WO2020025710A1 - Micro-arn destiné à être utilisé dans la prévention et/ou le traitement du cancer invasif - Google Patents

Micro-arn destiné à être utilisé dans la prévention et/ou le traitement du cancer invasif Download PDF

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WO2020025710A1
WO2020025710A1 PCT/EP2019/070704 EP2019070704W WO2020025710A1 WO 2020025710 A1 WO2020025710 A1 WO 2020025710A1 EP 2019070704 W EP2019070704 W EP 2019070704W WO 2020025710 A1 WO2020025710 A1 WO 2020025710A1
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mir
compound
cancer
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cell
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Stefan EICHMÜLLER
Theresa KORDASS
Wolfram Osen
David EISEL
Rainer KÖNIG
Volker AST
Marcus OSWALD
Alexander Berndt
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Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts
Universitätsklinikum Jena
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/14—Type of nucleic acid interfering N.A.
    • C12N2310/141—MicroRNAs, miRNAs

Definitions

  • the present invention seeks to provide novel therapeutic strategies to prevent and treat proliferative diseases, such as colorectal cancer.
  • the invention is based on the idea of targeting microRNAs (miRNA or miR) which regulate the expression of extracellular matrix (ECM) genes involved in tumorigenesis as strategy for the treatment of cancer diseases.
  • the present invention is particularly based on modulating the expression or activity of a miR, preferably a human miR (hsa-miR) selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, which were identified to down- regulate target genes for ECM remodeling in the tumor environment in several tumor sets.
  • miR preferably a human miR (hsa-miR) selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16
  • the present invention hence provides compounds, which are agonists or mimics of the disclosed miR, which are particularly useful in the prevention and/or treatment of proliferative diseases, such as cancer diseases, preferably colorectal cancer.
  • CRC Colorectal carcinoma
  • CRC cancer diseases
  • Current therapeutic options for cancer diseases include surgeiy to remove the tumorous tissue during an operation, radiation therapy to destroy cancer cells, chemotherapy to prevent tumorous cells from growing and dividing further, and immunotherapy to boost the body's natural defense system to fight the cancer.
  • all of these treatment strategies have severe side effects.
  • many cancers become resistant to treatment, or patients relapse following treatment.
  • a continuous selection pressure exists to develop cancer cell clones which are resistant to the chemotherapeutic used.
  • miRNAs are short endogenous RNA molecules that are well-established players in the complex network of post-transcriptional regulation of gene expression in eukaryotic cells. It has been shown that miRNAs regulate their target genes in an expression-dependent, non-linear repression mechanism. Aberrantly expressed miRNAs can participate in tumor initiation, development, progression and invasion in multiple human cancer types, including colorectal cancer. The regulating effect of miRNAs can depend on the cell type, tissue, RNA- induced silencing complex (RISC) availability, binding site abundance and seed sequence complementarity.
  • RISC RNA- induced silencing complex
  • the miR-200 family contains 5 members, called miR-200a, miR-200b, miR-200c, miR-141, and miR-429.
  • the five members of the miR-200 family are found in two clusters.
  • miR-200a, miR-200b, and miR-429 are located on chromosome 1
  • miR-200c and miR- 141 are located on chromosome 12.
  • Members of the miR-200 family are highly enriched in epithelial tissues.
  • the miR-200 family, especially miR-200c is believed to play an essential role in metastasis and invasion of carcinoma due to its functional regulation of epithelial-to- mesenchymal transition (EMT), the initiating step of metastasis.
  • EMT epithelial-to- mesenchymal transition
  • the miR-i7 ⁇ 92 cluster maps to human chromosome 13 and is a well-characterized polycistronic miRNA cluster. This cluster encodes for six individual miRNAs, namely miR-17, miR-i8a, miR-i9a, miR-20a, miR-i9b-i, and miR-92a. Interestingly, the organization and sequences of the miR-i7 ⁇ 92 family are highly conserved among all vertebrates. miR-192 is located on human chromosome 11 and is involved in proliferation, cell cycle progression and apoptosis. It has been shown to have effects in several different cell types, including tumor cells and surrounding stromal cells.
  • the tumor microenvironment is a heterogeneous population of cells consisting of the tumor bulk plus supporting cells. It is becoming increasingly evident that these supporting cells are recruited by cancer cells from nearby endogenous host stroma and promote events such as tumor angiogenesis, proliferation, invasion, and metastasis.
  • recruited stromal cells include vascular endothelial cells, pericytes, adipocytes, fibroblasts, and bone-marrow mesenchymal stromal cells.
  • TASCs tumor- associated stromal cells
  • the extracellular matrix is a complex network of macromolecules with distinctive physical, biochemical, and biomechanical properties.
  • the ECM serves not only as the scaffold upon which tissues are organized but provides critical biochemical and biomechanical cues that direct cell growth, survival, migration and differentiation and modulate vascular development and immune function.
  • the ECM is commonly deregulated and becomes disorganized in diseases such as cancer.
  • Biochemical cues from the tumor-associated ECM modulate tumor cells and surrounding TASCs. Importantly, such cues between the evolving ECM, tumor cells and its TASCs affect cancer progression by directly promoting cellular transformation and metastasis.
  • the present invention seeks to provide treatment for a proliferative disease, preferably a proliferative disease that is characterized by high expression of at least one gene for an ECM protein, such as for FGF2, DST, PLODi, SPARC, L0XL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl or FBLN5.
  • an ECM protein such as for FGF2, DST, PLODi, SPARC, L0XL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl or FBLN5.
  • the growth factor FGF2 is part of many cellular processes and particularly known as an inducer of angiogenesis.
  • FGF2 is part of the FGFR-SRC-ITGAV/ITGB5 cascade that induces tumor cell adhesion to fibroblasts and tumor cell motility.
  • the linker protein DST is a component of hemidesmosomes and connects intermediate filaments to the actin cytoskeleton and helps to form actin bundles around the nucleus.
  • PLODi is another important collagen crosslinking enzyme necessary for the biogenesis and stability of collagens.
  • An increased expression of PLODi has been reported in breast cancer tissue compared to normal breast tissue.
  • LOXL2 is a member of the lysyl oxidase gene family which catalyzes the crosslinking of extracellular collagen and elastin resulting in increased ECM stiffness and subsequent activation of the kinases FAK and Src, which enable tumor cells to proliferate and invade. Furthermore, LOXL2 is involved in metastasis. LOXL2 expression is increased in a number of tumor types, including colorectal cancer. LOXL2 has been reported to be highly expressed in tumor-associated fibroblasts and is a marker for poor prognosis in colorectal cancer.
  • the genes ITGBi and ITGAV are members of the integrin gene family.
  • the cell surface peroxidase PXDN is, when secreted into the extracellular space, involved in ECM formation by crosslinking collagen IV. It has been shown that silencing of PXDN leads to significant reduction of cancer cell invasion in melanoma.
  • FBNi is an extracellular glycoprotein secreted by fibroblasts that forms long, elastic microfibrils as important components of the extracellular matrix. It has been shown to promote ovarian tumorigenesis and metastasis in mouse models. When highly expressed, FBNi indicates poor overall survival in ovarian cancer. Inhibition of E-cadherin and b- catenin and stimulation of matrix metallopeptidase 2, 9 and 13 are further characteristics of FBNi.
  • MMPs matrix metallopeptidases
  • TIMP2 matrix metallopeptidases
  • LAM Cl is a member of a large family of extracellular glycoproteins which form large polymers with other laminin isoforms and form a major constituent of the basement membrane besides collagen.
  • LAMCi is essential during embryonic development. In the context of tumor development, it is known that LAMCi promotes cancer cell migration and invasion in prostate cancer and enhances tumor cell motility and invasion in uterine and ovarian carcinomas.
  • the cytokine transforming growth factor beta 1 (TGFBi) is a well-known ligand and driver of TGF-b signaling which is central for tumor development and progression, angiogenesis, epithelial-to-mesenchymal transition and metastasis.
  • Tumor cell-derived TGF-b is actively promoting the transdifferentiation of fibroblasts to a myofibroblast- or tumor-associated fibroblast phenotype.
  • TGF-b is known to increase its own expression and the expression of MMP2.
  • the transcription factor ETSi plays a role in numerous tumor types and is linked to tumor progression, invasion, metastasis and angiogenesis. It is active in cancer cells, cancer- associated fibroblasts and endothelial cells. In particular in colorectal cancer, stromal protein levels of ETSi were significantly associated with the formation of lung metastasis.
  • SERPINHi (often referred to as HSP47) is involved in the synthesis and deposition of collagen as a chaperone present in the endoplasmatic reticulum.
  • SERPINHi is linked to the progression of breast cancer, cell migration and invasion of cancer cells in cervical squamous cell carcinoma, tumor growth and invasion in glioma.
  • Tissue Inhibitor of Metalloproteinases 2 can bind various matrix metalloproteinases (MMPs), a large family of ECM-degrading endopeptidases. TIMP2 inactivates them irreversibly to prevent excessive destruction of the ECM network and to maintain tissue homeostasis. TIMP2 in colorectal cancer samples shows a specific staining not only in epithelial tumor cells but also in the ECM of the stromal compartment. High expression of TIMP2 was associated with bad outcome in colorectal cancer and shortened disease-free and overall survival in human breast cancer.
  • MMPs matrix metalloproteinases
  • FNi mediates cell-matrix adhesion and is directly involved in cell migration and invasion by binding to the a 5 b i integrin transmembrane receptor dimer.
  • Overexpression of FNi was observed in myofibroblasts and in epithelial cells in samples of colorectal cancer patients. Elevated epithelial expression of FNi can be an indicator of lymph node metastasis in primary colorectal cancers.
  • the extracellular glycoprotein FBLN5 is involved in the formation of elastic fibers and antagonizes fibronectin-mediated signaling by binding to the same integrin receptors.
  • FBLN5 is known to be induced by TGF-b signaling in fibroblasts and endothelial cells and plays a role in initiating and enhancing epithelial-to-mesenchymal transition in normal and malignant mammary epithelial cells, explaining its tumor promoting role.
  • ECM components ECM degradation
  • ECM synthesis ECM - cell signaling
  • ECM degradation summarizes candidate genes like matrix metallopeptidases that are directly or indirectly involved in ECM decay.
  • Target genes that are involved in the synthesis of ECM structures or maintain its integrity like lysyl oxidase can be assigned to the category "ECM synthesis”.
  • ECM synthesis genes that are part of direct ECM - cell interactions like integrins or are involved in ECM - cell communication like TGF-b can be assigned to the category "ECM - cell signaling”.
  • a pharmaceutically modified regulation of ECM proteins could enable the development of better treatment strategies for proliferative diseases such as cancer diseases, which is urgently needed.
  • a compound for use in the prevention and/or treatment of a proliferative disease in a subject wherein the compound is an agonist or mimic of at least one micro RNA (miRNA, miR) selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • miRNA micro RNA
  • miR micro RNA
  • stromal subgroup is also used herein for the CMS4 subgroup.
  • the invention is based on the surprising finding that miR-200, miR-17, miR- 192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2 down-regulate target genes for ECM remodeling, such as the ones described above. Further, restoring miR- 200, miR-17 and/or miR-192 expression in tumor stroma cells reduces the invasive capacity of cancer cells, which supports the therapeutic application of the compounds of the invention.
  • micro RNA refers to a particular class of "small RNA molecules" as defined above.
  • micro RNA refers to a non-coding RNA comprising from about 3 to about 200, from about 4 to about 180, from about 5 to about 150, from about 6 to about 120, from about 7 to about 90, from about 8 to about 80, from about 10 to about 60, from about 20 to about 40 or from about 20 to about 30 nucleotides in length, which hybridizes to and regulates the expression of a coding RNA.
  • the mi-RNAs as referred to may be single or double-stranded and may be obtained from a micro-RNA precursor, such as a hairpin RNA precursor, by natural processing routes (e.g. using intact cells or cell lysates) or by synthetic routes (e.g. using isolated processing enzymes, such as the dicer enzyme or RNAase III).
  • the mi-RNAs may be obtained directly by biological or chemical synthesis without the involvement of a precursor.
  • the mi-RNAs can silence the activity of an oligonucleotide encoding for a protein by blocking its translation.
  • mi-RNA micro-RNA
  • miR mi-RNA sequence
  • a pre-miRNA is the product of cleavage of a primary mi-RNA transcript, or "pri-miR” by the double-stranded RNA-specific ribonuclease known as Drosha, but a pre- miRNAs can also be produced directly by biological or chemical synthesis without having been processed from a pri-miR.
  • a miR precursor according to the invention refers to a precursor of a herein references mature miR sequence.
  • miR base http://www.mirbase.org/
  • miRBase 22 release all sequences of precursor miR of such mature miR sequence shall be incorporated herein by reference (see also: miRBase: annotating high confidence microRNAs using deep sequencing data.
  • miRBase annotating high confidence microRNAs using deep sequencing data.
  • Kozomara A Griffiths-Jones S. NAR 2014 42:D68-D73
  • miRBase integrating microRNA annotation and deep-sequencing data.
  • Kozomara A Griffiths-Jones S. NAR 2011 39:1)152- D157; miRBase: tools for microRNA genomics.
  • seed sequence refers to the sequence that may have a role in the binding of the miRNA to the mRNA.
  • the seed sequence or seed region may be a conserved heptametrical sequence that is mostly situated at positions 2-7 from the miRNA 5 '-end. Even though base pairing of miRNA and its target mRNA does not match perfect, the “seed sequence” may be perfectly complementaiy.
  • the invention provides compounds useful in aiding the treatment of a cancer disease by reducing the invasive capacity of tumor cells.
  • the compounds of the invention are therefore useful for treating an invasive tumor disease, preferably a solid tumor disease such as colon cancer.
  • invasive tumor disease shall refer to a cancer that has spread beyond the layer of tissue in which it started into the normal surrounding tissues. Invasive cancers may or may not be metastatic.
  • prevention or treatment in particular is a reduction of tumor invasion.
  • the key process of extracellular matrix remodeling that the inventors identified is a hallmark of tumor cell migration, invasion and metastasis and may at least partially underlie the poor clinical course of patients with stromal subgroup tumors.
  • a medical“use” of the compounds of the invention shall preferably refer to a method for preventing or treating a disease in a subject, wherein the method comprises a step of administering to the subject a therapeutically effective amount of a compound comprising an agonist or mimic of at least one microRNA (miRNA, miR) selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • miRNA microRNA
  • Preferred compounds of the invention are agonists of the miR selected from miR-200c, miR-17, or miR-192.
  • Treatment is meant to include, e.g., treating, delaying or alleviating disease progression, reducing the symptoms of, or curing the disease or condition.
  • An“effective amount” is an amount of the compound(s) or the pharmaceutical composition as described herein that alleviates symptoms as found for the disease to be treated, such as a cancer disease. Alleviating is meant to include, e.g., preventing, treating, reducing the symptoms of, or curing the disease or condition.
  • the invention also includes a method for treating a subject at risk for a development and/or progression of a cancer disease, wherein a therapeutically effective amount of a compound as described above is provided.
  • the term“prevention” or“preventing” when used in the context of a subject refers to stopping, hindering, and/or slowing down the development or onset of a proliferative disease and in in particular the symptoms associated with the proliferative disease.
  • A“proliferative disease” in the context of the present invention shall preferably refer to a disease such as a cancer or a tumor disease.
  • Cancer diseases that can be treated by the compound of the present invention include, but are not limited to, lung cancer, bladder cancer, ovarian cancer, uterine cancer, endometrial cancer, breast cancer, liver cancer, pancreatic cancer, stomach cancer, cervical cancer, lymphoma, leukemia, acute myeloid leukemia, acute lymphocytic leukemia, salivary gland cancer, bone cancer, brain cancer, colon cancer, rectal cancer, colorectal cancer, kidney cancer, skin cancer, melanoma, squamous cell carcinoma, pleomorphic adenoma, hepatocellular carcinoma, and/or adenocarcinoma.
  • the proliferative disease is characterized by expression of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, L0XL2, ITGBl, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10.
  • the at least one extracellular matrix protein is preferably expressed in the tumor or tumor surrounding tissue (tumor environment), such as a stromal cell.
  • the term“subject” or“patient” preferably refers to a mammal, such as a mouse, rat, guinea pig, rabbit, cat, dog, monkey, or preferably a human, for example a human patient.
  • the subject of the invention may be at danger of suffering from a proliferative disease such as a cancer or a tumor disease, or suffer from a cancer or tumor disease, preferably, wherein the cancer disease is colorectal cancer.
  • a proliferative disease such as a cancer or a tumor disease
  • the cancer disease is colorectal cancer.
  • agonist in the context of the present invention shall include any kind of agonistic molecule.
  • Preferred agonists are, in the context of the invention, agonistic molecules that enhance the expression, function and/or stability of miRNAs, such as miR-200, miR-17, miR- 192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • Agonists of expression, function and/or stability of miRNAs such as miR-200, miR-17, miR-192, miR- 29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, in the context of the present invention, also include agonists of expression, function and/or stability that interact with one or more other components of the signaling pathway as disclosed herein and/or with one or more other genes that control the expression, function and/or stability of miRNAs, such as of miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR- 92a-i, and miR-92a-2.
  • the agonist of the invention may agonize the expression, function and/or stability of another gene that itself modulates the expression, function and/or stability of miRNAs of the invention.
  • modulators that act, for example, as an anti-sense nucleotide molecule against certain mRNA, preferably one that binds to, such as specifically binds to, a nucleic acid that regulates the expression of miRNAs of the invention.
  • agonist shall also refer to modulators that act as an anti-sense nucleotide molecule against mRNA of a molecule that regulates the expression of a gene that controls the expression, function and/or stability of miRNAs, such as miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2 for example a transcription factor for or activator protein of certain miRNAs, such as of miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • Mechanisms by which such modulation may be brought about, and/or the effects of such modulation can include one or more of those as described elsewhere herein.
  • the term“expression” refers to the process by which a polynucleotide is transcribed from a DNA template (e.g. into mRNA or other RNA transcripts), and later on translated into a peptide or protein.
  • the term“transcription” refers to the process of producing RNA from a DNA template.
  • “In vitro transcription” refers to the process of transcription of a DNA sequence into RNA molecules using a laboratory medium which contains an RNA polymerase and RNA precursors.
  • “in vivo transcription” refers to the process of transcription of a DNA sequence into RNA molecules, within a living organism.
  • the agonist of expression, function and/or stability of certain miRNAs is, in some embodiments, selected from a compound which is a polypeptide, peptide, glycoprotein, a peptidomimetic, a small molecular compound, a polynucleotide molecule, such as a molecule comprising a DNA molecule, an RNA molecule, a DNA aptamer, an RNA aptamer, a PNA (peptide nucleic acid), an LNA (locked nucleic acid), and the like, including i RNA (miR), dsRNA, easyRNA, siRNA, shRNA, and other RNAi constructs. Also included shall begenetic constructs for targeted gene editing, such as a CRISPR/Cas9 construct and/
  • the compounds of the present invention specifically and/or selectively influence the expression, function and/or stability of miRNAs, such as miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2 in cells showing a pathological form of cell proliferation, growth or survival, or in surrounding cells of such pathological cells, such as stromal cells.
  • miRNAs such as miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2 in cells showing a pathological form of cell proliferation, growth or survival, or in surrounding cells of such pathological cells, such as stromal cells.
  • Preferred examples are cells that have become a tumor cell, or originate from a tumor cell, and their surrounding tumor cells.
  • i RNA mimics refer to double-stranded, synthetic replicates of endogenous miRNAs, which augment the intracellular concentration and function of a specific endogenous i RNA, such as miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • i RNA mimics of the present invention are, in some embodiments, specifically and/or selectively useful in cells showing a pathological form of cell proliferation, growth or survival, or in surrounding stromal cells of such pathological cells.
  • Preferred examples are cells that have become a tumor cell, or originate from a tumor cell, and their surrounding tumor cells, such as tumor stromal cells.
  • the miRNAs which are preferably selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, as used in context of the herein disclosed invention shall pertain to a miRNA molecule, comprising, or consisting essentially of a sequence of the human variants of the aforementioned miRs, as annotated in miRbase fhttp://www.mirbase.org/ ⁇ ) in the database version of March 2018 (release 22).
  • the sequences of the miR of the invention are provided in any of SEQ ID Nos: 1-15.
  • the term shall also refer to a miRNA comprising the ribonucleotide sequence according to any of SEQ ID Nos: 1-15 with any modifications. Such modifications preferably do not affect the function of the miRNAs of the present invention, preferably of miR-200, miR-17, and miR-192.
  • Modifications in the context of the invention may be selected from any chemical modification, such as a modified internucleoside linkage, for example a phosphorothiate linkage, a modified nucleobase, for example a Cs-thiazole uracil, a Cs-propynyl-cytosine, a phenoxazine-modified cytosine, or a 2’-fluoro nucleotide, and a modified sugar moiety, or any modifications known to the person skilled in the art.
  • a modified internucleoside linkage for example a phosphorothiate linkage
  • a modified nucleobase for example a Cs-thiazole uracil, a Cs-propynyl-cytosine, a phenoxazine-modified cytosine, or a 2’-fluoro nucleotide
  • a modified sugar moiety or any modifications known to the person skilled in the art.
  • a miR or miR agonist of the invention may be provided as RNA, DNA or any hybrid or variant thereof.
  • the agonist or mimic is a compound that agonizes or mimics the expression, stability, and/or function of the at least one miRNA selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • An“agonist” of expression, function and/or stability of miRNAs selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, in the context of the invention may be any compound that affects the expression, function and/or stability of miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and/or miR-92a-2or derivatives thereof.
  • the compound agonizes or mimics the expression, stability, and/or function of the at least one miRNA in a cell associated with the proliferative disease to be treated, such as a tumor cell and/or a tumor stromal cell.
  • a tumor cell and/or a tumor stromal cell.
  • the term“cell” shall mean any mammalian cell and can include any mammalian cell type. In particular embodiments the term“cell” shall refer preferentially to a tumor cell or a tumor stromal cell.
  • tumor cell shall refer to any cell showing a pathological form of cell proliferation, growth or survival.
  • stromal cell shall refer to any stromal cell, such as a cell selected from vascular endothelial cells, pericytes, adipocytes, fibroblasts, and bone-marrow mesenchymal stromal cells.
  • the term“tumor stromal cell” shall refer to stromal cells surrounding tumor cells. Particularly, such cells are recruited by tumor cells from nearby endogenous host stroma and are then involved in events such as tumor angiogenesis, proliferation, invasion, and metastasis. Particularly, tumor cells can co-opt reactive stromal cells and transition them into tumor- associated stromal cells (TASCs). These TASCs are characterized by higher expression of proteins, such as matrix metalloproteinases, compared with their normal, non-reactive counterparts.
  • proteins such as matrix metalloproteinases
  • the compound for use is selected from a polypeptide, a peptide, a glycoprotein, a peptidomimetic, a small molecular compound, and a polynucleotide molecule.
  • Polynucleotide refers to a polymeric form of nucleotides of any length.
  • Polynucleotides may have any three dimensional structure and may perform any function, known or unknown.
  • polynucleotide molecule as compound for use, which comprises a DNA molecule, an RNA molecule, a DNA aptamer, an RNA aptamer, a PNA (peptide nucleic acid), an LNA (locked nucleic acid), or combinations thereof.
  • a further preferred embodiment of the present invention then relates to a polynucleotide molecule as compound for use, which is selected from i RNA (miR), dsRNA, easyRNA, siRNA, shRNA, and other RNAi constructs.
  • the polynucleotide molecule is described in one particular embodiment of the present invention to be a i RNA molecule selected from miR-200, miR-17, miR-192, a miR- derivative thereof, and a miR-precursor thereof.
  • a polynucleotide molecule comprising, consisting essentially of, or consisting of a sequence as shown in any one of SEQ ID Nos: 1-15, or a miR precursor thereof.
  • a miR-derivative of miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and/or miR-92a-2 which is preferably an agonist in context of the invention, as used in some embodiments, comprises, consists essentially of or consists of a nucleotide sequence having at least 60%, 70%, preferably at least 80%, such as at least 90% sequence identity to any of SEQ ID Nos: 1-15, and most preferably at least 95% (such as at least 99%) sequence identity to any of SEQ ID Nos: 1-15, or of a miR precursor thereof.
  • the derivative of miR-200, miR-17, miR-192, miR-29b-i, miR- 29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2 comprises, consists essentially of or consists of a nucleotide sequence having at least 80% sequence identity to the sequence shown in any of SEQ ID Nos: 1-15, or of a miR precursor thereof.
  • the miR derivative in accordance with the invention comprises a sequence comprising at least the seed sequence of any one of the miR shown in SEQ ID NO: 1 to 15, or of a miR precursor thereof.
  • the seed seqeuences are known to the skilled artisan and may be derived from the miRbase using the herein used identifiers with the database version of July 16, 2018.
  • miR-200 shall be selected from miR-200c, miR-200a, and miR-200b. Particularly preferred is the compound for use of the present invention, wherein the miR-200 is miR-200c.
  • a miR-derivative shall include any structural derivative of the miRs of the present invention, particularly miR-200, miR-17, miR-192, miR- 29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, but preferably such derivatives of the invention are characterized by that they maintain the biological function/activity of the parent miR molecule (with regard to the miR target), or at least in part, or preferably have an increased activity compared to the parent miR molecule (preferably the silencing activity towards the miR-target).
  • the derivative of the invention comprises at least one chemical modification, for example a chemical modification selected from a modified internucleoside linkage, a modified nucleobase, and a modified sugar moiety.
  • a further embodiment then relates to a derivative comprising at least one modified internucleoside linkage, which is a phosphorothiate linkage.
  • the derivative comprises at least one modified nucleobase, which is a Cs-thiazole uracil, a Cs-propynyl-cytosine, a phenoxazine-modified cytosine, and/or a 2’-fluoro nucleotide.
  • modified nucleobase which is a Cs-thiazole uracil, a Cs-propynyl-cytosine, a phenoxazine-modified cytosine, and/or a 2’-fluoro nucleotide.
  • a derivative that comprises at least one modified sugar, which is a 2'-0- methyl ribose, a 2'-0-methoxyethyl ribose, a 2'-0-propyl ribose, and/or a bicyclic sugar moiety.
  • An additional embodiment of the present invention then relates to an agonist or mimic, which is a DNA molecule encoding for the at least one miRNA selected from miR-200, miR- 17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, a miR- derivative thereof, and a miR-precursor thereof.
  • the DNA molecule encoding for the at least one miRNA selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, the miR-derivative thereof, or the miR-precursor thereof is a vector comprising a nucleic acid sequence, preferably an expressible sequence, encoding miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, the miR-derivative thereof, and/or the miR-precursor thereof.
  • vector shall include, without being limited thereto, any nucleic acid molecules that are single-stranded, double-stranded, or partially double-stranded.
  • the nucleic acid molecules of the present invention can comprise one or more free ends, or no free ends (such as circular molecules).
  • vector refers to a circular double stranded DNA loop into which additional DNA segments can be inserted, such as by standard molecular cloning techniques known to the person skilled in the art.
  • viral vector refers to a viral vector, wherein virally-derived DNA or RNA sequences are present in the vector for packaging into a virus, such as a retrovirus, adenovirus, or adeno-associated virus.
  • Viral vectors also include polynucleotides carried by a virus for transfection into a host cell.
  • Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g. bacterial vectors having a bacterial origin of replication). Other vectors (e.g.
  • non-episomal mammalian vectors are integrated into the genome of a host cell upon introduction into the host cell, and thereby replicated along with the host genome. Moreover, certain vectors are able to direct the expression of genes to which they are operatively linked. Such vectors are referred to as“expression vectors”. Common expression vectors of utility in recombinant DNA techniques are often in the form of plasmids.
  • the vector is a viral vector, a non-viral vector, an integrating vector, a non-integrating vector, a cosmid, a phagemid, and/or a plasmid.
  • vector which is an expression construct comprising a promoter sequence operably linked to a coding sequence.
  • Recombinant expression constructs can comprise a nucleic acid in a form suitable for expression of the nucleic acid in a host cell.
  • the recombinant expression vectors include one or more regulatoiy elements, which may be selected on the basis of the host cells to be used for expression. Such regulatoiy elements need to be operatively-linked to the nucleic acid sequence to be expressed.
  • “operably linked” is intended to mean that the nucleotide sequence of interest is linked to the regulatory element(s) in a manner that allows for expression of the nucleotide sequence (e.g. in an in vitro transcription/translation system or in a host cell when the vector is introduced into the host cell).
  • regulatory element is intended to include promoters, enhancers, internal ribosomal entry sites (IRES), and other expression control elements (e.g. transcription termination signals, such as polyadenylation signals and poly-U sequences). Regulatoiy elements include those that direct constitutive expression of a nucleotide sequence in many types of host cells.
  • coding sequence shall refer to the portion of DNA and the transcribed mRNA, which is translated by a ribosome into peptide or protein. Such a coding region has a start codon close to its 5’ end and a stop codon close to its 3’ end.
  • the coding region in mRNA has a five prime untranslated region (s'-UTR) and a three prime untranslated region (3'-UTR).
  • the expression construct comprising a coding sequence is under a transcriptional control of a cis acting regulatoiy element.
  • Cis acting regulatoiy element shall refer to a region of non-coding DNA which regulates the transcription of neighboring genes. CREs are present on the same molecule of DNA as the gene they regulate.
  • One example of a CRE is the operator in the lac operon. This DNA sequence is bound by the lac repressor, which, in turn, prevents transcription of the adjacent genes on the same DNA molecule. The lac operator is, thus, considered to "act in cis" on the regulation of the nearby genes. The operator itself does not code for any protein or RNA.
  • trans-regulatory elements are diffusible factors, usually proteins that may modify the expression of genes distant from the gene from which they were transcribed.
  • a further embodiment of the present invention then relates to the agonists or mimics of the invention, which are DNA molecules encoding for miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and/or miR-92a-2, wherein the miR-200 is selected from miR-200c, miR-200a, and miR-200b, preferably wherein the miR-200 is miR- 200c.
  • agonists or mimics of the invention which are DNA molecules encoding for miRNAs that comprise, consist essentially of, or consist of a sequence shown in any one of, or a multitude of, any of SEQ ID Nos: 1-15, or of a miR precursor thereof.
  • An additional embodiment of the invention relates to agonists or mimics, which are DNA molecules encoding for miR-derivatives .
  • miR-derivatives as used in some embodiments of the present invention, comprise, consist essentially of or consist of a nucleotide sequence having at least 60%, 70%, preferably at least 80%, such as at least 90% sequence identity to any of SEQ ID Nos: 1-15, and most preferably at least 95% (such as at least 99%) sequence identity to any of SEQ ID Nos: 1-15, or of a miR precursor thereof.
  • the derivative of miR-200, miR-17, and/or miR-192 comprises, consists essentially of or consists of a nucleotide sequence having at least 80% sequence identity to the sequence shown in any of SEQ ID Nos: 1-15, or of a miR precursor thereof.
  • the proliferative disease to be prevented and/or treated can be characterized by expression of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5,
  • an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5,
  • extracellular matrix protein shall refer to any protein present in the ECM, which is a complex network of macromolecules with distinctive physical, biochemical, and biomechanical properties that serves as a scaffold upon which tissues are organized. Many of such ECM proteins can serve as cues that direct growth, survival, migration and differentiation of surrounding cells.
  • the expression of SPARC, FGF2, DST, PLODi, LOXL2, ITGBl, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10 is preferably occurring in a cell associated with the proliferative disease, such as a tumor cell and/or an associated tumor stromal cell.
  • the expression of at least one gene for an extracellular matrix protein is a detectable expression, optionally an increased expression in the cell associated with the proliferative disease compared to a cell not associated with the disease, such as a healthy cell.
  • the expression can be detected by any standard method known to the person skilled in the art, such as by a polymerase chain reaction (PCR), or by determining the density of a spot on a microarray, on a hybridization blot, such as a Northern blot, or on an immunoblot, such as a Western blot, or on a bead array
  • PCR polymerase chain reaction
  • the compounds of the present invention can be used to treat any proliferative disease.
  • Preferred proliferative diseases to be treated are cancer diseases.
  • the cancer disease to be prevented and/or treated is selected from lung cancer, bladder cancer, ovarian cancer, uterine cancer, endometrial cancer, breast cancer, liver cancer, pancreatic cancer, stomach cancer, cervical cancer, lymphoma, leukemia, acute myeloid leukemia, acute lymphocytic leukemia, salivaiy gland cancer, bone cancer, brain cancer, colon cancer, rectal cancer, colorectal cancer, kidney cancer, skin cancer, melanoma, squamous cell carcinoma, pleomorphic adenoma, hepatocellular carcinoma, and/or adenocarcinoma.
  • the miR and miR agonists are useful for the prevention and/treatment of the disorders: hsa-miR-29b-i for a cancer selected from Colon, Gastric, Melanoma, Breast, Esophagel, HNSCC, Pancreas, Testicular; hsa-miR-29b-2 for the following cancers: Colon, Gastric, Breast, Uveal_Melanoma, Melanoma, Esophagel, HNSCC, Pancreas, Testicular; hsa- miR-92a-i for the the following cancers: Bladder, Gastric, Melanoma, Esophagel, HNSCC, Lung squamous cell, Colon, Prostate, HNSCC; hsa-miR-92a-2 for the following cancers: Bladder, Gastric, Rectum, Thyroid, Testicular, Colon, Prostate; hsa-miR-16-1 for the following cancers: Bladder,
  • the cancer disease to be prevented and/or treated is colorectal cancer, such as preferably a colorectal cancer of the stromal tumor subtype (CMS4).
  • CMS4 stromal tumor subtype
  • colorectal cancer which is localized colorectal cancer (stages o, I, or II), or advanced adenomas (stages III, or IV).
  • One embodiment relates to the compound of the present invention, which is reducing the expression of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNi, LAMCi, TGFBi, ETSi, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLi, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10.
  • an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGB
  • a compound which reduces the expression of the at least one gene in a cell, such as a tumor cell and/or a tumor stromal cell, optionally wherein the tumor stromal cell is surrounding the tumor cell.
  • An additional embodiment relates to the compound for use of the present invention, wherein the compound is administered to the subject in a therapeutically effective amount for preventing and/or treating the proliferative disease as described above.
  • Administration of an agent can be accomplished by any method, which allows the agent to reach the target cells. These methods include, e.g., injection, deposition, implantation, suppositories, oral ingestion, inhalation, topical administration, or any other method of administration where access to the target cells by the agent is obtained. Injections can be, e.g., intravenous, intradermal, subcutaneous, intramuscular or intraperitoneal.
  • Implantation includes inserting implantable drug delivery systems, e.g., microspheres, hydrogels, polymeric reservoirs, cholesterol matrices, polymeric systems, e.g., matrix erosion and/or diffusion systems and non-polymeric systems, e.g., compressed, fused or partially fused pellets.
  • Suppositories include glycerin suppositories.
  • Oral ingestion doses can be enterically coated.
  • Inhalation includes administering the agent with an aerosol in an inhalator, either alone or attached to a carrier that can be absorbed.
  • the agent can be suspended in liquid, e.g., in dissolved or colloidal form.
  • the liquid can be a solvent, partial solvent or non-solvent. In many cases, water or an organic liquid can be used.
  • Another embodiment relates to the compound for use of the present invention, wherein the compound is administered to a subject, which is a mammal, such as a mouse, a rat, a guinea pig, a rabbit, a cat, a dog, a monkey, or a human, preferably a human patient.
  • a subject which is a mammal, such as a mouse, a rat, a guinea pig, a rabbit, a cat, a dog, a monkey, or a human, preferably a human patient.
  • the prevention and/or treatment comprises the administration of the compound to the subject, preferably wherein the compound is administered to the subject by oral, intranasal, topical, rectal, bronchial, vaginal or parenteral administration, such as, for example, by intravascular administration, intravenous bolus injection, intravenous infusion, intra-arterial bolus injection, intra-arterial infusion, peri- tissue injection, such as peri-tumoral injection, intra-tissue injection, such as intra-tumoral injection, intra-retinal injection, subretinal injection, subcutaneous injection, direct application, inhalation, or by any clinically/medically accepted method.
  • intravascular administration intravenous bolus injection, intravenous infusion, intra-arterial bolus injection, intra-arterial infusion, peri- tissue injection, such as peri-tumoral injection, intra-tissue injection, such as intra-tumoral injection, intra-retinal injection, subretinal injection, subcutaneous injection, direct application, in
  • the compound is administered to the subject, in some embodiments, as a naked polynucleotide molecule, or in conjunction with a delivery reagent.
  • the compound is administered to the subject in conjunction with a lipophilic reagent, lipofectin, lipofectamine, cellfectin, polycations, and/or liposomes.
  • the object of the present invention is further solved by a combination for use in the prevention and/or treatment of a proliferative disease, comprising at least two agonists and/or mimics as described herein before, wherein said agonists or mimics are agonists or mimics of one or more miR selected from miR-200, miR-17, and miR-192.
  • Another embodiment of the invention relates to a combination for use in the prevention and/or treatment of a proliferative disease, comprising:
  • an agonist or mimic of miR-17 and an agonist or mimic of miRi92 or d. an agonist or mimic of miR-200 and an agonist or mimic of miR-17 and an agonist or mimic of miRi92.
  • the combination for use comprises, in some embodiments, miR-200, which is selected from miR-200c, miR-200a, and miR-200b.
  • miR-200 is miR-200c.
  • the combination alternatively or additionally may comprise any of miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and/or miR-92a-2.
  • the combination is preferably a combination of two, three or more individual miRs, or miR agonists selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • a pharmaceutical composition for use in the prevention and/or treatment of a proliferative disease comprising
  • At least one compound which is an agonist or mimic of at least one miRNA selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, or a combination for use in the prevention and/or treatment of a proliferative disease as described above; and
  • Carriers, excipients and strategies to formulate a pharmaceutical composition for example to be administered systemically or topically, by any conventional route, in particular enterally, e.g. orally, e.g. in the form of tablets or capsules, parenterally, e.g. in the form of injectable solutions or suspensions, topically, e.g. in the form of lotions, gels, ointments or creams, or in nasal or a suppositoiy form are well known to the person of skill and described in the respective literature.
  • the object of the present invention is further solved by providing a method of preventing and/or treating a proliferative disease in a subject, the method comprising administering to the subject an effective amount of a compound, which is an agonist or mimic of at least one miRNA selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16- 2, miR-92a-i, and miR-92a-2, or a combination for use in the prevention and/or treatment of a proliferative disease as described above, or a pharmaceutical composition as described above, thereby preventing and/or treating the proliferative disease in the subject.
  • a compound which is an agonist or mimic of at least one miRNA selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16- 2, miR-92a-i, and miR-92a-2, or a combination for use in
  • the above-described method causes a reduction of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, L0XL2, ITGBl, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10, preferably in a cell, which is associated with a tumor disease, more preferably wherein the cell is a tumor cell or a cell
  • the reduction of the at least one gene for an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBl, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10 preferably in a cell, which is associated with a tumor disease, more preferably wherein the cell is a tumor cell or a cell surrounding a
  • the object of the present invention is additionally solved by providing a method of reducing the expression of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBl, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10 in a cell of a subject, the method comprising introducing into the cell an effective amount of a compound, which is an
  • An additional embodiment relates to the method as described above, wherein the cell is a cell associated with a tumor disease, preferably wherein the cell is a tumor cell or a cell surrounding a tumor cell such as a tumor stromal cell.
  • the method as described above comprising the administration of an effective amount of the compound of the present invention, a combination according to the present invention, or a pharmaceutical composition according to the present invention, to the subject.
  • the object of the present invention is further solved by the use of an agonist or mimic of at least one miRNA selected from miR-200, miR-17, and miR-192 for the manufacture of a medicament for use in the prevention and/or treatment of a proliferative disease.
  • An additional embodiment of the invention relates to the use as described above, wherein a reduction of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, L0XL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNi, LAMCi, TGFBi, ETSi, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLi, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10, is therapeutically beneficial.
  • an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi
  • Item 1 A compound for use in the prevention and/or treatment of a proliferative disease in a subject, wherein the compound is an agonist or mimic of at least one micro RNA (miRNA, miR) selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • miRNA micro RNA
  • Item 2 The compound for use according to item 1, wherein the agonist or mimic is a compound that agonizes or mimics the expression, stability, and/or function of the at least one miRNA.
  • Item 3 The compound for use according to item 1 or 2, wherein the compound agonizes or mimics the expression, stability, and/or function of the at least one miRNA in a cell associated with the proliferative disease, such as a tumor cell and/or a tumor stromal cell.
  • Item 4 The compound for use according to any one of items 1 to 3, wherein the compound is selected from a polypeptide, a peptide, a glycoprotein, a peptidomimetic, a small molecular compound, and a polynucleotide molecule.
  • Item 5 The compound for use according to any one of items 1 to 4, wherein the compound is a polynucleotide molecule and comprises a DNA molecule, an RNA molecule, a DNA aptamer, an RNA aptamer, a PNA (peptide nucleic acid), an LNA (locked nucleic acid), or combinations thereof.
  • Item 6 The compound for use according to item 4 or 5, wherein the polynucleotide molecule is selected from miRNA (miR), dsRNA, easyRNA, siRNA, shRNA, and other RNAi constructs.
  • Item 7 The compound for use according to any one of items 4 to 6, wherein the polynucleotide molecule is a miRNA molecule selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, a miR-derivative thereof, and a miR-precursor thereof.
  • the polynucleotide molecule is a miRNA molecule selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, a miR-derivative thereof, and a miR-precursor thereof.
  • Item 8 The compound for use according to any one of items 1 to 7, wherein the miR-200 is selected from miR-200c, miR-200a, and miR-200b, and preferably is miR-200c.
  • Item 9 The compound for use according to any one of items 1 to 8, wherein the miR is a human miR (hsa-miR).
  • Item 10 The compound for use according to any one of items 4 to 9, wherein the polynucleotide molecule comprises, consists essentially of, or consists of a sequence shown in any one of SEQ ID Nos: 1-15, or a miR precursor thereof.
  • Item 11 The compound for use according to any one of items 7 to 10, wherein the miR- derivative comprises, consists essentially of or consists of a nucleotide sequence having at least 80% sequence identity to the sequence shown in SEQ ID Nos: 1-15, or a miR precursor thereof.
  • Item i2 The compound for use according to any one of items 7 to 11, wherein the miR- derivative comprises at least one chemical modification, for example a chemical modification selected from a modified internucleoside linkage, a modified nucleobase, and a modified sugar moiety.
  • Item 13 The compound for use according to item 12, wherein the at least one modified internucleoside linkage is a phosphorothiate linkage.
  • Item 14 The compound for use according to item 12 or 13, wherein the at least one modified nucleobase comprises a Cs-thiazole uracil, a Cs-propynyl-cytosine, a phenoxazine-modified cytosine, and/or a 2’-fluoro nucleotide.
  • Item 15 The compound for use according to any one of items 12 to 14, wherein the at least one modified sugar moiety comprises a 2'-0-methyl ribose, a 2'-0-methoxyethyl ribose, a 2'-
  • Item 16 The compound for use according to any one of items l to 5, wherein the agonist or mimic is a DNA molecule encoding for the at least one miRNA selected from miR-200, miR- 17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, the miR-derivative thereof, and the miR-precursor thereof.
  • the agonist or mimic is a DNA molecule encoding for the at least one miRNA selected from miR-200, miR- 17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, the miR-derivative thereof, and the miR-precursor thereof.
  • Item 17 The compound for use according to item 16, wherein the DNA molecule is a vector comprising a nucleic acid sequence, preferably an expressible sequence, encoding miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, the miR-derivative thereof, and/or the miR-precursor thereof.
  • the DNA molecule is a vector comprising a nucleic acid sequence, preferably an expressible sequence, encoding miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2, the miR-derivative thereof, and/or the miR-precursor thereof.
  • Item 18 The compound for use according to item 17, wherein the vector is a viral vector, a non-viral vector, an integrating vector, a non-integrating vector, a cosmid, a phagemid, and/or a plasmid.
  • the vector is a viral vector, a non-viral vector, an integrating vector, a non-integrating vector, a cosmid, a phagemid, and/or a plasmid.
  • Item 19 The compound for use according to item 17 or 18, wherein the vector is an expression construct comprising a promoter sequence operably linked to a coding sequence.
  • Item 20 The compound for use according to any one of items 17 to 19, wherein the coding sequence is under a transcriptional control of a cis acting regulatoiy element.
  • Item 21 The compound for use according to any one of items 16 to 20, wherein the miR-200 is selected from miR-200c, miR-200a, and miR-200b, preferably wherein the miR-200 is miR-200c.
  • Item 22 The compound for use according to any one of items 16 to 21, wherein the miRNA comprises, consists essentially of, or consists of a sequence shown in any one of SEQ ID Nos: 1-15, or a miR precursor thereof.
  • Item 23 The compound for use according to any one of items 16 to 22, wherein the miR- derivative comprises, consists essentially of or consists of a nucleotide sequence having at least 80% sequence identity to the sequence shown in any one of SEQ ID Nos: 1-15, or a miR precursor thereof.
  • Item 24 The compound for use according to any one of items 1 to 23, wherein the proliferative disease is characterized by expression of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNi,
  • Item 25 The compound for use according to item 24, wherein the expression is an expression in a cell associated with the proliferative disease, such as a tumor cell and/or tumor stromal cell.
  • Item 26 The compound for use according to item 25, wherein the expression is a detectable expression, optionally an increased expression in the cell associated with the proliferative disease compared to a cell not associated with the disease, such as a healthy cell.
  • Item 27 The compound for use according to any one of items 1 to 26, wherein the proliferative disease is a cancer disease.
  • Item 28 The compound for use according to item 27, wherein the cancer disease is selected from lung cancer, bladder cancer, ovarian cancer, uterine cancer, endometrial cancer, breast cancer, liver cancer, pancreatic cancer, stomach cancer, cervical cancer, lymphoma, leukemia, acute myeloid leukemia, acute lymphocytic leukemia, salivary gland cancer, bone cancer, brain cancer, colon cancer, rectal cancer, colorectal cancer, kidney cancer, skin cancer, melanoma, squamous cell carcinoma, pleomorphic adenoma, hepatocellular carcinoma, and/or adenocarcinoma.
  • the cancer disease is selected from lung cancer, bladder cancer, ovarian cancer, uterine cancer, endometrial cancer, breast cancer, liver cancer, pancreatic cancer, stomach cancer, cervical cancer, lymphoma, leukemia, acute myeloid leukemia, acute lymphocytic leukemia, salivary gland cancer, bone cancer, brain cancer, colon cancer, rectal cancer, colorec
  • Item 29 The compound for use according to item 27 or 28, wherein the cancer disease is colorectal cancer.
  • Item 30 The compound for use according to item 28 or 29, wherein the colorectal cancer is localized colorectal cancer (stages o, I, or II), or advanced adenomas (stages III, or IV).
  • Item 31 The compound for use according to any of items 1 to 30, wherein the compound is reducing the expression of the at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNi, MMP2, FSCNi, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBi, CLASPi, FURIN, TLLi, CTSD, MMP16, COL3A1, NIDi, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or AD AM10.
  • an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi,
  • Item 32 The compound for use according to any of items 1 to 31, wherein the compound reduces the expression of the at least one gene in a cell, such as a tumor cell and/or a tumor stromal cell, optionally wherein the tumor stromal cell is surrounding the tumor cell.
  • a cell such as a tumor cell and/or a tumor stromal cell, optionally wherein the tumor stromal cell is surrounding the tumor cell.
  • Item 33 The compound for use according to any one of items 1 to 32, wherein the compound is administered to the subject in a therapeutically effective amount for preventing and/or treating the proliferative disease.
  • Item 34 The compound for use according to any one of items 1 to 33, wherein the subject is a mammal, such as a mouse, a rat, a guinea pig, a rabbit, a cat, a dog, a monkey, or a human, preferably a human patient.
  • a mammal such as a mouse, a rat, a guinea pig, a rabbit, a cat, a dog, a monkey, or a human, preferably a human patient.
  • Item 35 The compound for use according to any one of items 1 to 34, wherein the prevention and/or treatment comprises the administration of the compound to the subject, preferably wherein the compound is administered to the subject by oral, intranasal, topical, rectal, bronchial, vaginal or parenteral administration, such as, for example, by intravascular administration, intravenous bolus injection, intravenous infusion, intra-arterial bolus injection, intra-arterial infusion, peri-tissue injection, such as peri-tumoral injection, intra- tissue injection, such as intra-tumoral injection, intra-retinal injection, subretinal injection, subcutaneous injection, direct application, inhalation, or by any clinically/medically accepted method.
  • intravascular administration intravenous bolus injection, intravenous infusion, intra-arterial bolus injection, intra-arterial infusion, peri-tissue injection, such as peri-tumoral injection, intra- tissue injection, such as intra-tumoral injection, intra-
  • Item 36 The compound for use according to any one of items 1 to 35, wherein the compound is administered to the subject as a naked polynucleotide molecule, or in conjunction with a deliveiy reagent.
  • Item 37 The compound for use according to any one of items 1 to 36, wherein the compound is administered to the subject in conjunction with a lipophilic reagent, lipofectin, lipofectamine, cellfectin, polycations, and/or liposomes.
  • Item 38 A combination for use in the prevention and/or treatment of a proliferative disease, comprising at least two agonists and/or mimics according to any one of items 1 to 37 selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2.
  • Item 39 The combination for use according to item 38, comprising:
  • Item 40 The combination for use according to item 38 or 39, wherein the miR-200 is selected from miR-200c, miR-200a, and miR-200b, preferably is miR-200c.
  • Item 41 A pharmaceutical composition for use in the prevention and/or treatment of a proliferative disease, comprising
  • Item 42 A method of preventing and/or treating a proliferative disease in a subject, the method comprising administering to the subject an effective amount of a compound according to any one of items 1 to 37, a combination according to any one of items 38 to 40, or a pharmaceutical composition according to item 41, thereby preventing and/or treating the proliferative disease in the subject.
  • Item 43 The method according to item 42, in which a reduction of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDl, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or AD AM10, is therapeutically beneficial in the subject.
  • an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi,
  • Item 44 A method of reducing the expression of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDl, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10 in a cell of a subject, the method comprising introducing into the cell an effective amount of a compound according to any one of items 1 to 37, a combination according
  • Item 46 The method according to item 44 or 45, comprising the administration of an effective amount of a compound according to any one of items 1 to 37, a combination according to any one of items 38 to 40, or a pharmaceutical composition according to item 41, to the subject.
  • Item 47 Use of an agonist or mimic of at least one micro RNA (miR) selected from miR-200, miR-17, miR-192, miR-29b-i, miR-29b-2, miR-16-1, miR-16-2, miR-92a-i, and miR-92a-2for the manufacture of a medicament for use in the prevention and/or treatment of a proliferative disease.
  • miR micro RNA
  • Item 48 The use according to item 47, in which a reduction of at least one gene for an extracellular matrix protein, such as for SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN, FBNl, MMP2, FSCNl, LAMCl, TGFBl, ETSl, KDR, SERPINHl, TIMP2, NCAMl, FNl, FBLN5, COL4A2, COL4A1, LAMBl, CLASPl, FURIN, TLLl, CTSD, MMP16, COL3A1, NIDl, COL1A1, COL5A2, BMPi, COL10A1, COL15A1, COL4A6, COL4A5, COL2A1, MMP9, COL5A3, COL7A1, COL18A1, CTSB, and/or ADAM10, is therapeutically beneficial.
  • an extracellular matrix protein such as for SPARC, FGF2, DST, PLODi, LOXL
  • Figure 1 shows enriched gene sets of differentially expressed genes and miRNAs in the stromal subgroup. Top 10 gene sets are listed according to the number of regulating miRNAs (left) and the number of enriched target genes (right). The color indicates the category to which the single gene sets were assigned to. a) MiRNAs down-regulated, target genes up-regulated
  • Figure 3 shows the invasion rates of colon cancer cells and fibroblasts with and without miRNA transfection. Shown are bar plots of invasion rates as relative mean fluorescence intensities (MFI) in arbitrary units [A.U.].
  • MFI mean fluorescence intensities
  • Figure 4 shows the conceptional summaiy of the invention.
  • the stromal subgroup of colorectal cancer exhibits high metastatic potential of cancer cells and thus this subgroup has the worst survival prognosis compared to all other subgroups.
  • miR-200c, miR-17, and miR-192 tumor- associated fibroblasts can produce more ECM-related and invasion-promoting factors leading to an increased invasion of cancer cells and thus a highly invasive phenotype. Admission of these three miRNAs to tumor-associated fibroblasts in turn blocks transcription or translation of invasion-promoting proteins leading to a less invasive phenotype.
  • the inventors explored multiple miRNAs and their target gene regulation to define actors in colorectal cancer biology. They focused their investigation on the regulatory role of miRNAs in extracellular matrix remodeling and found that extracellular matrix (ECM)-related genes are more strongly expressed in the stromal rather than epithelial cancer cell component of tumors ( Figure l). The inventors particularly identified the most prominently regulated functional gene sets to be extracellular matrix organization (including miR-200c and miR- 192, associated with 89 target genes) and extracellular matrix disassembly (including miR- 17, associated with 19 target genes) genes.
  • ECM extracellular matrix
  • miR-200c, miR-17 and miR-192 regulate genes crucial for extracellular matrix remodeling, since enforced expression of either miR-200c, miR-17 or miR-192 in untransformed human colon fibroblasts down-regulated 80% of the ECM remodeling target genes.
  • Enforced miR-200c, miR-17 and/or miR-192 expression in colon fibroblasts reduces invasive capacity of co-cultured colorectal cancer cells
  • the inventors set up a Boyden-chamber assay with HCT-116 colon cancer cells in the inner chambers and CCD-18C0 colon fibroblasts in the outer chambers and measured cancer cell invasion as mean fluorescent intensity. Fibroblasts were selectively transfected with mimics of miR-200c, miR-17, and miR-192, a combination of all three mimics or a mock control mimic.
  • the inventors’ data shows that miR-200c, miR-17, and miR-192, expression in fibroblasts impacts the migratory capacity of co-cultured colon tumor cells. Expressing these miRNAs singly or in combination in human colon fibroblasts co-cultured with colon cancer cells considerably reduced cancer cell invasion in Boyden-chamber assays, i.e. the invasive activity of co-cultured colorectal cancer cells. Consequently, the inventors identified miR- 200c, miR-17 and miR-192 as central regulators of extracellular matrix remodeling by tumor- associated fibroblasts, which suppress migratory activity of colon cancer cells. The inventors’ data support a tumor-suppressive function for these three miRNAs in colorectal cancer.
  • miR-200c, miR-17 and miR-192 down-regulate target genes associated with ECM remodeling in fibroblasts, and the down-regulation of these miRNA regulators in stromal subgroup colorectal tumors could account for the heightened invasive or metastatic capacity of cancer cells (Figure 4).
  • the inventors also observed a reduction of fibroblast migration after enforced expression of miR-192 or miR-17.
  • the inventors identified SPARC, FGF2, DST, PLODi, LOXL2, ITGBi, ITGAV, PXDN and FBNi as potential target genes of miR-192, MMP2, FSCNi, LAMCi and TGFBi as potential miR-17 target genes and ETSi, KDR, SERPINHi, TIMP2, NCAMi, FNi and FBLN5 as potential miR-200c target genes.
  • These 3 miRNA and their 20 target genes are very likely involved in ECM remodeling in stromal colorectal tumor, and are highly relevant for cancer cell migration and metastasis.
  • - hsa-miR-29b-i relevant in the following cancers: Colon, Gastric, Melanoma, Breast, Esophagel, HNSCC, Pancreas, Testicular
  • - hsa-miR-29b-2 relevant in the following cancers: Colon, Gastric, Breast, Uveal_Melanoma, Melanoma, Esophagel, HNSCC, Pancreas, Testicular
  • - hsa-miR-16-1 relevant in the following cancers: Bladder, Gastric, Rectum, Lower grade glioma, HNSCC, Colon, Pancreas, Lung squamous cell
  • the aforementioned miRNAs were found down-regulated in samples of stromal-like, cancer-specific subgroups compared to samples of other subgroups.
  • the human colon fibroblast cell line CCD-18C0 was purchased from ATCC. Cells were cultured in DMEM medium with high glucose (1 g/L) and sodium pyruvate (110 mg/L) (Gibco, Carlsbad, CA, USA) supplemented with 10 % FCS superior (Biochrom, Berlin, Germany) without antibiotics at 37°C and 5 % CO2. The cells were genotyped and tested for mycoplasma contamination.
  • the human colon carcinoma cell line HCT-116 was kindly provided by Evi Frei. HCT-116 cells were cultured in McCoy's 5a Medium Modified (Gibco) + 10% FCS superior without antibiotics at 37°C and 5 % CO2. Cells were tested for mycoplasma contamination.
  • Fibroblasts were seeded in 6 well plates and cultured until they reached 80 % confluence. Transfection was performed using Lipofectamine RNAiMAX reagent (Life technologies, CA, USA), which resulted in high transfection efficacy of 80 % and low effect on cell viability. Cells were transfected with 50 nM mimic control-i (ath-miR4i6), miR-192, miR-17 or miR- 200c, respectively. Two days post transfection cells were harvested and cell pellets were shock-frozen and stored at -8o°C until further usage. Experimental validation with qPCR
  • TBP normalization control gene for miR-192 and miR- 17 and UBC as a control gene for miR-200c.
  • ACt-values were calculated by subtracting the Ct-value of the candidate gene from the Ct-value of the house-keeping gene for each replicate. Unpaired Student's t- test was performed on the ACt-values of the miRNA transfection and the mimic-control experiment. As the inventors tested for down-modulation, the t-test parameter "alternative" was set to "greater". Upper and lower error values were calculated as log2 fold expression value +/- log2-transformed AACt standard error.
  • CCD-18C0 cells were seeded in 6 well plates and grown to 80 % confluency until they were transfected with 50 nM miRNA (mock, miR-192, miR-17 or miR-200c) and cultured for 3 days.
  • the invasion assay was performed using 96 well Boyden-chamber plates (8 pm pore size, Corning, Big Flats, NY, USA), which were coated with 10 pg matrigel per well. Fibroblasts were harvested and 3 individually transfected wells were pooled for each condition.
  • 1.5x105 CCD-18C0 cells were seeded in outer wells of the Boyden-chamber in 150 m ⁇ DMEM medium supplemented with 10 % FCS.
  • HCT-116 colon cancer cells were seeded in the inner wells with a cell number of 5x104 in 50 m ⁇ per well. HCT-116 cells were kept in medium without serum. After 24 h incubation at 37°C invasion was measured. Cells were lysed with lx cell dissociation solution (CDS) (Trevigen, Gaithersburg, MD, USA) supplemented with calcein-AM. HTS Transwell 96 well black receiver plates (Corning, Big Flats, NY, USA) were pre-incubated with PBS at 37 °C. Meanwhile, the medium was removed from the outer wells of the Boyden-chamber plates. Washing of outer wells was performed very carefully by pipetting 150 m ⁇ PBS in and out.
  • CDS lx cell dissociation solution
  • the PBS from the black receiver plate was discarded and 100 m ⁇ of lx CDS-calcein was pipetted in each well.
  • the inlay from the transwell plate was transferred carefully to the receiver plate avoiding any contacts.
  • the plates were incubated for 1 h at 37 °C and knocking at each side of the plate was performed every 15 min to ensure complete dissociation of cells attached to the lower side of the gel membrane. Finally, fluorescence was measured using 485 nm as excitation wavelength, 538 nm as the emission wavelength and integration of 60 ms.
  • n.s. not significant
  • hnscc head and neck squamous cell carcinoma
  • lung see lung squamous cell carcinoma
  • lg glioma lower grade glioma
  • HNSCC head and neck squamous cell carcinoma
  • ESCC Oesophageal squamous cell carcinoma
  • AC Oesophageal adenocarcinoma

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Abstract

La présente invention vise à fournir de nouvelles stratégies thérapeutiques permettant de prévenir et de traiter des maladies prolifératives, telles que le cancer colorectal. L'invention repose sur l'idée de cibler des micro-ARN (miARN) qui régulent les protéines de matrice extracellulaire (ECM) impliquées dans la tumorigenèse en tant que stratégie pour le traitement de maladies cancéreuses. La présente invention est plus particulièrement basée sur la modulation de l'expression ou de l'activité de miR-200, miR-17, et miR-192, qui ont été identifiés comme régulant à la baisse des gènes cibles en vue d'un remodelage ECM dans l'environnement tumoral. De manière surprenante, la restauration de l'expression de miR-200, miR-17 et/ou miR-192 dans des cellules du stroma tumoral réduit la capacité invasive des cellules du cancer du côlon. La présente invention concerne donc des composés, qui sont des agonistes ou des mimétiques de miR-200, miR-17, et miR-192, qui sont particulièrement utiles dans la prévention et/ou le traitement de maladies prolifératives, telles que les maladies cancéreuses, de préférence le cancer colorectal.
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WO2024040126A3 (fr) * 2022-08-16 2024-04-18 The Regents Of The University Of California Régulation glycémique améliorée par administration de micro-arn 192

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