性人類75kDa TNF受體連接物與IgG1之Fc部分組合來製成。其為人工設計之二聚體融合蛋白質。
本發明提供一種包含貴金屬奈米粒子、複數個連接物結合物及複數個PEG之複合物，其中連接物具有下式：，其中m為1至4；其中連接物結合物包含連接物及抗TNFα人工融合蛋白質或抗TNFα抗體，其中融合蛋白質及抗體添加有醛基，且連接物之NH2基結合至融合蛋白質或抗體之醛基上；其中抗TNFα抗體為賽妥珠單抗(certolizumab)、英利昔單抗(infliximab)、利妥昔單抗(rituximab)、阿達木單抗(adalimumab)或托珠單抗(tocilizumab)或其生物類似物，且抗TNFα人工融合蛋白質為依那西普(etanercept)或其生物類似物；且其中連接物經由連接物中之1,2-二硫雜環戊烷基之硫原子連接至貴金屬奈米粒子，且PEG直接連接至貴金屬奈米粒子；且其中負載於貴金屬奈米粒子上之融合蛋白質或抗體的比率為約1.0%至約6.0%(w/w)，且負載於貴金屬奈米粒子上之PEG的比率為約0.5%(w/w)至約12%(w/w)。
在一個實施例中，貴金屬奈米粒子為Au奈米粒子、Ag奈米粒 子、Pd奈米粒子及Pt奈米粒子。較佳地，貴金屬奈米粒子為Au奈米粒子(AuNP)。在一個實施例中，貴金屬奈米粒子為Au奈米粒子、Ag奈米粒子、Pd奈米粒子及Pt奈米粒子。較佳地，貴金屬奈米粒子為Au奈米粒子。在一個實施例中，本發明中所使用之PEG的分子量在約1000至約20,000Da之範圍內。較佳地，PEG之分子量為約2000Da。在一個實施例中，連接物為連接物I-1(LK)。
在一個實施例中，負載於貴金屬奈米粒子上之抗TNFα融合粒子或抗體的比率為約1.0%至約5.0%(w/w)或約1%至約4%(w/w)。
在一些實施例中，對於尺寸為約20nm至約30nm之貴金屬奈米粒子，負載於貴金屬奈米粒子上之抗TNFα融合粒子或抗體的比率為約3.0%(w/w)至約6.0%(w/w)、約3.0%(w/w)至約5.0%(w/w)或約3.0%(w/w)至約4.0%(w/w)；更佳地，約3.5%(w/w)。對於尺寸為約40nm之貴金屬奈米粒子，負載於貴金屬奈米粒子上之抗TNFα抗體的比率為約2.0%(w/w)至約3%(w/w)；較佳地，約2.5%(w/w)至約3%(w/w)；更佳地，約2.63%(w/w)。
在一個實施例中，負載於貴金屬奈米粒子上之PEG的比率為約0.75%(w/w)至約10%(w/w)。
本發明亦提供一種醫藥組合物，其包含本發明之複合物及醫藥學上可接受之載劑。本發明亦提供用於治療自體免疫及免疫調節障礙疾病之方法，其包含向個體投與有效量之本發明的複合物。
圖1(A)至(F)顯示連接至不同粒度之AuNP的ETA/LK之飽和量。(A)及(B)占AuNP(20-30nm)之ETA/LK的飽和量為3.5%(w/w)。低於0.35μg之ETA/LK量導致AuNP(20-30nm)凝聚，呈現灰色-紫色；(C)及(D)占AuNP(40nm)之ETA/LK的飽和量在1.75%(w/w)至3.5%(w/w)之範圍內，且平均值為2.625%(w/w)；及(E)及(F)占AuNP(80 nm)之ETA/LK的飽和量在0.875%(w/w)至1.75%(w/w)之範圍內，且平均值為1.31%(w/w)。
圖2顯示LK成功地連接至ETA。OD536值自1.8391降低至背景。
圖3顯示AuNP/LK/ETA複合物(與ETA結合之20-30nm AuNP)之電漿子峰(λmax)中的2nm紅移。
圖4顯示在表面上結合有ETA融合蛋白質之各金奈米粒子。
圖5顯示AuNP/LK/ETA複合物之TEM圖像。觀察到1-5nm以金標記之二級抗人類IgG抗體與位於AuNP表面上之ETA結合。
圖6顯示AuNP/LK/ETA/PEG複合物之實例的示意圖。
圖7顯示AuNP/ETA/LK及AuNP/ETA/2KLK中之不同連接物長度不會影響ETA之活性。各自封端有或不含PEG之AuNP/ETA/LK及AuNP/ETA/2KLK不會影響ETA之活性。
本發明係基於至少一種金屬奈米粒子之發現，其可同時結合pH敏感性連接物、融合蛋白或抗體及PEG以形成複合物。本發明之複合物可將融合蛋白質或抗體傳遞至疾病過程中所涉及之靶細胞或接近細胞，以使融合蛋白質或抗體可選擇性地靶向及作用於細胞。
除非另外定義，否則本文所用之所有技術及科學術語均具有如一般技術者通常理解相同之含義。除非另外說明，否則本文中所提及之所有專利、申請案、公開之申請案及其他公開案均以全文引用之方式併入。除非另外說明，否則在本文術語存在複數個定義之情況下，以此部分中之定義為準。
從最廣之意義上使用術語「抗體」，且其包含單株抗體(例如全長或完好單株抗體)、多株抗體、單價、多價抗體、多特異性抗體(例如雙特異性抗體，只要其呈現所要之生物活性即可)，且亦可包括某些抗體片段(如本文中更詳細所描述)。抗體可為嵌合、人類、人類化及/ 或親和力成熟的。
如本文所使用，術語「腫瘤」係指所有瘤性細胞生長及增殖(無論惡性或良性)，及所有癌前及癌性細胞及組織。術語「癌症」、「癌性」、「細胞增生性病症」、「增生性病症」及「腫瘤」在本文中提及時不相互排斥。
如本文所使用，術語「癌症」係指哺乳動物中之生理病況，其典型特徵在於失調之細胞生長/增殖。癌症之實例包括(但不限於)癌瘤、淋巴瘤(例如霍奇金氏(Hodgkin's)及非霍奇金氏淋巴瘤(non-Hodgkin's lymphoma))、胚細胞瘤、肉瘤及白血病、鱗狀細胞癌、小細胞肺癌、非小細胞肺癌、肺腺癌、肺鱗狀癌、腹膜癌、肝細胞癌、腸胃癌、胰臟癌、神經膠母細胞瘤、子宮頸癌、卵巢癌、肝癌(liver cancer)、膀胱癌、肝癌(hepatoma)、乳癌、結腸癌、結腸直腸癌、子宮內膜或子宮癌、唾液腺癌、腎臟癌、肝癌(liver cancer)、前列腺癌、外陰癌、甲狀腺癌、肝癌(hepatic carcinoma)、白血病及其他淋巴組織增生病症及各種類型之頭頸癌。
如本文所使用，術語「治療(treating)」、「處理(treatment)」、「醫療(therapeutic)」或「療法(therapy)」未必意謂完全治癒或消除疾病或病況。疾病或病況之任何不期望之徵候或症狀達到任何程度上之任何緩解均可視為治療及/或療法。此外，治療可能包括可能使患者之總體健康或外觀惡化之作用。
「有效量」係指在所需劑量及時段下，可有效達成所需治療或預防結果之量。
本發明之物質/分子的「治療有效量」可根據諸如個體之疾病病況、年齡、性別、及重量、及物質/分子在個體中引起所需反應之能力等因素而變化。治療有效量亦為治療有益效應超過物質/分子之任何毒性或有害效應之量。「預防有效量」係指在所需劑量及時段下， 可有效達成所需預防結果之量。通常但並非必需，因為預防劑量係用在個體疾病之前或在疾病早期，所以預防有效量將低於治療有效量。
術語「醫藥學上可接受之鹽」係指對其所投與之生物體不產生顯著刺激且不消除化合物之生物活性及特性的化合物之鹽。
術語「生物類似物」(亦稱為後續生物製品或後入生物製品)係指近似為原始產物之相同複製物的生物醫學產物。
在一個態樣中，本發明提供包含貴金屬奈米粒子、複數個連接物結合物及複數個PEG之複合物，其中連接物具有下式：，其中m為1至4；其中連接物結合物包含連接物及抗TNFα人工融合蛋白質或抗體，其中融合蛋白質及抗體添加有醛基，且連接物之NH2基結合至融合蛋白質或抗體或生物類似物之醛基上；其中抗TNFα抗體為賽妥珠單抗(certolizumab)、英利昔單抗(infliximab)、利妥昔單抗(rituximab)、阿達木單抗(adalimumab)或托珠單抗(tocilizumab)或其生物類似物，且抗TNFα人工融合蛋白質為依那西普(etanercept)或其生物類似物；且其中連接物經由連接物中之1,2-二硫雜環戊烷基之硫原子連接至貴金屬奈米粒子，且PEG直接連接至貴金屬奈米粒子；且其中負載於貴金屬奈米粒子上之融合蛋白質或抗體的比率為約1.0%至約6.0%(w/w)，且負載於貴金屬奈米粒子上之PEG的比率為約0.5%(w/w)至約12%(w/w)。
在一個實施例中，貴金屬奈米粒子為Au奈米粒子、Ag奈米粒子、Pd奈米粒子及Pt奈米粒子。較佳地，貴金屬奈米粒子為Au奈米粒子。
在一個實施例中，貴金屬奈米粒子之尺寸在約20nm至約100nm之範圍內。在一些實施例中，尺寸為約20nm至約80nm、約20nm至約50nm、約20nm至約40nm或約20nm至約30nm。
在一個實施例中，連接物具有下式：
在一個實施例中，本發明中所使用之PEG的分子量在約1,000至約20,000Da之範圍內。較佳地，PEG之分子量為約1,000至約5,000Da或約2,000Da。
在一個實施例中，負載於貴金屬奈米粒子上之抗TNFα融合蛋白質或抗體的比率為約1.0%至約5%(w/w)、約1.0%至約4%(w/w)、約1%至約3.5%(w/w)、約1.3%至約3.5%(w/w)、約1.3%至約3%(w/w)或2.5%至3.5%。在一些實施例中，對於尺寸為約20nm至約30nm之貴金屬奈米粒子，負載於貴金屬奈米粒子上之抗TNFα融合蛋白質或抗體為約3.0%(w/w)至約6%(w/w)、約3.0%(w/w)至約5%(w/w)或約3.0%(w/w)至約4%(w/w)；更佳地，約3.5%(w/w)。對於尺寸為約40nm之貴金屬奈米粒子，負載於貴金屬奈米粒子上之抗TNFα融合蛋白質或抗體為約2.0%(w/w)至約3%(w/w)；較佳地，約2.5%(w/w)至約3%(w/w)；更佳地，約2.63%(w/w)。對於尺寸為約80nm之貴金屬奈米粒子，負載於貴金屬奈米粒子上之抗TNFα融合蛋白質或抗體為約1.0%(w/w)至約2%(w/w)；較佳地，約1.0%(w/w)至約1.5%(w/w)；更佳地，約1.31%(w/w)。
在一個實施例中，負載於貴金屬奈米粒子上之PEG的比率為約0.75%(w/w)至約10%(w/w)；較佳地，0.75%(w/w)至約9%(w/w)、0.75%(w/w)至約8%(w/w)、0.75%(w/w)至約7%(w/w)、0.75%(w/w) 至約6%(w/w)、0.75%(w/w)至約5%(w/w)、0.75%(w/w)至約4%(w/w)、約1%(w/w)至約10%(w/w)、約1%(w/w)至約9%(w/w)、約1%(w/w)至約8%(w/w)、約1%(w/w)至約7%(w/w)、約1%(w/w)至約6%(w/w)、約1%(w/w)至約5%(w/w)、約1%(w/w)至約4%(w/w)、約2%(w/w)至約6%(w/w)、約2%(w/w)至約10%(w/w)、約2%(w/w)至約8%(w/w)、約2%(w/w)至約5%(w/w)或約2%(w/w)至約4%(w/w)。
如本文所描述，抗TNFα抗體或融合蛋白質經氧化以具有醛基。可藉由首先在連接物末尾之NH2基處使連接物與抗TNFα融合蛋白質或抗體結合，且將連接物結合物及PEG連接至金屬奈米粒子以形成複合物來製備本發明之複合物。連接物經由連接物中之1,2-二硫雜環戊烷基之硫原子連接至貴金屬奈米粒子，且PEG直接連接至貴金屬奈米粒子。抗TNFα融合蛋白質或抗體以飽和狀態負載於貴金屬奈米粒子上。用於連接物與抗TNFα融合蛋白質或抗體之結合及連接物結合物及PEG與金屬奈米粒子之連接的方法為此項技術中已知的。
如本文所使用，抗TNFα抗體為賽妥珠單抗、英利昔單抗、利妥昔單抗、阿達木單抗或托珠單抗或其生物類似物。抗TNFα融合蛋白質為依那西普或其生物類似物。
鑒於本發明，可使用熟習此項技術者已知之方法製備本發明之連接物I-醯胺連接物。舉例而言，可如以下方案中顯示製備本發明之較佳連接物。
在一些情況下，亦可使用聚乙二醇化以減小連接物或奈米粒子及生物部分之間的電荷相互作用，例如藉由在連接物或奈米粒子之表面上產生可避免聚合物與生物部分相互作用之親水層。在一些情況下，添加聚(乙二醇)重複單元可延長聚合性結合物之電漿半衰期，例如藉由減少由吞噬細胞系統導致之聚合性結合物的吸收同時減少由細胞導致之轉染/吸收功效。本領域普通技術人員將知曉用於聚乙二醇化聚合物之方法及技術，舉例而言，如下文實例中所論述，藉由開環聚合技術(ROMP)或類似者，藉由使用EDC(1-乙基-3-(3-二甲胺基丙基)氫氯化碳化二亞胺)及NHS(N-羥基丁二醯亞胺)以使聚合物與以胺收端之PEG基反應。
在一個態樣中，本發明提供一種醫藥組合物，其包含本發明之複合物及醫藥學上可接受之載劑。
在另一態樣中，本發明提供用於治療自體免疫及免疫調節障礙疾病之方法，其包含向個體投與有效量之本發明的複合物。在一些實施例中，疾病為發炎性腸病、牛皮癬、化膿性汗腺炎及頑固性哮喘類風濕性關節炎、幼年型類風濕性關節炎及牛皮癬性關節炎、斑狀牛皮癬或僵直性脊椎炎。
本文所描述之一些實施例係關於一種組合物，其可包含連接本 發明之抗TNFα融合蛋白質或抗體的金屬奈米粒子複合物及醫藥學上可接受之載劑。醫藥組合物有助於向生物體投與連接抗TNFα融合蛋白質或抗體之金屬奈米粒子複合物。用於經口、經直腸、局部或非經腸(包括皮下、腹膜內、肌內及靜脈內)投與之合適醫藥學上可接受之載劑為熟習此項技術者所已知的。載劑必須在與調配物之其他成分相容且不對其受體有害之意義上為醫藥學上可接受的。存在於此項技術中之投與化合物的多種技術包括(但不限於)經口、注射、噴霧、非經腸及局部投與。
舉例而言，適於非經腸投與之組合物宜包括活性化合物之無菌水性製劑，其較佳與接受者之血液呈等張性。適用調配物亦包括含有本發明之複合物的濃縮溶液或固體，其一旦用適當溶劑稀釋，即產生適於上述非經腸投與之溶液。
舉例而言，對於經口投與，本發明之複合物可併入至惰性載劑中，呈離散單位形式，諸如膠囊、扁膠劑、錠劑或口含劑，各自含有預定量之抗TNFα抗體；呈粉末或顆粒狀；或呈懸浮液或水性液態溶液或非水性液體狀，例如糖漿、酏劑、乳液或飲劑。合適載劑可為澱粉或糖且包括崩解劑、潤滑劑、調味劑、黏合劑及其他相同性質之物質。
舉例而言，經直腸投與之調配物可使用習知載劑，例如可可豆油或Witepsol S55(Dynamite Nobel Chemical,Germany之商標)作為栓劑基質，形成栓劑。舉例而言，本文所描述之某些實施例係關於一種藥物傳遞系統，其包含連接一或多種相同或不同治療劑或診斷劑之一或多種金屬奈米粒子複合物。
組合物可宜呈單位劑型，且可藉由藥學技術中熟知之任何方法來製備。
任何組合物可視情況包括一或多種其他組分，諸如鎮痛劑(諸如 NSAID)、COX-2抑制劑及改善疾病之抗風濕藥(DMARD)。DMARD之實例包括(但不限於)甲胺喋呤(methotrexate)、羥基氯喹(hydroxychloroquine)、柳氮磺胺吡啶(sulfasalazine)、來氟米特(1eflunomide)、TNFα抑制劑(賽妥珠單抗、英利昔單抗及依那西普)、利妥昔單抗、托珠單抗、阿巴西普(abatacept)及阿那白滯素(anakinra)。NSAID之實例包括(但不限於)阿司匹林(aspirin)、二氟尼柳(diflunisal)、雙水楊酸酯(salsalate)、布洛芬(ibuprofen)、右布洛芬(dexibuprofen)、非諾洛芬(fenoprofen)、酮基布洛芬(ketoprofen)、右酮洛芬(dexketoprofen)、氟比洛芬(flurbiprofen)、奧沙普嗪(oxaprozin)、洛索洛芬(Loxoprofen)萘普生(naproxen)。COX-2抑制劑之實例包括(但不限於)塞內昔布(celecoxib)。
投與連接抗TNFα融合蛋白質或抗體之金屬奈米粒子複合物的多種技術存在於此項技術中，其包括(但不限於)經口、經直腸、局部、噴霧、注射及非經腸傳遞，包括肌內、皮下、靜脈內、髓內注射、鞘內、直接室內、腹膜內、鼻內及眼內注射。
實例 實例1 製備連接物I-1
步驟1 將乙醇胺(1.28g，1.05當量)及類脂酸(4.13g，1.0當量)溶解於80mL DCM中，且緩慢添加EDC．HCl(4.98g，1.3當量)、羥基丁二醯亞胺(NHS)(2.30g，1.0當量)及三乙胺(6.4mL，2.3當量)。在室溫下進行反應達至少5小時且反應之後為TLC。隨後，藉由添加ddH2O來淬滅混合物，且用DCM萃取2-3次。收集有機層，用無水硫酸鎂乾燥，用MgSO4過濾且隨後移除DCM。藉由使用DCM：MeOH=9.8：0.2用管柱來純化萃取物。產物為淺黃色液體。產量：4.04g；81.0%。
步驟2 將氯甲酸4-硝基苯酯(3.35g，1.15當量)置於雙頸瓶中。在真空下達1小時，將三通閥轉向至氮氣器件且用注射器添加60mL DCM。用10-15mL DCM溶解經稱重之步驟1產物1，用注射器將其注入至反應燒瓶中，且隨後緩慢添加三乙胺(4.5mL，2.3當量)。在冰浴中進行反應約1小時且隨後回到室溫下持續整夜。反應完成後，藉由添加ddH2O來淬滅混合物，且用DCM萃取3次。收集有機層，用無水硫酸鎂乾燥，用MgSO4過濾且隨後移除DCM。藉由使用EtOAc：Hex：MeOH=1：1：0.2用管柱來純化萃取物。產物為淺黃色液體。純化後產率：51.6%。
步驟3 將經稱重之步驟2產物2(3.5g，1.0當量)溶解於120mL DCM中，且隨後緩慢添加水合肼(6.5mL，10.0當量)。在室溫下進行反應約24小時。溶液顏色自淺黃色變為橙色。反應完成後，藉由添加ddH2O來淬滅混合物，且用DCM萃取2-3次。收集有機層，用無水硫酸鎂乾燥，用MgSO4過濾且隨後移除DCM。藉由使用DCM：MeOH=9.5：0.5用管柱來純化萃取物。在此步驟中，產物阻塞於管柱中且可提高MeOH至5%來洗滌最終產物。產物為淺黃色黏性固體。純化後產率：19.5%。
連接物I-1(連接物I-醯胺，m=2)。 1 H(400MHz)δ 1.23-1.31(2H,m),1.40-1.51(3H,m),1.57-1.64(1H,m),1.78-1.86(1H,m),2.05(2H,t),2.32-2.39(1H,m),3.01-3.18(4H,m),3.39(2H,t),3.52-3.57(3H,m), 4.03-4.19(4H,m),7.94(1H,s),8.14(1H,s)。 13 C(100MHz)δ 25.9,29.0,34.8,36.1,38.9,39.5,40.9,57.2,64.7,69.3,69.6,159.5,174.9ppm。H-質量(m/z)：(M+Na)=374.1171
連接物I-2(連接物I-醯胺，m=1)。 1 H(300MHz)δ 1.30-1.38(2H,m),1.45-1.57(3H,m),1.60-1.72(1H,m),1.80-1.92(1H,m),2.06(2H,s),2.36-2.46(1H,m),3.07-3.24(4H,m),3.56-3.65(1H,m),3.95(2H,t),4.02(2H,s),7.88(1H,t),8.11(1H,s)。 13 C(100MHz)δ 25.6,28.8,34.6,35.9,38.8,39.4,40.7,56.9,63.7,163.8,174.3ppm。H-質量(m/z)：(M+Na)=330.0894。
連接物I-3(連接物I-醯胺，m=3)。 1 H(300MHz)δ 1.30-1.37(2H, m),1.45-1.57(3H,m),1.60-1.72(1H,m),1.80-1.91(1H,m),2.06(2H,t),2.35-2.46(1H,m),3.07-3.23(4H,m),3.39(2H,t),3.47-3.65(7H,m),4.02(2H,s),4.05-4.08(2H,m),7.84(1H,t),8.18(1H,s)。 13 C(100MHz)δ 25.7,28.9,34.7,35.9,38.8,39.4,40.7,60.0,64.4,69.4,69.6,70.1,70.3,159.2,174.2ppm。H-質量(m/z)：(M+Na)=418.1466。
連接物I-4(連接物I-醯胺，m=4)。 1 H(300MHz)δ 1.30-1.38(2H,m),1.45-1.57(3H,m),1.60-1.72(1H,m),1.80-1.91(1H,m),2.06(2H,t),2.36-2.46(1H,m),3.07-3.23(4H,m),3.39(2H,t),3.49-3.65(11H,m),4.02(2H,s),4.05-4.08(2H,m),7.83(1H,t),8.18(1H,s)。 13 C(100MHz)δ 25.7,28.9,34.7,35.9,38.8,39.4,40.7,56.9,64.4,69.4,69.6,70.2,70.3,70.4,159.2,174.1ppm。H-質量(m/z)：(M+Na)=440.1875。
實例2 製備本發明之複合物
使用10kDa MWCO離心式過濾器(Millipore,UFC501024)濃縮依那西普(Etanercept，ETA)(Mycenax biotech,TuNEX)，且以1mg/mL將其溶解於100mM Na2HPO4(pH值為7.4之緩衝劑)中。隨後，將20μL含有100mM NaIO4之水溶液添加至100μL依那西普溶液中，且在暗處 保溫混合物達30分鐘。藉由添加250μL 1×PBS來淬滅反應物。此時，氧化依那西普Fc部分上之碳水化合物部分以使其具有醛基。隨後，將LK(連接物I-1)添加至依那西普溶液中。LK在分子相對位置上具有醯肼及二硫醇基。醯肼部分與改質依那西普分子之Fc部分的醛基相互作用。在室溫下保溫反應混合物達2小時，且隨後使用離心式過濾器收集硫醇化依那西普，且在0.1M磷酸鈉緩衝劑(pH值為7.4)中對其進行再懸浮。將硫醇化依那西普與0.5mL金奈米粒子混合，且在室溫下保溫懸浮液達1小時以獲得AuNP/LK/ETA複合物。
與AuNP連接之ETA/LK的飽和度及藥物負載量。在溶液中，將以不同量(0μg、0.0438μg、0.0875μg、0.175μg、0.35μg、0.7μg及1.4μg)存在於0.1M磷酸鹽緩衝劑中之ETA/LK與10μg AuNP(20-30nm)混合，且隨後在室溫下保溫達60分鐘以用於飽和度測試。使用UV-Vis分光光度計量測與AuNP連接之ETA/LK的飽和度。使用AuNP之顏色測定與AuNP連接之ETA/LK的飽和度。
將14μg ETA/LK添加至1mL之50ppm AuNP(其具有不同尺寸)中，且隨後混合。在室溫下保溫所得混合物達60分鐘。隨後，以15000rpm離心所得混合物達20分鐘。對所得上清液進行BCA分析以量測藥物負載量。根據下式計算藥物負載量：
結果顯示占AuNP(20-30nm)之ETA/LK的飽和量為3.5%(w/w)。低於0.35μg之ETA/LK量導致AuNP凝聚，呈現灰色-紫色(參見圖1(A)及(B))。根據上文所提及之量及步驟，在飽和度測試中使用尺寸為40nm之AuNP。結果顯示占AuNP(40nm)之ETA/LK的飽和量在1.75%(w/w)至3.5%(w/w)之範圍內，且平均值為2.625%(w/w)(參見圖1(C)及(D))。類似地，根據上文所提及之量及步驟，在飽和度測試中使用 尺寸為80nm之AuNP。結果顯示占AuNP(80nm)之ETA/LK的飽和量在0.875%(w/w)至1.75%(w/w)之範圍內，且平均值為1.31%(w/w)(參見圖1(E)及(F))。上文所提及之結果概括於下表中。
ETA連接至LK(連接物I-1)之證明。
用UV-Vis分光光度計(Beckman,DU 800)分析LK與ETA之醛基的連接。在室溫下，將ETA之醛基與10mg/mL purpald溶液混合達2小時。醛基之吸收係在536nm處。LK連接至ETA之醛基以成為ETA/LK。當在室溫下將ETA/LK與10mg/mL purpald溶液混合達2小時時，無法在536nm處量測到吸收值。圖2顯示LK成功地連接至ETA。
ETA/LK連接至AuNP之證明。用UV-Vis分光光度計(Beckman,DU 800)分析AuNP/LK/ETA複合物。所記錄之表面電漿子共振譜顯示AuNP/LK/ETA複合物之電漿子峰(λmax)中的2nm紅移(圖3)。使用螢光顯微鏡(Nikon,TE2000-U)量測以AuNP/LK/ETA複合物標記之Alex Fluor 568二級抗體的紅色螢光圖像。各金奈米粒子顯示紅色螢光，其表明各金奈米粒子在表面上與ETA抗體結合(圖4)。吾等藉由與結合有1-5nm AuNP之二級抗體結合來進一步檢測AuNP/LK/ETA複合物。在AuNP/LK/ETA複合物之TEM圖像中，觀察到1-5nm以金標記之二級抗人類IgG抗體結合至位於AuNP表面上之ETA(圖5)。
連接至AuNP/LK/ETA複合物之AuNP的PEG。將具有2000Da分子量之PEG添加至AuNP/LK/ETA/PEG複合物溶液中。將所得混合物置放於室溫下達20分鐘，且隨後以15000rpm離心20分鐘以移除溶液。將所得沈澱物添加至水中以調配所需濃度之AuNP/LK/ETA/PEG 複合物溶液。AuNP/LK/ETA/PEG複合物之實例的示意圖顯示於圖6中。將AuNP/LK/ETA/PEG複合物保持在4℃來儲存以供進一步使用。所得AuNP/LK/ETA/PEG複合物具有以下特徵，其列於下表中。
實例3 AuNP尺寸、連接物長度及封端有或不含PEG對阻斷TNFα治療之影響
AuNP尺寸。使用尺寸為20-90nm之AuNP連接至ETA/LK以測試AuNP尺寸對阻斷TNF-α治療之影響。用以下混合物治療MCF-7細胞(5x105)：(A)僅50ng/ml TNF-α，(B)30μg/mL尺寸為20-50nm之AuNP，(C)50ng/ml TNF-α及30μg/mL尺寸為20-50nm之AuNP，(D)50ng/ml TNF-α及4.2μg/mL AuNP(20-50nm)/LK/ETA複合物，(E)50ng/ml TNF-α及與AuNP(20-50nm)/LK/ETA複合物之ETA相等的ETA。在5%CO2下在37℃下培養經治療之細胞達48小時。結果顯示將尺寸為20-50nm之AuNP連接至ETA/LK不會影響ETA之活性。類似地，亦使用尺寸為70-90nm之AuNP且顯示相同結果，亦即將尺寸為20-50nm之AuNP連接至ETA/LK不會降低ETA之活性。
連接物長度及封端有或不含PEG。第一，使用AuNP連接至ETA/LK或ETA/2KLK(2000Da連接物)，且測試所得AuNP/ETA/LK及AuNP/ETA/2KLK以評估連接物長度對阻斷TNF-α治療之影響。第二，使用AuNP/LK/ETA複合物及AuNP/LK/ETA/PEG複合物測試封端有或不含PEG對阻斷TNF-α治療之影響。用2.1μg/mL AuNP/LK/ETA複合物、2.1μg/mL AuNP/LK/ETA/PEG複合物、2.1μg/mL AuNP/2KLK/ETA複合物及2.1μg/mL AuNP/2KLK/ETA/PEG複合物之混合物治療MCF-7細胞。將上文所提及之複合物、30μg/mL尺寸為20-50nm之AuNP及50ng/ml TNF-α添加至6孔培養盤(各孔接種有5X105MCF-7細胞)中，且在5%CO2下在37℃下培養48小時。結果顯示AuNP/ETA/LK及AuNP/ETA/2KLK中之不同連接物長度不會影響ETA之活性，以及各自封端有或不含PEG之AuNP/ETA/LK及AuNP/ETA/2KLK不會影響ETA之活性(參見圖7)。
AuNP/LK/ETA/PEG複合物之例示性生物活性
將500μL、0.025ng/μL TNF-α及500μL、1ng/μL AuNP/LK/ETA/PEG複合物或游離ETA混合達2小時。將所得混合物添加至6孔板(各孔接種有5x105MCF-7細胞)中以分別在5% CO2下在37℃下進行培養。兩天後，對細胞進行統計。比較AuNP/LK/ETA/PEG複合物及游離ETA之生物活性，發現AuNP/LK/ETA/PEG複合物中和TNF-α之阻斷能力與游離ETA之阻斷能力相同，因此AuNP、LK及PEG之結合不會影響 ETA活性。游離ETA及在5%CO2下在37℃下之複合物的相對細胞存活率(%)顯示於下表中。

Claims (15)

  1. 一種複合物其包含貴金屬奈米粒子、視數個連接物結合物及複數個PEG，其中該連接物具有下式：，其中m為1至4；其中連接物結合物包含連接物及抗TNFα人工融合蛋白質或抗體，其中該融合蛋白質及該抗體已添加醛基，且該連接物之NH2基結合至該融合蛋白質或該抗體或生物類似物之醛基上；其中該抗TNFα抗體為賽妥珠單抗(certolizumab)、英利昔單抗(infliximab)、利妥昔單抗(rituximab)、阿達木單抗(adalimumab)或托珠單抗(tocilizumab)或其生物類似物，且該抗TNFα人工融合蛋白質為依那西普(etanercept)或其生物類似物；且其中該連接物經由該連接物中之1,2-二硫雜環戊烷基之硫原子連接至該貴金屬奈米粒子，且該等PEG直接連接至該貴金屬奈米粒子；且其中負載於該貴金屬奈米粒子上之該融合蛋白質或抗體的比率為約1.0%至約6.0%(w/w)，且負載於該貴金屬奈米粒子上之該PEG的比率為約0.5%(w/w)至約12%(w/w)。
  2. 如請求項1之複合物，其中該貴金屬奈米粒子為Au奈米粒子、Ag奈米粒子、Pd奈米粒子及Pt奈米粒子。
  3. 如請求項1之複合物，其中該貴金屬奈米粒子為Au奈米粒子。
  4. 如請求項1之複合物，其中該貴金屬奈米粒子之尺寸在約20nm至約100nm之範圍內。
  5. 如請求項1之複合物，其中該連接物具有下式：
  6. 如請求項1之複合物，其中用於本發明中之PEG的分子量在約1,000至約20,000Da之範圍內。
  7. 如請求項1之複合物，其中負載於該貴金屬奈米粒子上之該抗TNFα融合蛋白質或抗體的比率為約1%至約4%(w/w)。
  8. 如請求項1之複合物，其中對於尺寸為約20nm至約30nm之貴金屬奈米粒子，負載於該貴金屬奈米粒子上之該抗TNFα抗體或融合蛋白質為約3.0%(w/w)至約6.0%(w/w)；對於尺寸為約40nm之貴金屬奈米粒子，負載於該貴金屬奈米粒子上之該抗TNFα抗體為約2.0%(w/w)至約5%(w/w)；或對於尺寸為約80nm之貴金屬奈米粒子，負載於該貴金屬奈米粒子上之該抗TNFα抗體為約1.0%(w/w)至約3%(w/w)。
  9. 如請求項1之複合物，其中負載於該貴金屬奈米粒子上之該PEG的比率為約0.75%(w/w)至約10%(w/w)。
  10. 如請求項1之複合物，其中負載於該貴金屬奈米粒子上之該PEG的比率為約2%(w/w)至約5%(w/w)。
  11. 一種醫藥組合物，其包含如請求項1之複合物及醫藥學上可接受之載劑。
  12. 如請求項11之醫藥組合物，其可進一步包含一或多種其他組分。
  13. 如請求項14之醫藥組合物，其中該其他組分為鎮痛劑(NSAID)、COX-2抑制劑或改善疾病之抗風濕藥(DMARD)。
  14. 一種用於治療自體免疫及免疫調節障礙疾病之方法，其包含向個體投與有效量之本發明的複合物。
  15. 如請求項19之方法，其中該疾病為發炎性腸病、牛皮癬、化膿 性汗腺炎及頑固性哮喘類風濕性關節炎、幼年型類風濕性關節炎及牛皮癬性關節炎、斑狀牛皮癬或僵直性脊椎炎。
TW105123349A 2015-07-22 2016-07-22 包含金屬奈米粒子、連接物及抗體之複合物 TW201718021A (zh)

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
US201562195654P 2015-07-22 2015-07-22

Publications (1)

Publication Number Publication Date
TW201718021A true TW201718021A (zh) 2017-06-01

Family

ID=57833804

Family Applications (2)

Application Number Title Priority Date Filing Date
TW105123126A TW201713365A (zh) 2015-07-22 2016-07-21 遞送治療藥物之ｐｈ－敏感性的連接物
TW105123349A TW201718021A (zh) 2015-07-22 2016-07-22 包含金屬奈米粒子、連接物及抗體之複合物

Family Applications Before (1)

Application Number Title Priority Date Filing Date
TW105123126A TW201713365A (zh) 2015-07-22 2016-07-21 遞送治療藥物之ｐｈ－敏感性的連接物

Country Status (10)

Country Link
US (1) US10688193B2 (zh)
EP (1) EP3325440B1 (zh)
JP (1) JP6875371B2 (zh)
KR (1) KR102187410B1 (zh)
CN (2) CN107848957B (zh)
AU (1) AU2016295602B2 (zh)
ES (1) ES2797923T3 (zh)
HK (1) HK1246773A1 (zh)
TW (2) TW201713365A (zh)
WO (2) WO2017012568A1 (zh)

Families Citing this family (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU2016295602B2 (en) 2015-07-22 2019-11-14 Gnt Biotech & Medicals Corporation PH-sensitive linkers for delivering a therapeutic agent
CN106831853B (zh) * 2017-02-15 2019-02-22 浙江海正药业股份有限公司 7-乙基-10-o-叔丁基二苯基硅基喜树碱-20-o-甘氨酸盐酸盐的制备工艺
GB201819430D0 (en) 2018-11-29 2019-01-16 Midatech Ltd Therapeutic compounds, nanoparticles and uses thereof
RS64992B1 (sr) 2018-12-17 2024-01-31 Gi Cell Inc Blok kopolimer koji sadrži hidrofilni prvi blok, hidrofobni drugi blok i funkcionalnu grupu sposobnu da se specifično vezuje za tiol
KR20200087335A (ko) 2018-12-31 2020-07-21 강원대학교산학협력단 합산곱신경망을 이용한 영상 처리 방법 및 장치
KR20230009407A (ko) * 2020-06-12 2023-01-17 애피컬 몰레큘러 바이오테크 씨오., 엘티디. 비알코올성 지방간 질환 및 비알코올성 지방간염 치료용 카보네이트를 함유한 프테로스틸벤 아미노산의 신규 유사체
CZ309422B6 (cs) * 2020-09-29 2022-12-28 Ústav organické chemie a biochemie AV ČR, v. v. i. Povrchově modifikované částice
CN114377126A (zh) * 2021-12-16 2022-04-22 天津市泌尿外科研究所 纳米钯片负载妥珠单抗的纳米复合体、制备方法及其在制备治疗炎症相关性贫血药物的应用
GB202305212D0 (en) * 2023-04-06 2023-05-24 Elixa Mediscience Ltd Compounds

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110077581A1 (en) * 2009-09-25 2011-03-31 Georgia Tech Research Corporation Targeted cellular delivery of nanoparticles
KR101196667B1 (ko) * 2010-04-15 2012-11-02 포항공과대학교 산학협력단 피에이치 민감성 금속 나노 입자를 이용한 항암제 전달 시스템
US8697129B2 (en) * 2011-03-02 2014-04-15 Imra America, Inc. Stable colloidal gold nanoparticles with controllable surface modification and functionalization
US8717746B2 (en) 2012-03-22 2014-05-06 Abb Technology Ag Cooling apparatus for switchgear with enhanced busbar joint cooling
AU2016295602B2 (en) 2015-07-22 2019-11-14 Gnt Biotech & Medicals Corporation PH-sensitive linkers for delivering a therapeutic agent

Also Published As

Publication number Publication date
TW201713365A (zh) 2017-04-16
CN108348599A (zh) 2018-07-31
KR20180033533A (ko) 2018-04-03
US20180221498A1 (en) 2018-08-09
US10688193B2 (en) 2020-06-23
AU2016295602A1 (en) 2018-02-15
JP2018524389A (ja) 2018-08-30
HK1246773A1 (zh) 2018-09-14
CN107848957B (zh) 2021-05-11
WO2017012591A1 (en) 2017-01-26
CN107848957A (zh) 2018-03-27
EP3325440A4 (en) 2019-01-23
AU2016295602B2 (en) 2019-11-14
KR102187410B1 (ko) 2020-12-07
JP6875371B2 (ja) 2021-05-26
WO2017012568A1 (en) 2017-01-26
EP3325440B1 (en) 2020-04-08
EP3325440A1 (en) 2018-05-30
ES2797923T3 (es) 2020-12-04

Similar Documents

Publication Publication Date Title
TW201718021A (zh) 包含金屬奈米粒子、連接物及抗體之複合物
WO2023078273A1 (en) Specific conjugation for an antibody-drug conjugate
TWI804499B (zh) Ror1抗體免疫接合物
JP2021104030A (ja) 抗ｅｇｆｒ抗体及び抗体薬物コンジュゲート
US10576161B2 (en) Compositions and methods for the depletion of CD137+ cells
TW200539855A (en) Calicheamicin conjugates
JP7449583B2 (ja) 標的特異的複合体及びその用途
US20210283267A1 (en) Therapeutic methods using antibody drug conjugates (adcs)
CN117482261A (zh) 放射性免疫缀合物与检查点抑制剂组合疗法
CN110049999A (zh) 抗psma抗体、其用途及其缀合物
TWI726217B (zh) 含有抗globo h抗體之抗體－藥物共軛物及其用途
US20240058467A1 (en) Anti-ror1 antibody conjugates, compositions comprising anti ror1 antibody conjugates, and methods of making and using anti-ror1 antibody conjugates
JP2019534706A (ja) 抗ｅｄｂ抗体および抗体−薬物コンジュゲート
JP2022515885A (ja) 切断可能なリンカーを含む化合物及びその使用
US20220118104A1 (en) Compounds comprising cleavable linker and uses thereof
TWI807320B (zh) 含有α-烯醇酶抗體之藥物共軛物及其用途
WO2022068775A1 (zh) 抗pd-l1抗体及其用途
JP2023506158A (ja) Ｃｄ２７６に特異的な抗体－薬物コンジュゲートおよびその使用
CN114173822A (zh) 用胶原结合药物载体治疗癌症的方法和组合物
US20210095039A1 (en) Anti-cd252 antibodies, conjugates, and methods of use
WO2023190827A1 (ja) ｐＨ依存性抗硫酸化グリコサミノグリカン抗体及び抗体薬物複合体
CN118922210A (zh) B7h4抗体药物偶联物及其用途