EP1112080A1 - REGULATION DE L'EXPRESSION D'ONCOGENE HER2/neu A L'AIDE DE POLYAMIDES SYNTHETIQUES - Google Patents

REGULATION DE L'EXPRESSION D'ONCOGENE HER2/neu A L'AIDE DE POLYAMIDES SYNTHETIQUES

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Publication number
EP1112080A1
EP1112080A1 EP99952908A EP99952908A EP1112080A1 EP 1112080 A1 EP1112080 A1 EP 1112080A1 EP 99952908 A EP99952908 A EP 99952908A EP 99952908 A EP99952908 A EP 99952908A EP 1112080 A1 EP1112080 A1 EP 1112080A1
Authority
EP
European Patent Office
Prior art keywords
polyamide
correspond
composition
her2
polyamides
Prior art date
Legal status (The legal status 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 status listed.)
Withdrawn
Application number
EP99952908A
Other languages
German (de)
English (en)
Other versions
EP1112080A4 (fr
Inventor
Peter B. California Inst. of Techology DERVAN
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California Institute of Technology CalTech
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California Institute of Technology CalTech
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Publication date
Application filed by California Institute of Technology CalTech filed Critical California Institute of Technology CalTech
Publication of EP1112080A1 publication Critical patent/EP1112080A1/fr
Publication of EP1112080A4 publication Critical patent/EP1112080A4/fr
Withdrawn legal-status Critical Current

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Classifications

    • 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
    • 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
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4164—1,3-Diazoles
    • A61K31/4178—1,3-Diazoles not condensed 1,3-diazoles and containing further heterocyclic rings, e.g. pilocarpine, nitrofurantoin
    • 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
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Definitions

  • Her-2/neu protein likely plays a role in cell motility and hence in metastasis.
  • inhibition of Her-2/neu gene expression by direct interference at the DNA level may be a potent therapeutic approach for metastatic disease.
  • transcription factors such as ESX, AP-2, and the TATA binding protein ("TBP") - play an important role in the regulation of the expression of the gene for the HER2/neu growth factor receptor. See Baert, J.-L. et al., Benz, C, et al; supra; Chang, C- H., et al., supra; Bosher, J. M., et al., Proc. Natl. Acad. Sci. USA 92, 744-747 (1995). These transcription factors activate the expression of pl85"ER2 U p on binding to sites within the HER2/neu promoter.
  • the nucleotide sequence of the HER2/neu promoter and a schematic representation are shown in Figure 1.
  • the present invention relates to and includes methods and compositions for the modulation, or regulation, of gene expression or overexpression by reducing the transcription of genes.
  • the transcription of specific individual target genes is reduced.
  • Such reductions result from the application of polyamides that bind or interact with the minor groove of double-stranded DNA (dsDNA) within the promoter regions of target genes.
  • the binding or interaction is with a predetermined target nucleic acid sequence within the promoter regions to inhibit or down-regulate transcription.
  • the polyamides bind to the minor groove of double-stranded DNA in a promoter region that controls the transcription and expression of a gene.
  • the transcription of the gene is inhibited by modulating the binding of a protein transcription factor to dsDNA.
  • the transcription factors are ESX, ETS, and TBP.
  • both classes of transcription factors can be inhibited by polyamides that contact or bind the minor groove of dsDNA.
  • DNA complexation of proteins contacting the minor groove may be inhibited by direct steric hinderance, repulsion, or exclusion or, alternatively, byallosteric efforts.
  • the binding of major groove binding proteins may be suppressed by a polyamide-induced change of the DNA conformation.
  • inhibition can also be achieved other ways, for example, by conjugating a DNA cleavage agent to a polyamide targeted to a desired site, or by chemically modifying DNA.
  • the expression or overexpression of oncogenes is targeted.
  • the oncogenes are endogenous cellular oncogenes involved in cancer, particularly human breast cancer.
  • One oncogene target of the invention is the HER-2/neu gene, which may be down-regulated or inhibited by the use of polyamides that bind to target sequences within the HER-2/neu promoter region.
  • these sequences are, or are proximal to, transcription factor binding sites within the HER2/neu promoter. Interactions or binding between the polyamide and the target sequence can inhibit the transcription of the HER2/neu gene.
  • the degree of inhibition of HER2/neu expression may be extensive and includes the inhibition of HER2/neu overexpression.
  • the invention further encompasses application of polyamides for the treatment of various tumors or cancers, including breast cancer.
  • Suitable polyamides have a binding affinity at the dsDNA target sequence of at least 10 9 M "1 and a selectivity of at least about two. Selectivity is defined as the ratio of the binding affinity for the identified dsDNA target sequence to the binding affinity for a single base-pair mismatch dsDNA sequence. In preferred embodiments, selectivity against at least 90% of single base mismatch sequences is greater than about 10.
  • the polyamides additionaly comprise at least two aliphatic amino acid residues chosen from the group consisting of glycine, ⁇ -alanine, ⁇ -aminobutyric acid, R-2,4-diaminobutyric acid, and 5 -amino valeric acid, and at least one terminal alkylamino residue, the polyamide having a binding affinity at the target dsDNA sequence of at least 10 9 M "! and a selectivity of at least about two, selectivity being defined as the ratio of the binding affinity for the identified target dsDNA sequence to the binding affinity for a single base- pair mismatch dsDNA sequence.
  • the invention further provides methods suitable for treating a subject having a condition associated with abnormal expression of a cellular oncogene.
  • the subject is preferably a human patient and, more particularly, one afflicted with breast cancer or other diseases or conditions associated with aberrant Her-2/neu oncogene expression.
  • Figure 1 depicts the HER2/neu promoter, showing the nucleotide sequence in A, including binding sites of Ets, AP-2, and TBP ("TATA") transcription factors and the "CCAAT box", and in B, a schematic diagram, not to scale.
  • Figure 2 A is a graphical representation of the results of a DNase I footprint titration of polyamide HER2-1 (left) and the mismatch polyamide ImPy- ⁇ -Pylm- ⁇ - PyPyPyPyPy- ⁇ -Dp (right) and 2B, the schematic structures and association constants of the polyamides, where the polyamides are represented by closed circles for imidazole rings, open circles for pyrrole rings, curved lines for ⁇ -aminobutyric acid, diamonds for ⁇ - alanine, and a half circle with a positive charge for dimethylaminopropylamide.
  • Figure 3 compares the sequence of the HER2/neu promoter and polyamide structures and binding sites; the binding site for the TATA binding protein (TBP) is indicated along with the structures of the polyamides HER2-A, HER2-1, 70, and the mismatch polyamide 86.
  • TATA binding protein TATA binding protein
  • Figure 4 is a graphical representation of the results of experiments showing the effects of polyamides Her2-1 (A) and 70 on TBP binding.
  • Figure 5 is a graphical representation of the results of experiments showing the effects of the polyamide HER2-1 on HER2/neu transcription in vitro in a cell free system.
  • Figure 6 is a graphical representation of the results of experiments showing the effects of the polyamides HER2-1 and 70 on HER2/neu mRNA production in the human breast cancer cell line SK-BR-2.
  • the polyamides are preferably cell-permeable and capable of inhibiting gene transcription in vivo, in vitro, or in cell free systems. Appropriate application of such polyamide molecules may be used to inhibit expression or overexpression of endogenous oncogenes as a treatment of various diseases, including cancer.
  • the polyamides bind to the minor groove of double stranded DNA in a promoter region that controls the transcription and expression of a target gene.
  • Preferred target genes are endogenous oncogenes involved in cancer formation or progression.
  • the transcription of the gene is inhibited by modulating the binding of a protein, such a transcription factor, to the same promoter region with which the polyamide binds or interacts.
  • the transcription factors are one or more of the following: ESX; ETS; and TBP.
  • the expression or overexpression of oncogenes is targeted.
  • the oncogenes are those implicated in human breast cancer, and their expression or overexpression is inhibited by polyamides that contact or bind the minor groove in the region of the oncogene promoter.
  • the contacted or bound portions of the promoter region are, or are proximal to, transcription factor binding sites.
  • the degree of inhibition is preferably large and more preferably enough to inhibit even overexpression of the oncogene, in when the copy number of the gene increases.
  • the polyamides used in the present invention comprise N-methylimidazole and N-methylpyrrole carboxamides. These polyamides generally have a crescent-shaped structure that permits interaction and complexation the minor groove of double-stranded DNA. NMR studies have confirmed that these compounds can bind to DNA in a 2:1 ratio by a motif in which two polyamide ligands are arranged in an antiparallel way, side- by-side to each other (Pelton, J., et al., Proc. Natl. Acad. Sci. USA 86, 5723-5727 (1986); Mrksich, M., et al., Proc. Natl. Acad. Sci. USA, 89, 7586-7590 (1992); Wade, W.
  • polyamides are called "hairpin polyamides", as they adopt a hairpin-like conformation in the DNA complex.
  • the sequence of the imidazole and the pyrrole carboxamides in the polyamide determines the DNA sequence specificity of the ligand, according to the scheme of carboxamide pairs that recognize nucleotide pairs described above.
  • the polyamides also preferably have a binding affinity at the target dsDNA sequence of at least 10 9 M "1 and a selectivity of at least about two, selectivity being defined as the ratio of the binding affinity for the identified target dsDNA sequence to the binding affinity for a single base-pair mismatch dsDNA sequence.
  • a mismatch polyamide of sequence composition Imlm- ⁇ -Imlm- ⁇ -PyPy- ⁇ -PyPy- ⁇ -Dp was also used in these studies.
  • Polyamide 70 of sequence composition ImPyPyPy- ⁇ -PyPyPyPy- ⁇ -Dp, binds the sequence 5'-AGTATA-3' overlapping the TATA box, while polyamide 86, of sequence composition ImPylmPy- ⁇ - PyPyPyPy- ⁇ -Dp, is a mismatch polyamide, with a single atom substitution from polyamide 70.
  • Figure 1 shows the sequence of the Her-2/neu promoter region and the binding sites of several transcription factors.
  • the hairpin polyamide ImPy- ⁇ -Pylm- ⁇ -PyPy- ⁇ - PyPy- ⁇ -Dp was synthesized to bind immediately downstream of the TBP binding site ( Figure 2B, left).
  • the polyamide compounds of the invention possess the ability to inhibit gene expression or overexpression, properties that are exploited in the treatment of any of a number of diseases or conditions, most notably cancer and especially breast cancer.
  • a composition of this invention may be active er se, or may act as a "pro-drug" that is converted in vivo to an active form.
  • Solid carriers include starch, lactose, calcium sulfate dihydrate, terra alba, sucrose, talc, gelatin, agar, pectin, acacia, magnesium stearate and stearic acid.
  • Liquid carriers include syrup, peanut oil, olive oil, saline, water, dextrose, glycerol and the like.
  • the carrier or diluent may include any prolonged release material, such as glyceryl monostearate or glyceryl distearate, alone or with a wax.
  • the preparation may be in the form of a syrup, elixir, emulsion, soft gelatin capsule, liquid containing capsule, sterile injectable liquid (e.g., a solution), such as an ampoule, or an aqueous or nonaqueous liquid suspension.
  • sterile injectable liquid e.g., a solution
  • an ampoule or an aqueous or nonaqueous liquid suspension.
  • the relative amounts of HER2/neu mRNA from the various cells can be determined using reverse transcriptase (RT)-polymerase chain reaction (PCR).
  • PCR reverse transcriptase
  • concentration of total RNA is determined by spectrophotometry (using the optical density at 260 nM) for each different cell type and polyamide concentration.
  • An equal amount of total RNA (10 ng) is used for each RT-PCR.
  • RT-PCR was carried out using the Reverse Transcription System kit (Promega).

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  • Health & Medical Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Epidemiology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

La présente invention concerne des méthodes et des compositions comprenant des polyamides pouvant se lier au sillon mineur d'ADN double brin destinées à inhiber ou à réduire la transcription et l'expression de gène. Ces polyamides comprennent au moins quatre paires de résidus carboxamides aromatiques choisis afin de contacter et/ou de se lier spécifiquement à la séquence de nucléotides d'une cible ADN à double brin dans la région promoteur du gène de la cible. Ces méthodes, compositions et polyamides sont destinés à l'inhibition de la transcription et de l'expression d'oncogène aussi bien qu'au traitement de cancer.
EP19990952908 1998-09-11 1999-09-10 REGULATION DE L'EXPRESSION D'ONCOGENE HER2/neu A L'AIDE DE POLYAMIDES SYNTHETIQUES Withdrawn EP1112080A4 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US9990698P 1998-09-11 1998-09-11
US99906P 1998-09-11
PCT/US1999/020971 WO2000015242A1 (fr) 1998-09-11 1999-09-10 REGULATION DE L'EXPRESSION D'ONCOGENE HER2/neu A L'AIDE DE POLYAMIDES SYNTHETIQUES

Publications (2)

Publication Number Publication Date
EP1112080A1 true EP1112080A1 (fr) 2001-07-04
EP1112080A4 EP1112080A4 (fr) 2002-10-24

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Family Applications (2)

Application Number Title Priority Date Filing Date
EP19990952908 Withdrawn EP1112080A4 (fr) 1998-09-11 1999-09-10 REGULATION DE L'EXPRESSION D'ONCOGENE HER2/neu A L'AIDE DE POLYAMIDES SYNTHETIQUES
EP99969022A Withdrawn EP1162961A2 (fr) 1998-09-11 1999-09-10 Regulation de l'expression oncongene de her2/neu par des polyamides synthetiques

Family Applications After (1)

Application Number Title Priority Date Filing Date
EP99969022A Withdrawn EP1162961A2 (fr) 1998-09-11 1999-09-10 Regulation de l'expression oncongene de her2/neu par des polyamides synthetiques

Country Status (5)

Country Link
EP (2) EP1112080A4 (fr)
JP (2) JP2002524501A (fr)
AU (2) AU768405B2 (fr)
CA (2) CA2343289A1 (fr)
WO (2) WO2000015209A2 (fr)

Families Citing this family (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1762614A3 (fr) * 2000-07-21 2007-05-30 Research Corporation Technologies, Inc. Promoteurs spécifiques de tumeur
US7122626B2 (en) 2001-04-26 2006-10-17 Genesoft Pharmceuticals, Inc. Halogen-substitued thienyl compounds
AU2002312371A1 (en) 2001-06-13 2002-12-23 Roland W. Burli Aryl-benzimidazole compounds having antiinfective activity
EP1470119A4 (fr) 2001-06-13 2005-10-19 Genesoft Pharmaceuticals Inc Composes benzothiophene presentant une activite anti-infectieuse
WO2002101007A2 (fr) 2001-06-13 2002-12-19 Genesoft Pharmaceuticals, Inc Composes benzamides anti-pathogenes
US6777425B2 (en) 2001-06-13 2004-08-17 Genesoft Pharmaceuticals, Inc. Isoquinoline compounds having antiinfective activity
DE60326735D1 (de) 2002-08-02 2009-04-30 Genesoft Pharmaceuticals Inc Biaryl-verbindungen mit antiinfektiver wirkung
US7265129B2 (en) 2002-10-25 2007-09-04 Genesoft Pharmaceuticals, Inc. Anti-infective biaryl compounds
JP2006509027A (ja) 2002-12-10 2006-03-16 オーシェント ファーマシューティカルズ コーポレーション （ピロールカルボキサミド）−（ベンズアミド）−（イミダゾールカルボキサミド）モチーフを有する抗菌化合物
JP3792655B2 (ja) 2003-01-20 2006-07-05 日本電気株式会社 新規な癌遺伝子、該癌遺伝子由来の組換えタンパク質、およびそれらの用途
JP2006022063A (ja) * 2004-07-09 2006-01-26 Univ Nihon Ｌｏｘ−１遺伝子発現抑制剤
US10307401B2 (en) * 2016-08-29 2019-06-04 California Institute Of Technology Compositions and methods for treatment of prostate cancer
US10723716B2 (en) 2016-12-21 2020-07-28 New York University Alpha-helix mimetics as modulators of Abeta self-assembly

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998035702A1 (fr) * 1996-08-01 1998-08-20 California Institute Of Technology Inhibition de la transcription de genes par des ligands de polyamide se fixant a l'adn

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998035702A1 (fr) * 1996-08-01 1998-08-20 California Institute Of Technology Inhibition de la transcription de genes par des ligands de polyamide se fixant a l'adn

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
See also references of WO0015242A1 *
WHITE S ET AL: "RECOGNITION OF THE FOUR WATSON-CRICK BASE PAIRS IN THE DNA MINOR GROOVE BY SYNTHETIC LIGANDS" NATURE, MACMILLAN JOURNALS LTD. LONDON, GB, vol. 391, no. 6666, 29 January 1998 (1998-01-29), pages 468-471, XP002072062 ISSN: 0028-0836 *

Also Published As

Publication number Publication date
EP1162961A2 (fr) 2001-12-19
JP2002524501A (ja) 2002-08-06
AU768405B2 (en) 2003-12-11
CA2344654A1 (fr) 2000-03-23
AU6496599A (en) 2000-04-03
WO2000015242A1 (fr) 2000-03-23
JP2002524525A (ja) 2002-08-06
EP1112080A4 (fr) 2002-10-24
WO2000015209A2 (fr) 2000-03-23
WO2000015209A3 (fr) 2000-05-25
AU6034199A (en) 2000-04-03
CA2343289A1 (fr) 2000-03-23

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