WO2011002077A1 - Agent de thérapie génique tnf-α contenu dans une micelle polymère - Google Patents

Agent de thérapie génique tnf-α contenu dans une micelle polymère Download PDF

Info

Publication number
WO2011002077A1
WO2011002077A1 PCT/JP2010/061312 JP2010061312W WO2011002077A1 WO 2011002077 A1 WO2011002077 A1 WO 2011002077A1 JP 2010061312 W JP2010061312 W JP 2010061312W WO 2011002077 A1 WO2011002077 A1 WO 2011002077A1
Authority
WO
WIPO (PCT)
Prior art keywords
group
integer
pharmaceutical composition
cancer
general formula
Prior art date
Application number
PCT/JP2010/061312
Other languages
English (en)
Japanese (ja)
Other versions
WO2011002077A8 (fr
Inventor
賢二 中野
一則 片岡
伸宏 西山
Original Assignee
国立大学法人 東京大学
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 国立大学法人 東京大学 filed Critical 国立大学法人 東京大学
Publication of WO2011002077A1 publication Critical patent/WO2011002077A1/fr
Publication of WO2011002077A8 publication Critical patent/WO2011002077A8/fr

Links

Images

Classifications

    • 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
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • A61K48/0008—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition
    • A61K48/0025—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition wherein the non-active part clearly interacts with the delivered nucleic acid
    • A61K48/0041—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition wherein the non-active part clearly interacts with the delivered nucleic acid the non-active part being polymeric
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • A61K48/005—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'active' part of the composition delivered, i.e. the nucleic acid delivered
    • 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
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia

Definitions

  • the present invention relates to a TNF- ⁇ gene therapeutic agent, and more particularly to the field of cancer treatment using a TNF- ⁇ expression plasmid.
  • Pancreatic cancer which is said to be a representative of intractable cancer, is difficult to detect abnormalities because the pancreas is in the back of the body, and has the property of growing invasively and prone to metastasis early as a characteristic of pancreatic cancer, and anticancer drugs are also ineffective Therefore, there is currently no satisfactory effective treatment.
  • peritoneal dissemination that has metastasized from stomach cancer, colon cancer, pancreatic cancer, biliary tract cancer, etc. is difficult to detect at an early stage.
  • the establishment of an effective treatment for such intractable cancer is eagerly desired, but the possibility that gene therapy born from a new concept is effective has not yet been fully examined.
  • the present invention has been made in view of the above problems, and the problem to be solved is a gene delivery system that is effective in the treatment of intractable cancer including pancreatic cancer and peritoneal dissemination, and has higher safety. And so on.
  • the present inventors have found that a cultured pancreatic cancer cell resistant to cell growth inhibition using a recombinant TNF- ⁇ protein has a high TNF- ⁇ expression plasmid. It has been found that the introduction of molecular micelles has an unexpected effect on cell proliferation. The present inventors have further studied and succeeded in suppressing the growth of pancreatic cancer and peritoneal dissemination by delivering the polymer micelle to the cancer tissue in the living body, and have completed the present invention.
  • a pharmaceutical composition comprising a TNF- ⁇ expression plasmid as an active ingredient and encapsulating the expression plasmid in a polymer micelle comprising the following block copolymer: (1) a block copolymer having a hydrophilic segment made of polyethylene glycol or a derivative thereof and a polycationic segment made of a polypeptide, or (2) a hydrophilic segment made of polyethylene glycol or a derivative thereof and a polypeptide A block copolymer in which the hydrophilic segment and the polycationic segment are bonded via a disulfide bond.
  • R 1 represents a hydrogen atom or an optionally substituted linear or branched C 1-12 alkyl group
  • L 1 and L 2 represent a linking group
  • R 2 represents a methylene group or an ethylene group
  • R 3 represents a hydrogen atom, a protecting group, a hydrophobic group or a polymerizable group
  • R 4 is the same as R 5 or is an initiator residue
  • R 5 each independently represents a hydroxyl group, an oxybenzyl group, or a —NH— (CH 2 ) a —X group, wherein X each independently represents a bulky amine compound residue having a pKa value of 7.4 or less.
  • the TNF- ⁇ gene may be any animal-derived gene.
  • the human TNF- ⁇ gene is published in Genbank Accession No. NM_000594.2 and can be isolated by a method known per se.
  • the mouse TNF- ⁇ gene is published in Genbank Accession No. NM_013693.2 and can be isolated by a method known per se.
  • TNF- ⁇ gene or “DNA encoding TNF- ⁇ ” refers to a human TNF- ⁇ gene (DNA) represented by a specific base sequence (SEQ ID NO: 1), a homologue of TNF- ⁇ , It is used for the purpose of including genes (DNA) encoding mutants and matured bodies. Specifically, the human TNF- ⁇ gene described in SEQ ID NO: 1 and mouse homologs and rat homologs thereof are included. The gene or DNA does not ask whether the functional region is different, and can include, for example, an expression control region, a coding region, an exon, or an intron.
  • Such variants include naturally occurring allelic variants, non-naturally occurring variants, and variants having amino acid sequences that have been altered by artificial deletions, substitutions, additions and insertions.
  • Examples of the mutant include those that are at least 70%, preferably 80%, more preferably 95%, and still more preferably 97% homologous to a protein or (poly) peptide without mutation.
  • the derivatives include those in which the amino terminal or carboxy terminal or side chain of the protein is substituted.
  • the matured body includes a protein obtained by cleaving the N-terminal signal peptide, and examples thereof include a polypeptide comprising an amino acid sequence at positions 77 to 233 of the amino acid sequence set forth in SEQ ID NO: 2.
  • the amino acid modifications include naturally occurring amino acid modifications and non-naturally occurring amino acid modifications, and specifically include post-translational modifications of amino acids.
  • RGD peptide for example, ACDC RGD CFCG (SEQ ID NO: 3; also referred to as RGD4C), DGARYC RGD CFDG (SEQ ID NO: 4; also referred to as RGD10), C RGD CF (K [H-] KKK) 6 (SEQ ID NO: 5; also referred to as cRGD-hK), However, it is not limited to these.
  • Block Copolymer The block copolymer used in the present invention has a block in which a hydrophilic segment composed of polyethylene glycol (PEG) or a derivative thereof and a polycationic segment composed of a polypeptide are bound via a disulfide bond.
  • a copolymer (1) and a block copolymer (2) bonded via a disulfide bond are included.
  • Derivatives of PEG are shown in the following formulas (I) to (V).
  • the hydrophilic segment and the polycationic segment the structure (for example, the degree of polymerization) is not limited, and an arbitrary structure can be selected.
  • the polycation is a polycation having a cationic group in the side chain.
  • a peptide is preferred.
  • a hydrogen atom or a protecting group and m is from 5 to 20,000.
  • N is an integer from 2 to 5,000
  • y is an integer from 0 to 4,999
  • z is an integer from 1 to 4,999
  • z is less than n
  • y + z is n Suppose it is not larger.
  • R 1 represents an hydrogen atom or a substituted C 1-12 good straight or branched even if alkyl, C 1- Examples of 12 alkyl include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, n-hexyl, decyl, undecyl, dodecyl and the like.
  • R 1 is preferably methyl or acetalized formyl group, cyano group, formyl group, carboxyl group, amino group, C 1-6 alkoxycarbonyl group, C 2-7 acylamide group, the same or different tri-C 1-6 A linear or branched C 1-12 alkyl group substituted with an alkylsiloxy group, a siloxy group or a silylamino group.
  • the substituent is an acetalized formyl group, it can be converted to another formyl group (—CHO; aldehyde group) by hydrolysis under acidic mild conditions.
  • Such formyl group, or carboxyl group or amino group can be present, for example, in the shell part of the polymer micelle of the copolymer and nucleic acid according to the present invention, and through these groups, the antibody or its specific Fragments having binding properties (F (ab ′) 2 , F (ab), etc.) and proteins that can impart other functionalities or targeting properties to the micelles can be used to covalently bind to the micelles.
  • a PEG segment having a functional group at one end can be conveniently formed by, for example, a method for producing a PEG segment portion of a block copolymer described in WO96 / 32434, WO96 / 33233, and WO97 / 06202.
  • the PEG or derivative portion thus formed and the polypeptide portion have any linkage mode depending on the method for producing the copolymer of the general formula (I), (II), (III) or (IV). As long as it meets the object of the present invention, it may be bonded with any linking group.
  • Examples of the linear or branched C 1-20 alkyl group include C 1-6 lower alkyl such as methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, n-pentyl, n-hexyl, and the like.
  • C 7-12 intermediate alkyl such as heptyl, octyl, nonyl, decyl, undecyl, dodecyl and the like
  • C 13-20 higher alkyl such as tridecyl, tetradecyl, hexadecyl, octadecyl, icosyl and the like.
  • These groups may optionally be substituted with one or more halogens (eg, fluorine, base, bromine), and may be substituted with one hydroxyl group in middle to higher alkyls. Good.
  • R 13 represents a hydroxyl group, an oxybenzyl group, or an NH—R 20 group, wherein R 20 represents a linear or branched C 1-20 alkyl group which may be substituted, R 14 and R 15 each independently represents a methylene group or an ethylene group, R 16 represents a hydrogen atom, a protecting group, a hydrophobic group or a polymerizable group, R 17 and R 18 each independently represents a hydroxyl group, an oxybenzyl group, or an NH— (CH 2 ) a —X group, wherein a is an integer of 1 to 5, and X is independently Represents an amine compound residue containing at least one group derived from a primary, secondary, or tertiary amine compound, or a quaternary ammonium salt, or a non-amine compound residue, and among the total number of R 17 and R 18 , ⁇ There are at least two groups that are NH— (CH 2 ) a —X groups (where
  • mice Six-week-old female nude mice were purchased from CLEA Japan. All mice were bred with free intake of sterilized feed and sterilized water. All animal studies were conducted in accordance with the principles of the University of Tokyo guidelines on animal experiments.
  • the rTNF group (middle) to which recombinant hTNF was added at a concentration of 20 ng / mL compared to the control group (right) into which the Mock gene was introduced in the PANC-1 and Suite-2 cells, showed cell growth. Mildly suppressed. In the hTNF gene-introduced group (left), more cytotoxicity was observed than in the rTNF group.
  • the TNF- ⁇ concentration in the medium measured by ELISA was 8.4 ng / mL for PANC-1 cells and 15.9 ng / mL for Suite-2 cells, which was lower than that of recombinant hTNF 20 ng / mL. Nevertheless, the cytotoxic effect was high. From this result, it was suggested that the cell growth of pancreatic cancer can be further suppressed by continuously expressing human TNF- ⁇ in pancreatic cancer cells.
  • Example 9 Effect of RGD-hTNF gene therapy in colon cancer-derived peritoneal dissemination 1 ⁇ 10 6 HCT116-Luc colon cancer cells that stably express luciferase were transplanted together with Matrigel into the abdominal cavity of nude mice, and a peritoneal dissemination model was established. Produced. One week after transplantation, the hTNF expression plasmid-encapsulated PEG-SS-P [Asp (DET)] polymer micelle and RGD-hTNF expression plasmid-encapsulated PEG-SS-P [Asp (DET)] produced in Example 3 were high.

Abstract

L'invention porte sur un système de distribution de gène ou similaire, qui est utile pour le traitement de cancers réfractaires comprenant le cancer du pancréas et une dissémination péritonéale. De façon spécifique, l'invention porte sur une composition pharmaceutique contenue dans une micelle polymère, ladite composition pharmaceutique contenant un plasmide d'expression pour TNF-a en tant qu'ingrédient actif. Le plasmide d'expression est composé de l'un des polymères à blocs suivants : (1) un copolymère à blocs qui a un segment hydrophile qui est composé d'un polyéthylène glycol ou d'un dérivé de celui-ci et un segment polycationique qui est composé d'un polypeptide ; ou (2) un copolymère à blocs qui a un segment hydrophile qui est composé d'un polyéthylène glycol ou d'un dérivé de celui-ci et un segment polycationique qui est composé d'un polypeptide, le segment hydrophile et le segment cationique étant liés par une liaison disulfure.
PCT/JP2010/061312 2009-07-02 2010-07-02 Agent de thérapie génique tnf-α contenu dans une micelle polymère WO2011002077A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2009-158282 2009-07-02
JP2009158282 2009-07-02

Publications (2)

Publication Number Publication Date
WO2011002077A1 true WO2011002077A1 (fr) 2011-01-06
WO2011002077A8 WO2011002077A8 (fr) 2011-04-07

Family

ID=43411145

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2010/061312 WO2011002077A1 (fr) 2009-07-02 2010-07-02 Agent de thérapie génique tnf-α contenu dans une micelle polymère

Country Status (1)

Country Link
WO (1) WO2011002077A1 (fr)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004099287A1 (fr) * 2003-05-08 2004-11-18 Japan Science And Technology Agency Copolymere sequence polyethyleneglycol/polycation
WO2006085664A1 (fr) * 2005-02-10 2006-08-17 The University Of Tokyo Polymere chargeable de polycations et utilisation comme vecteur d'acides nucleiques
WO2007099660A1 (fr) * 2006-03-01 2007-09-07 The University Of Tokyo complexe de micelles polymeres contenant de l'acide nucleique

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004099287A1 (fr) * 2003-05-08 2004-11-18 Japan Science And Technology Agency Copolymere sequence polyethyleneglycol/polycation
WO2006085664A1 (fr) * 2005-02-10 2006-08-17 The University Of Tokyo Polymere chargeable de polycations et utilisation comme vecteur d'acides nucleiques
WO2007099660A1 (fr) * 2006-03-01 2007-09-07 The University Of Tokyo complexe de micelles polymeres contenant de l'acide nucleique

Non-Patent Citations (6)

* Cited by examiner, † Cited by third party
Title
"Meeting Info.: Digestive Disease Week/105th Annual Meeting of the American- Gastroenterological-Association. New Orleans, LA, USA. May 16 -20, 2004.", AMER GASTROENTEROL ASSOC *
"Nucleic Acids Symposium Series", 2004, article Y. TSUYAMA ET AL.: "Anti-tumor effects of naked plasmid DNA using a hydrodynamics-based procedure", pages: 239 - 240 *
K. J. CHANG ET AL.: "Effect of TNFerade Local Gene Therapy on Expression of Tumor Necrosis Factor Alpha (TNF-a) and Survivin in Tumors of Patients with Pancreatic Cancer : Can We Prove and Predict Response to Therapy?", GASTROENTEROLOGY, vol. 126, no. 4 SUPP, 2004, pages A623, W1372 *
N. ZAROVNI ET AL.: "Inhibition of Tumor Growth by Intramuscular Injection of cDNA Encoding Tumor Necrosis Factor a Coupled to NGR and RGD Tumor-homing Peptides", HUMAN GENE THERAPY, vol. 15, 2004, pages 373 - 382 *
SHI JIN ET AL.: "TNF-a/cycloheximide-induced apoptosis in intestinal epithelial cells requires Racl-regulated reactive oxygen species", AMERICAN JOURNAL OF PHYSIOLOGY - GASTROINTESTINAL AND LIVER PHYSIOLOGY, vol. 294, 2008, pages 928 - 937 *
Y. S. KIM ET AL.: "TNF-Induced Activation of the Noxl NADPH Oxidase and Its Role in the Induction of Necrotic Cell Death", MOLECULAR CELL, vol. 26, 2007, pages 675 - 687 *

Also Published As

Publication number Publication date
WO2011002077A8 (fr) 2011-04-07

Similar Documents

Publication Publication Date Title
Erel-Akbaba et al. Radiation-induced targeted nanoparticle-based gene delivery for brain tumor therapy
US20190062748A1 (en) Polyconjugates for Delivery of RNAi triggers to Tumor Cells In Vivo
Zhang et al. Cell-permeable NF-κB inhibitor-conjugated liposomes for treatment of glioma
ES2907486T3 (es) Métodos y composiciones para reducir metástasis
Wang et al. A self-assembled system for tumor-targeted co-delivery of drug and gene
US11213540B2 (en) Compositions and methods for inducing apoptosis
US20140349944A1 (en) Chaperone-based integrin inhibitors for the treatment of cancer and inflammatory diseases
JP5463531B2 (ja) Ｐｈｄ２発現抑制物質搭載ポリイオンコンプレックス
JP2006508635A (ja) 治療用または診断用試薬を送達するためのターゲティングタンパク質
JP5817055B2 (ja) ＴＮＦ−α、ＣＤ４０Ｌ及びＧＭ−ＣＳＦ併用遺伝子治療剤
WO2020144317A1 (fr) Peptides de pénétration cellulaire
WO2011002077A1 (fr) Agent de thérapie génique tnf-α contenu dans une micelle polymère
US20240139104A1 (en) Micelle releasing thermosensitive hydrogels as a therapeutic delivery system
WO2020158863A1 (fr) Médicament combinant une préparation de micelles contenant un composé anthracycline avec un immunostimulant
EP3889263A1 (fr) Gène codant pour la protéine humaine 2 ig-b7-h3 muté, vecteur recombiné, cellule hôte la contenant, composition pharmaceutique et application correspondante
WO2010100781A1 (fr) Composition pour l'administration d'acide nucléique
Deng et al. Long-term combined blockade of CXCR4 and PD-L1 with in vivo reassembly for intensive tumor interference
Liu et al. Unlocking the power of immunotherapy: Combinatorial delivery of plasmid IL-15 and gemcitabine to synergistically remodeling the tumor microenvironment
KR101327181B1 (ko) 신장세포특이적인 펩타이드 및 폴리에스테르아민 중합체를 포함하는 유전자 전달체 및 이를 포함한 신장 섬유증 예방 또는 치료용 조성물.
Chu Drug-free macromolecular therapeutics for treatment of B-cell malignancies
Klauber Harnessing Endogenous Systems for Cancer Therapy

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 10794245

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 10794245

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: JP