WO2019012030A1 - Inhibiteur de dhodh et inhibiteur de chk1 pour le traitement du cancer - Google Patents

Inhibiteur de dhodh et inhibiteur de chk1 pour le traitement du cancer Download PDF

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WO2019012030A1
WO2019012030A1 PCT/EP2018/068903 EP2018068903W WO2019012030A1 WO 2019012030 A1 WO2019012030 A1 WO 2019012030A1 EP 2018068903 W EP2018068903 W EP 2018068903W WO 2019012030 A1 WO2019012030 A1 WO 2019012030A1
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inhibitor
dhodh
chkl
cancer
cells
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PCT/EP2018/068903
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English (en)
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Claude SARDET
Stéphanie ARNOULD
Geneviève RODIER
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INSERM (Institut National de la Santé et de la Recherche Médicale)
Université De Montpellier
Institut Régional Du Cancer De Montpellier
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Publication of WO2019012030A1 publication Critical patent/WO2019012030A1/fr

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    • 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
    • A61K31/275—Nitriles; Isonitriles
    • A61K31/277—Nitriles; Isonitriles having a ring, e.g. verapamil
    • 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
    • A61K31/42—Oxazoles
    • 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
    • A61K31/55—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having seven-membered rings, e.g. azelastine, pentylenetetrazole
    • A61K31/551—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having seven-membered rings, e.g. azelastine, pentylenetetrazole having two nitrogen atoms, e.g. dilazep
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to an inhibitor of DHODH and an inhibitor of Chkl for use in the treatment of a cancer in a subject in need thereof.
  • Dihydroorotate dehydrogenase (EC 1.3.5.2; DHODH) is the one mitochondrial enzyme that is located on the outer surface of the inner membrane and takes part in the fourth and rate-limiting step of de novo pyrimidine biosynthesis [1]. It converts dihydroorotic acid to orotic acid whilst reducing ubiquinone to ubiquinol which makes DHODH a link between pyrimidine synthesis and respiratory electron transport chain.
  • DHODH has emerged as a new therapeutic target in a wide spectrum of pathologies as de novo pyrimidine synthesis is extensively used in rapidly proliferating human or parasitic cells.
  • Much effort has been devoted to designing new inhibitors in order to overcome widespread resistance to current antimalarial drugs [2-5] inasmuch as Plasmodium proliferation relies exclusively on this pathway [6].
  • Series of original compounds were also synthesised as part of a program aiming at identifying new antivirals [7-11] and a new compound is currently in clinical development for the treatment of fungal infection [12].
  • the immunosuppressant leflunomide has been prescribed for the treatment of inflammatory response associated with rheumatoid arthritis [13-16] and the immunomodulatory properties of its active metabolite teriflunomide (TFN) led to its recent approval for the treatment of relapsing remitting multiple sclerosis [17-19]. DHODH inhibition also effectively slowed down cancer cell and tumour growth of diverse tissue origins [20-24].
  • the main characteristic of this effect was the sustained accumulation of teriflunomide-induced DNA damage as cells displayed increased phospho serine 139 H2AX ( ⁇ 2 ⁇ ) levels and concentration-dependent phosphorylation of Chkl on serine 345 upon exposure to the combination as compared with either inhibitor alone. More, the combination of the DHODH inhibitor with the Chkl inhibitor in a significant lower dose allow to minimize the off-target effects of the Chkl inhibitor. Altogether these results suggest that combining DHODH and Chkl inhibitions may be a strategy worth considering as a potential alternative to conventional chemotherapies.
  • the present invention relates to an inhibitor of DHODH and an inhibitor of Chkl for use in the treatment of a cancer in a subject in need thereof.
  • the present invention relates to the combination of an inhibitor of DHODH and an inhibitor of Chkl for use in the treatment of a cancer in a subject in need thereof.
  • the invention relates to i) an inhibitor of DHODH and ii) an inhibitor of Chkl, as a combined preparation for simultaneous, separate or sequential use in the treatment of cancer in a subject in need thereof.
  • the invention relates to an inhibitor of DHODH protein and ii) an inhibitor of Chkl protein as a combined preparation for simultaneous use in the treatment of cancer.
  • DHODH for "dihydroorotate dehydrogenase” denotes an enzyme which catalyzes the fourth enzymatic step, the ubiquinone-mediated oxidation of dihydroorotate to orotate, in de novo pyrimidine biosynthesis.
  • This protein is a mitochondrial protein located on the outer surface of the inner mitochondrial membrane (IMM). Inhibitors of this enzyme are used to treat autoimmune diseases such as rheumatoid arthritis. (Entrez Gene ID number: 1723; mR A sequences references RefSeq: NM 001361.4; protein sequence reference RefSeq: NP 001352.2; Uniprot: Q02127).
  • CHK1 for "replication checkpoint 1" also known as CHEK1 refers to the human gene encoding a DNA replication checkpoint kinase that signals the DNA replication fork stalling and phosphorylates cdc25, an important phosphatase in cell cycle control, particularly for entry into mitosis (Entrez Gene ID number: 1111; mRNA sequences references RefSeq: NM_001114121, NM_001114122.2, NM_001244846.1, NM_001274.5 , NM_001330427.1, NM 001330428.1; protein sequence reference RefSeq: NP 001107593, NP 001107594.1, NP_001231775.1, NP_001265.2, NP_001317356.1, NP_001317357.1).
  • the cancer may be selected in the group consisting of adrenal cortical cancer, anal cancer, bile duct cancer, bladder cancer, bone cancer, brain and central nervous system cancer, breast cancer, Castleman disease, cervical cancer, colorectal cancer, endometrial cancer, esophagus cancer, gallbladder cancer, gastrointestinal carcinoid tumors, Hodgkin's disease, non-Hodgkin's lymphoma, Kaposi's sarcoma, kidney cancer, laryngeal and hypopharyngeal cancer, liver cancer, lung cancer, mesothelioma, plasmacytoma, nasal cavity and paranasal sinus cancer, nasopharyngeal cancer, neuroblastoma, oral cavity and oropharyngeal cancer, ovarian cancer, pancreatic cancer, penile cancer, pituitary cancer, prostate cancer, retinoblastoma, rhabdomyosarcoma, salivary gland cancer, skin cancer, stomach cancer, testicular
  • the cancer is a p53 -deficient cancer.
  • the cancer is a breast cancer. More particularly, triple negative breast cancer and high-grade serous ovarian carcinoma.
  • the term "subject” denotes a mammal, such as a rodent, a feline, a canine, and a primate. Particularly, the subject according to the invention is a human.
  • treatment refers to both prophylactic or preventive treatment as well as curative or disease modifying treatment, including treatment of subjects at risk of contracting the disease or suspected to have contracted the disease as well as subjects who are ill or have been diagnosed as suffering from a disease or medical condition, and includes suppression of clinical relapse.
  • the treatment may be administered to a subject having a medical disorder or who ultimately may acquire the disorder, in order to prevent, cure, delay the onset of, reduce the severity of, or ameliorate one or more symptoms of a disorder or recurring disorder, or in order to prolong the survival of a subject beyond that expected in the absence of such treatment.
  • therapeutic regimen is meant the pattern of treatment of an illness, e.g., the pattern of dosing used during therapy.
  • a therapeutic regimen may include an induction regimen and a maintenance regimen.
  • the phrase “induction regimen” or “induction period” refers to a therapeutic regimen (or the portion of a therapeutic regimen) that is used for the initial treatment of a disease.
  • the general goal of an induction regimen is to provide a high level of drug to a subject during the initial period of a treatment regimen.
  • An induction regimen may employ (in part or in whole) a "loading regimen", which may include administering a greater dose of the drug than a physician would employ during a maintenance regimen, administering a drug more frequently than a physician would administer the drug during a maintenance regimen, or both.
  • maintenance regimen refers to a therapeutic regimen (or the portion of a therapeutic regimen) that is used for the maintenance of a subject during treatment of an illness, e.g., to keep the subject in remission for long periods of time (months or years).
  • a maintenance regimen may employ continuous therapy (e.g., administering a drug at a regular intervals, e.g., weekly, monthly, yearly, etc.) or intermittent therapy (e.g., interrupted treatment, intermittent treatment, treatment at relapse, or treatment upon achievement of a particular predetermined criteria [e.g., disease manifestation, etc.]).
  • DHODH inhibitor or “Chkl inhibitor” denotes molecules or compound which can inhibit the activity of the proteins (e.g. inhibit the kinase, polymerase, reductase activity of the proteins) or a molecule or compound which destabilizes the proteins.
  • an inhibitor of DHODH can inhibit the pyrimidine synthesis and particularly inhibit the reduction of ubiquinone to ubiquinol.
  • an inhibitor of Chkl can inhibit the phosphorylation of cdc25.
  • DHODH inhibitor or “Chkl inhibitor” also denotes inhibitors of the expression of the gene coding for the proteins.
  • the inhibitors according to the invention may be a low molecular weight compound, e. g. a small organic molecule (natural or not).
  • small organic molecule refers to a molecule (natural or not) of a size comparable to those organic molecules generally used in pharmaceuticals.
  • Preferred small organic molecules range in size up to about 10000 Da, more preferably up to 5000 Da, more preferably up to 2000 Da and most preferably up to about 1000 Da.
  • the inhibitor of DHODH according to the invention may be the leflunomide or its active metabolite, the teriflunomide (TFN) of formula (I):
  • the inhibitor of DHODH according to the invention may be the Leflunomide or its active metabolite, the IPP- AO 17- A04 (TFN) of formula (II) :
  • the DHODH inhibitor is an inhibitor described in the articles: Davies M et al 2009; Booker ML et al 2010; Coteron JM et al 2011; Skerlj RT et al 2011 ; Reyes P et al 1982; Bonavia A et al 2011; Hoffmann H-H et al 2011; Munier-Lehmann H et al 2015; Lucas-Hourani M et al 2015; Munier-Lehmann H et al 2013.
  • the DHODH inhibitor is the compounds 2-(3' -ethoxy-3,5- difluorobiphenyl-4-ylamino)nicotinic acid; 2-(3,5-difluoro-3 -methoxybiphenyl-4- ylamino)nicotinic acid; 2-(3' -cyclopropoxy-3,5-difluorobiphenyl-4-ylamino)nicotinic acid; 2-(3,5-difluoro-3 -methoxybiphenyl-4-ylamino)-5-methylnicotinic acid; or a pharmaceutically acceptable salt or N-oxide thereof as described in the patent application WO2009153043.
  • the DHODH inhibitor is the ASLAN003 compound from the company ASLAN Pharma as described in aslanpharma.com/drug/aslan003/.
  • the DHODH inhibitor is a compound as described in the patent application WO2017037292.
  • the inhibitor of Chkl according to the invention may be the PF477736 compound of formula (III):
  • the inhibitor of Chkl according to the invention may be the Chkl inhibitor 7-hydroxystaurosporine as described in D Sampath, et al, 2005.
  • the Chkl inhibitor is an inhibitor described in PrudAppel, Recent Patents on Anti-Cancer Drug Discovery, 2006, 1, 55-68; Expert Opin. Ther. Patents (2011) 21(8): 1191-1210; and Cell Cycle (201 1) 10: 13, 2121-2128, each of which is incorporated herein by reference.
  • the Chkl inhibitor is selected from the group consisting of
  • the DHODH inhibitor is the GDC-0575 compound from the company genentech as described in gene.com/medical-professionals/pipeline/.
  • an inhibitor of the Chkl according to the invention may be a compound as described in the patent application WO2008132500.
  • the inhibitor according to the invention is an antibody.
  • Antibodies or directed against DHODH or Chkl can be raised according to known methods by administering the appropriate antigen or epitope to a host animal selected, e.g., from pigs, cows, horses, rabbits, goats, sheep, and mice, among others.
  • a host animal selected, e.g., from pigs, cows, horses, rabbits, goats, sheep, and mice, among others.
  • Various adjuvants known in the art can be used to enhance antibody production.
  • antibodies useful in practicing the invention can be polyclonal, monoclonal antibodies are preferred.
  • Monoclonal antibodies against DHODH or Chkl can be prepared and isolated using any technique that provides for the production of antibody molecules by continuous cell lines in culture.
  • Techniques for production and isolation include but are not limited to the hybridoma technique originally described by Ko filer and Milstein (1975); the human B-cell hybridoma technique (Cote et al, 1983); and the EBV-hybridoma technique (Cole et al. 1985).
  • techniques described for the production of single chain antibodies can be adapted to produce anti- DHODH or Chkl single chain antibodies.
  • Coumpounds useful in practicing the present invention also include anti- DHODH or Chkl antibody fragments including but not limited to F(ab')2 fragments, which can be generated by pepsin digestion of an intact antibody molecule, and Fab fragments, which can be generated by reducing the disulfide bridges of the F(ab')2 fragments.
  • F(ab')2 fragments which can be generated by pepsin digestion of an intact antibody molecule
  • Fab fragments which can be generated by reducing the disulfide bridges of the F(ab')2 fragments.
  • Fab and/or scFv expression libraries can be constructed to allow rapid identification of fragments having the desired specificity to DHODH or Chkl .
  • Humanized anti- DHODH or anti- Chkl antibodies and antibody fragments therefrom can also be prepared according to known techniques.
  • “Humanized antibodies” are forms of non-human (e.g., rodent) chimeric antibodies that contain minimal sequence derived from non-human immunoglobulin.
  • humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a hypervariable region (CDRs) of the recipient are replaced by residues from a hypervariable region of a non-human species (donor antibody) such as mouse, rat, rabbit or nonhuman primate having the desired specificity, affinity and capacity.
  • donor antibody such as mouse, rat, rabbit or nonhuman primate having the desired specificity, affinity and capacity.
  • framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues.
  • humanized antibodies may comprise residues that are not found in the recipient antibody or in the donor antibody. These modifications are made to further refine antibody performance.
  • the humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin and all or substantially all of the FRs are those of a human immunoglobulin sequence.
  • the humanized antibody optionally also will comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin.
  • Fc immunoglobulin constant region
  • the anti-Chkl antibody according to the invention may be the sc-8408 antibody as send by Santa Cruz biotechnology.
  • the antibody according to the invention is a single domain antibody against DHODH or Chkl .
  • the term "single domain antibody” (sdAb) or "VHH” refers to the single heavy chain variable domain of antibodies of the type that can be found in Camelid mammals which are naturally devoid of light chains. Such VHH are also called “nanobody®”. According to the invention, sdAb can particularly be llama sdAb.
  • VHH refers to the single heavy chain having 3 complementarity determining regions (CDRs): CDR1, CDR2 and CDR3.
  • CDRs complementarity determining regions
  • CDR complementarity determining region
  • VHH according to the invention can readily be prepared by an ordinarily skilled artisan using routine experimentation.
  • VHH variants and modified form thereof may be produced under any known technique in the art such as in- vitro maturation.
  • VHHs or sdAbs are usually generated by PCR cloning of the V-domain repertoire from blood, lymph node, or spleen cDNA obtained from immunized animals into a phage display vector, such as pHEN2.
  • Antigen- specific VHHs are commonly selected by panning phage libraries on immobilized antigen, e.g., antigen coated onto the plastic surface of a test tube, biotinylated antigens immobilized on streptavidin beads, or membrane proteins expressed on the surface of cells.
  • immobilized antigen e.g., antigen coated onto the plastic surface of a test tube, biotinylated antigens immobilized on streptavidin beads, or membrane proteins expressed on the surface of cells.
  • VHHs often show lower affinities for their antigen than VHHs derived from animals that have received several immunizations.
  • VHHs from immune libraries are attributed to the natural selection of variant VHHs during clonal expansion of B-cells in the lymphoid organs of immunized animals.
  • the affinity of VHHs from non-immune libraries can often be improved by mimicking this strategy in vitro, i.e., by site directed mutagenesis of the CDR regions and further rounds of panning on immobilized antigen under conditions of increased stringency (higher temperature, high or low salt concentration, high or low pH, and low antigen concentrations).
  • VHHs derived from camelid are readily expressed in and purified from the E. coli periplasm at much higher levels than the corresponding domains of conventional antibodies.
  • VHHs generally display high solubility and stability and can also be readily produced in yeast, plant, and mammalian cells.
  • the "Hamers patents” describe methods and techniques for generating VHH against any desired target (see for example US 5,800,988; US 5,874, 541 and US 6,015,695).
  • the "Hamers patents” more particularly describe production of VHHs in bacterial hosts such as E. coli (see for example US 6,765,087) and in lower eukaryotic hosts such as moulds (for example Aspergillus or Trichoderma) or in yeast (for example Saccharomyces, Kluyveromyces, Hansenula or Pichia) (see for example US 6,838,254).
  • the compound according to the invention is an aptamer.
  • Aptamers are a class of molecule that represents an alternative to antibodies in term of molecular recognition.
  • Aptamers are oligonucleotide or oligopeptide sequences with the capacity to recognize virtually any class of target molecules with high affinity and specificity.
  • Such ligands may be isolated through Systematic Evolution of Ligands by Exponential enrichment (SELEX) of a random sequence library, as described in Tuerk C. and Gold L., 1990.
  • the random sequence library is obtainable by combinatorial chemical synthesis of DNA. In this library, each member is a linear oligomer, eventually chemically modified, of a unique sequence.
  • Peptide aptamers consists of a conformationally constrained antibody variable region displayed by a platform protein, such as E. coli Thioredoxin A that are selected from combinatorial libraries by two hybrid methods (Colas et al., 1996).
  • the compound according to the invention is a polypeptide.
  • polypeptide is an antagonist of DHODH or Chkl and is capable to prevent the function of DHODH or Chkl .
  • polypeptide can be a mutated DHODH or Chkl protein or a similar protein without the function of DHODH or Chkl .
  • the polypeptide of the invention may be linked to a cell- penetrating peptide" to allow the penetration of the polypeptide in the cell.
  • cell-penetrating peptides are well known in the art and refers to cell permeable sequence or membranous penetrating sequence such as penetratin, TAT mitochondrial penetrating sequence and compounds (Bechara and Sagan, 2013; Jones and Sayers, 2012; Khafagy el and Morishita, 2012; Malhi and Murthy, 2012).
  • polypeptides of the invention may be produced by any suitable means, as will be apparent to those of skill in the art.
  • expression may conveniently be achieved by culturing under appropriate conditions recombinant host cells containing the polypeptide of the invention.
  • the polypeptide is produced by recombinant means, by expression from an encoding nucleic acid molecule.
  • Systems for cloning and expression of a polypeptide in a variety of different host cells are well known.
  • the polypeptide is preferably generated by expression from an encoding nucleic acid in a host cell.
  • Any host cell may be used, depending upon the individual requirements of a particular system. Suitable host cells include bacteria mammalian cells, plant cells, yeast and baculovirus systems. Mammalian cell lines available in the art for expression of a heterologous polypeptide include Chinese hamster ovary cells. HeLa cells, baby hamster kidney cells and many others. Bacteria are also preferred hosts for the production of recombinant protein, due to the ease with which bacteria may be manipulated and grown. A common, preferred bacterial host is E coli.
  • polypeptides used in the therapeutic methods of the present invention may be modified in order to improve their therapeutic efficacy.
  • modification of therapeutic compounds may be used to decrease toxicity, increase circulatory time, or modify biodistribution.
  • the toxicity of potentially important therapeutic compounds can be decreased significantly by combination with a variety of drug carrier vehicles that modify biodistribution.
  • adding dipeptides can improve the penetration of a circulating agent in the eye through the blood retinal barrier by using endogenous transporters.
  • a strategy for improving drug viability is the utilization of water-soluble polymers.
  • Various water-soluble polymers have been shown to modify biodistribution, improve the mode of cellular uptake, change the permeability through physiological barriers; and modify the rate of clearance from the body.
  • water-soluble polymers have been synthesized that contain drug moieties as terminal groups, as part of the backbone, or as pendent groups on the polymer chain.
  • PEG Polyethylene glycol
  • Attachment to various drugs, proteins, and liposomes has been shown to improve residence time and decrease toxicity.
  • PEG can be coupled to active agents through the hydroxyl groups at the ends of the chain and via other chemical methods; however, PEG itself is limited to at most two active agents per molecule.
  • copolymers of PEG and amino acids were explored as novel bio materials which would retain the biocompatibility properties of PEG, but which would have the added advantage of numerous attachment points per molecule (providing greater drug loading), and which could be synthetically designed to suit a variety of applications.
  • PEGylation techniques for the effective modification of drugs.
  • drug delivery polymers that consist of alternating polymers of PEG and tri- functional monomers such as lysine have been used by VectraMed (Plainsboro, N.J.).
  • the PEG chains typically 2000 daltons or less
  • Such copolymers retain the desirable properties of PEG, while providing reactive pendent groups (the carboxylic acid groups of lysine) at strictly controlled and predetermined intervals along the polymer chain.
  • the reactive pendent groups can be used for derivatization, cross-linking, or conjugation with other molecules.
  • These polymers are useful in producing stable, long-circulating pro-drugs by varying the molecular weight of the polymer, the molecular weight of the PEG segments, and the cleavable linkage between the drug and the polymer.
  • the molecular weight of the PEG segments affects the spacing of the drug/linking group complex and the amount of drug per molecular weight of conjugate (smaller PEG segments provides greater drug loading).
  • increasing the overall molecular weight of the block co-polymer conjugate will increase the circulatory half-life of the conjugate. Nevertheless, the conjugate must either be readily degradable or have a molecular weight below the threshold- limiting glomular filtration (e.g., less than 60 kDa).
  • linkers may be used to maintain the therapeutic agent in a pro-drug form until released from the backbone polymer by a specific trigger, typically enzyme activity in the targeted tissue.
  • a specific trigger typically enzyme activity in the targeted tissue.
  • tissue activated drug delivery is particularly useful where delivery to a specific site of biodistribution is required and the therapeutic agent is released at or near the site of pathology.
  • Linking group libraries for use in activated drug delivery are known to those of skill in the art and may be based on enzyme kinetics, prevalence of active enzyme, and cleavage specificity of the selected disease-specific enzymes. Such linkers may be used in modifying the protein or fragment of the protein described herein for therapeutic delivery.
  • the DHODH or Chkl inhibitor according to the invention is an inhibitor of DHODH or Chkl gene expression.
  • Small inhibitory RNAs can also function as inhibitors of DHODH or Chkl expression for use in the present invention.
  • DHODH or Chkl gene expression can be reduced by contacting a subject or cell with a small double stranded RNA (dsRNA), or a vector or construct causing the production of a small double stranded RNA, such that DHODH or CHkl gene expression is specifically inhibited (i.e. RNA interference or RNAi).
  • dsRNA small double stranded RNA
  • RNAi RNA interference
  • Methods for selecting an appropriate dsRNA or dsRNA-encoding vector are well known in the art for genes whose sequence is known (e.g. see for example Tuschl, T. et al. (1999); Elbashir, S. M. et al.
  • Ribozymes can also function as inhibitors of DHODH or CHkl gene expression for use in the present invention.
  • Ribozymes are enzymatic RNA molecules capable of catalyzing the specific cleavage of RNA.
  • the mechanism of ribozyme action involves sequence specific hybridization of the ribozyme molecule to complementary target RNA, followed by endonucleo lytic cleavage.
  • Engineered hairpin or hammerhead motif ribozyme molecules that specifically and efficiently catalyze endonucleo lytic cleavage of DHODH or CHkl mRNA sequences are thereby useful within the scope of the present invention.
  • ribozyme cleavage sites within any potential RNA target are initially identified by scanning the target molecule for ribozyme cleavage sites, which typically include the following sequences, GUA, GUU, and GUC. Once identified, short RNA sequences of between about 15 and 20 ribonucleotides corresponding to the region of the target gene containing the cleavage site can be evaluated for predicted structural features, such as secondary structure, that can render the oligonucleotide sequence unsuitable. The suitability of candidate targets can also be evaluated by testing their accessibility to hybridization with complementary oligonucleotides, using, e.g., ribonuclease protection assays.
  • antisense oligonucleotides and ribozymes useful as inhibitors of DHODH or CHkl gene expression can be prepared by known methods. These include techniques for chemical synthesis such as, e.g., by solid phase phosphoramadite chemical synthesis. Alternatively, anti-sense RNA molecules can be generated by in vitro or in vivo transcription of DNA sequences encoding the RNA molecule. Such DNA sequences can be incorporated into a wide variety of vectors that incorporate suitable RNA polymerase promoters such as the T7 or SP6 polymerase promoters. Various modifications to the oligonucleotides of the invention can be introduced as a means of increasing intracellular stability and half-life.
  • Possible modifications include but are not limited to the addition of flanking sequences of ribonucleotides or deoxyribonucleotides to the 5' and/or 3' ends of the molecule, or the use of phosphorothioate or 2'-0-methyl rather than phosphodiesterase linkages within the oligonucleotide backbone.
  • Antisense oligonucleotides siRNAs and ribozymes of the invention may be delivered in vivo alone or in association with a vector.
  • a "vector" is any vehicle capable of facilitating the transfer of the antisense oligonucleotide siRNA or ribozyme nucleic acid to the cells and preferably cells expressing DHODH or CHkl .
  • the vector transports the nucleic acid to cells with reduced degradation relative to the extent of degradation that would result in the absence of the vector.
  • the vectors useful in the invention include, but are not limited to, plasmids, phagemids, viruses, other vehicles derived from viral or bacterial sources that have been manipulated by the insertion or incorporation of the antisense oligonucleotide siR A or ribozyme nucleic acid sequences.
  • Viral vectors are a preferred type of vector and include, but are not limited to nucleic acid sequences from the following viruses: retrovirus, such as moloney murine leukemia virus, harvey murine sarcoma virus, murine mammary tumor virus, and rouse sarcoma virus; adenovirus, adeno-associated virus; SV40-type viruses; polyoma viruses; Epstein-Barr viruses; papilloma viruses; herpes virus; vaccinia virus; polio virus; and R A virus such as a retrovirus.
  • retrovirus such as moloney murine leukemia virus, harvey murine sarcoma virus, murine mammary tumor virus, and rouse sarcoma virus
  • retrovirus such as moloney murine leukemia virus, harvey murine sarcoma virus, murine mammary tumor virus, and rouse sarcoma virus
  • adenovirus adeno
  • Non-cytopathic viruses include retroviruses (e.g., lentivirus), the life cycle of which involves reverse transcription of genomic viral RNA into DNA with subsequent proviral integration into host cellular DNA. Retroviruses have been approved for human gene therapy trials. Most useful are those retroviruses that are replication-deficient (i.e., capable of directing synthesis of the desired proteins, but incapable of manufacturing an infectious particle). Such genetically altered retroviral expression vectors have general utility for the high-efficiency transduction of genes in vivo.
  • adeno-viruses and adeno-associated viruses are double-stranded DNA viruses that have already been approved for human use in gene therapy.
  • the adeno-associated virus can be engineered to be replication deficient and is capable of infecting a wide range of cell types and species. It further has advantages such as, heat and lipid solvent stability; high transduction frequencies in cells of diverse lineages, including hemopoietic cells; and lack of superinfection inhibition thus allowing multiple series of transductions.
  • the adeno-associated virus can integrate into human cellular DNA in a site-specific manner, thereby minimizing the possibility of insertional mutagenesis and variability of inserted gene expression characteristic of retroviral infection.
  • adeno-associated virus infections have been followed in tissue culture for greater than 100 passages in the absence of selective pressure, implying that the adeno-associated virus genomic integration is a relatively stable event.
  • the adeno- associated virus can also function in an extrachromosomal fashion.
  • Plasmid vectors have been extensively described in the art and are well known to those of skill in the art. See e.g. Sambrook et al, 1989. In the last few years, plasmid vectors have been used as DNA vaccines for delivering antigen-encoding genes to cells in vivo. They are particularly advantageous for this because they do not have the same safety concerns as with many of the viral vectors. These plasmids, however, having a promoter compatible with the host cell, can express a peptide from a gene operatively encoded within the plasmid.
  • Plasmids may be delivered by a variety of parenteral, mucosal and topical routes.
  • the DNA plasmid can be injected by intramuscular, eye, intradermal, subcutaneous, or other routes. It may also be administered by intranasal sprays or drops, rectal suppository and orally.
  • the plasmids may be given in an aqueous solution, dried onto gold particles or in association with another DNA delivery system including but not limited to liposomes, dendrimers, cochleate and microencapsulation.
  • the antisense oligonucleotide, siRNA, shRNA or ribozyme nucleic acid sequence is under the control of a heterologous regulatory region, e.g., a heterologous promoter.
  • the promoter may be specific for Muller glial cells, microglia cells, endothelial cells, pericyte cells and astrocytes
  • a specific expression in Muller glial cells may be obtained through the promoter of the glutamine synthetase gene is suitable.
  • the promoter can also be, e.g., a viral promoter, such as CMV promoter or any synthetic promoters.
  • the invention in another embodiment, relates to a method for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of an inhibitor of DHODH and an inhibitor of Chk 1.
  • a test is necessary.
  • DHODH inhibitors the pyrimidine synthesis or the reduction of ubiquinone to ubiquinol can be tested.
  • the impact of the inhibitors on DHODH enzymatic activity can also be tested using enzymatic assays as described in Knecht and Loffler, 1998; Miller et al, 1968 or in Yin S et al. Sci Rep. 2017 Jan 13;7:40670.
  • Chkl inhibitors the impact of these inhibitors on the phosphorylation of cdc25 and the auto- phosphorylation of Chkl can be tested.
  • Inhibition of Chkl kinase activity can also be tested by an in vitro Kinase assay performed with purified Chkl (immuno-precipitated from Human cells or recombinant protein) and a purified protein substrate of Chkl, in presence of labelled ATP.
  • Another object of the invention relates to a therapeutic composition comprising an inhibitor of DHODH and an inhibitor of Chkl according to the invention for use in the treatment of cancer in a subject in need thereof.
  • Any therapeutic agent of the invention may be combined with pharmaceutically acceptable excipients, and optionally sustained-release matrices, such as biodegradable polymers, to form therapeutic compositions.
  • “Pharmaceutically” or “pharmaceutically acceptable” refers to molecular entities and compositions that do not produce an adverse, allergic or other untoward reaction when administered to a mammal, especially a human, as appropriate.
  • a pharmaceutically acceptable carrier or excipient refers to a non-toxic solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type.
  • compositions for example, the route of administration, the dosage and the regimen naturally depend upon the condition to be treated, the severity of the illness, the age, weight, and sex of the patient, etc.
  • compositions of the invention can be formulated for a topical, oral, intranasal, parenteral, intraocular, intravenous, intramuscular or subcutaneous administration and the like.
  • the pharmaceutical compositions contain vehicles which are pharmaceutically acceptable for a formulation capable of being injected.
  • vehicles which are pharmaceutically acceptable for a formulation capable of being injected.
  • These may be in particular isotonic, sterile, saline solutions (monosodium or disodium phosphate, sodium, potassium, calcium or magnesium chloride and the like or mixtures of such salts), or dry, especially freeze-dried compositions which upon addition, depending on the case, of sterilized water or physiological saline, permit the constitution of injectable solutions.
  • the doses used for the administration can be adapted as a function of various parameters, and in particular as a function of the mode of administration used, of the relevant pathology, or alternatively of the desired duration of treatment.
  • other pharmaceutically acceptable forms include, e.g. tablets or other solids for oral administration; time release capsules; and any other form currently can be used.
  • compositions of the present invention may comprise a further therapeutic active agent.
  • the present invention also relates to a kit comprising an agonist, antagonist or inhibitor of the expression according to the invention and a further therapeutic active agent.
  • anti-cancer agents may be added to the pharmaceutical composition as described below.
  • Anti-cancer agents may be Melphalan, Vincristine (Oncovin), Cyclophosphamide
  • Others anti-cancer agents may be for example cytarabine, anthracyclines, fludarabine, gemcitabine, capecitabine, methotrexate, taxol, taxotere, mercaptopurine, thioguanine, hydroxyurea, cyclophosphamide, ifosfamide, nitrosoureas, platinum complexes such as cisplatin, carboplatin and oxaliplatin, mitomycin, dacarbazine, procarbazine, etoposide, teniposide, campathecins, bleomycin, doxorubicin, idarubicin, daunorubicin, dactinomycin, plicamycin, mitoxantrone, L-asparaginase, doxorubicin, epimbicm, 5-fluorouracil, taxanes such as docetaxel and paclitaxel, leucovorin, levami
  • additional anticancer agents may be selected from, but are not limited to, one or a combination of the following class of agents: alkylating agents, plant alkaloids, DNA topoisomerase inhibitors, anti-folates, pyrimidine analogs, purine analogs, DNA antimetabolites, taxanes, podophyllotoxin, hormonal therapies, retinoids, photosensitizers or photodynamic therapies, angiogenesis inhibitors, antimitotic agents, isoprenylation inhibitors, cell cycle inhibitors, actinomycins, bleomycins, MDR inhibitors and Ca2+ ATPase inhibitors.
  • Additional anti-cancer agents may be selected from, but are not limited to, cytokines, chemokines, growth factors, growth inhibitory factors, hormones, soluble receptors, decoy receptors, monoclonal or polyclonal antibodies, mono-specific, bi-specific or multi-specific antibodies, monobodies, polybodies.
  • Additional anti-cancer agent may be selected from, but are not limited to, growth or hematopoietic factors such as erythropoietin and thrombopoietin, and growth factor mimetics thereof.
  • the further therapeutic active agent can be an antiemetic agent.
  • Suitable antiemetic agents include, but are not limited to, metoclopromide, domperidone, prochlorperazine, promethazine, chlorpromazine, trimethobenzamide, ondansetron, granisetron, hydroxyzine, acethylleucine monoemanolamine, alizapride, azasetron, benzquinamide, bietanautine, bromopride, buclizine, clebopride, cyclizine, dunenhydrinate, diphenidol, dolasetron, meclizme, methallatal, metopimazine, nabilone, oxypemdyl, pipamazine, scopolamine, sulpiride, tetrahydrocannabinols, thiefhylperazine, thioproperazine and tropisetron.
  • the further therapeutic active agent can be an hematopoietic colony stimulating factor.
  • Suitable hematopoietic colony stimulating factors include, but are not limited to, filgrastim, sargramostim, molgramostim and epoietin alpha.
  • the other therapeutic active agent can be an opioid or non-opioid analgesic agent.
  • opioid analgesic agents include, but are not limited to, morphine, heroin, hydromorphone, hydrocodone, oxymorphone, oxycodone, metopon, apomorphine, nomioiphine, etoipbine, buprenorphine, mepeddine, lopermide, anileddine, ethoheptazine, piminidine, betaprodine, diphenoxylate, fentanil, sufentanil, alfentanil, remifentanil, levorphanol, dextromethorphan, phenazodne, pemazocine, cyclazocine, methadone, isomethadone and propoxyphene.
  • Suitable non-opioid analgesic agents include, but are not limited to, aspirin, celecoxib, rofecoxib, diclofinac, diflusinal, etodolac, fenoprofen, flurbiprofen, ibuprofen, ketoprofen, indomethacin, ketorolac, meclofenamate, mefanamic acid, nabumetone, naproxen, piroxicam and sulindac.
  • the further therapeutic active agent can be an anxiolytic agent.
  • Suitable anxiolytic agents include, but are not limited to, buspirone, and benzodiazepines such as diazepam, lorazepam, oxazapam, chlorazepate, clonazepam, chlordiazepoxide and alprazolam.
  • the further therapeutic active agent can be a checkpoint blockade cancer immunotherapy agent.
  • the checkpoint blockade cancer immunotherapy agent is an agent which blocks an immunosuppressive receptor expressed by activated T lymphocytes, such as cytotoxic T lymphocyte-associated protein 4 (CTLA4) and programmed cell death 1 (PDCDl, best known as PD-1), or by NK cells, like various members of the killer cell immunoglobulin- like receptor (KIR) family, or an agent which blocks the principal ligands of these receptors, such as PD-1 ligand CD274 (best known as PD-L1 or B7-H1).
  • CTL4 cytotoxic T lymphocyte-associated protein 4
  • PDCDl programmed cell death 1
  • NK cells like various members of the killer cell immunoglobulin- like receptor (KIR) family, or an agent which blocks the principal ligands of these receptors, such as PD-1 ligand CD274 (best known as PD-L1 or B7-H1).
  • the checkpoint blockade cancer immunotherapy agent is an antibody.
  • the checkpoint blockade cancer immunotherapy agent is an antibody selected from the group consisting of anti-CTLA4 antibodies, anti-PDl antibodies, anti-PDLl antibodies, anti-PDL2 antibodies, anti-TIM-3 antibodies, anti-LAG3 antibodies, anti-IDOl antibodies, anti-TIGIT antibodies, anti-B7H3 antibodies, anti-B7H4 antibodies, anti-BTLA antibodies, and anti-B7H6 antibodies.
  • FIGURES
  • FIG. 1 The combination of teriflunomide and PF477736 results in increased antiproliferative effect in transformed mouse embryonic fibroblasts.
  • Primary, immortalised or transformed mouse embryonic fibroblasts were exposed for 24 hours to increasing concentrations of teriflunomide ⁇ transformed MEF ICIO PF477736 (0.7 ⁇ ) (which was added 30 minutes after the beginning of exposure to TFN) (A) or IPP-A017-A04 ⁇ transformed MEF ICIO PF477736 (B), and grown in drug-free medium for three doubling times.
  • Mean ⁇ SD, n 3 independent experiments.
  • * p ⁇ 0.05, ** p ⁇ 0.01 as determined by two-tailed unpaired t-test.
  • FIG. 1 Pharmacological activity of PF477736 as a single agent or in combination with teriflunomide in triple negative breast cancer cell lines.
  • FIG. 3 The combination of teriflunomide and low dose of PF477736 (ICIO) reduces proliferation of SUM159 and HCC1937 triple negative breast cancer cell lines.
  • Figure 5 The combination of teriflunomide and PF477736 reduces proliferation of MDA-MB-231 triple negative breast cancer cell lines.
  • the active metabolite of leflunomide, teriflunomide (A771726, 2-cyano-3-hydroxy-N- (4-(trifluoromethyl)phenyl)but-2-enamide, see Suppl. Fig. 1), was purchased from Selleck Chemicals LLC (Houston, TX).
  • DHODH inhibitor IPP-A017-A04 (5-cyclopropyl-2-(4-(2,6- difluorophenoxy)-3-isopropoxy-5-methyl-lH-pyrazol-l-yl)-3-fluoropyridine, see Suppl. Fig.
  • Chkl inhibitor PF477736 ((2R)-2-Amino-2-cyclohexyl-N-[2-( 1 -methyl- 1 H-pyrazol-4-yl)-6-oxo-5 ,6-dihydro- 1 H- [l,2]diazepino[4,5,6-cd]indol-8-yl]-acetamide, see Suppl. Fig.l) was purchased from Axon Medchem (Groningen, Netherlands). Drugs were kept at -20°C as 10 mM stock solutions in DMSO. Paclitaxel was supplied by Fresenius-Kabi. Camptothecin, gemcitabine, hydroxyurea, sulforhodamine B, 7 AminoActinomycin D, propidium iodide and RNase A were purchased from Sigma Aldrich.
  • HCC1937 (ATCC-CRL-2336)
  • HCC38 (ATCC-CRL-2314)
  • BT 549 (ATCC HTB-122) triple negative breast cancer cells were obtained from American Type Culture Collection.
  • HCC1937 and HCC38 cells were grown in RPMI supplemented with 10 % fetal bovine serum, 100 ⁇ g/ml streptomycin and 100 units/ml penicillin.
  • BT-549 cells were grown in Dulbecco Modified Eagle Medium supplemented with 10 % fetal bovine serum, 100 ⁇ g/ml streptomycin and 100 units/ml penicillin. All cell types were grown at 37°C in a humidified atmosphere containing 5% C02 and were regularly checked for the absence of mycoplasma contamination.
  • Transformed mouse embryonic fibroblasts were transfected with 700 pmoles of either a mix of 4 target-specific 19-25 nt small interfering RNA (siRNA) for mouse Chkl (Santa Cruz Biotechnology sc 29270), or SASI_Mm01_00087610 and SASI_Mm01_00087610 MISSION® siRNA for mouse DHODH (Sigma Aldrich), or 1400 pmoles of the combination of Chkl and DHODH siRNA.
  • siRNA small interfering RNA
  • SUM159 cells were transfected with 700 pmoles of either SASI_Hs02_00326304 and SASI_Hs02_00326304_AS MISSION® siRNA for Human Kinase Chkl (Sigma Aldrich), or SASI_Hs01_00246561 and SASI HsOl 00246561 AS MISSION® siRNA for human DHODH (Sigma Aldrich) and 1400 pmoles of the combination of Chkl and DHODH siRNA. A 20-25 nt siRNA (Santa Cruz Biotechnology sc- 37007) was used as a control.
  • Exponentially growing cells were exposed to teriflunomide, IPP-A017-A04, PF4777736, 0.1 ⁇ camptothecin or 5 mM hydroxyurea (used as positive controls) or the combination of a DHODH inhibitor and PF477736 for 24 hours then grown in drug-free medium.
  • teriflunomide IPP-A017-A04, PF4777736, 0.1 ⁇ camptothecin or 5 mM hydroxyurea (used as positive controls) or the combination of a DHODH inhibitor and PF477736 for 24 hours then grown in drug-free medium.
  • cells were washed once with ice-cold PBS, trypsinised and counted.
  • One million cells per sample were fixed in ice-cold 70% ethanol and stored at -20°C until analysis.
  • Exponentially growing cells were exposed to teriflunomide, IPP-A017-A04, PF4777736, 0.1 ⁇ camptothecin, 5 mM hydroxyurea or the combination of a DHODH inhibitor and PF477736 for 24 hours then grown in drug-free medium. At each time point, cells were washed with PBS, trypsinised, pooled with floating cells and counted. The annexin V and 7-AAD (or propidium iodide) dual labelling of apoptotic cells was conducted using the annexin V-FLUOS Staining Kit from Roche Applied Science according to the manufacturer's instructions.
  • Exponentially growing cells were exposed to teriflunomide, IPP-A017-A04, PF4777736, 0.1 ⁇ camptothecin, 40 ⁇ gemcitabine, or the combination of a DHODH inhibitor and PF477736 for up to 24 hours.
  • lysis buffer 50 mM Tris HC1 pH 7.4 containing 100 mM NaCl, 50 mM NaF, 40 mM ⁇ -glycerophosphate, 5 mM EDTA, 1% Triton X-100, 1 mM sodium orthovanadate, 100 ⁇ PMSF, 1 ⁇ leupeptin, 1 ⁇ pepstatin A and 1 ⁇ aprotinin
  • Lysates were then centrifuged for 10 min at 13000 rpm and 4°C and supernatant protein concentrations were determined using the bicinchoninic acid assay.
  • mice Twenty-five microgram proteins were resolved in 7.5 % polyacrylamide gels and transferred onto nitrocellulose membranes (Whatman). Membranes were incubated overnight with either mouse monoclonal anti-Chkl (G4, Santa Cruz Biotechnology), rabbit anti- Phospho-Chkl (Ser345 or Ser296, Cell Signalling Technology), mouse monoclonal anti-Chk2 (clone 7, Upstate), mouse monoclonal anti-DHODH (E-8, Santa Cruz Biotechnology) or mouse monoclonal anti-D-actin (clone AC-15, Sigma) antibodies. For cleaved caspase-3 and gamma-H2AX assays, 15 microgram proteins were resolved in 15 % polyacrylamide gels.
  • Membranes were incubated overnight with either anti-phospho-histone (Serl39) H2AX (1/500, Merck Millipore), cleaved caspase-3 (Aspl75) (Cell Signalling Technology) or anti- tubulin (Sigma) rabbit antibodies. Signals were visualised using horseradish-conjugated antibodies and Luminata Chemiluminescent detection substrate (Millipore).
  • Exponentially growing cells were seeded in chamber slides, exposed to teriflunomide, IPP A017-A04, PF4777736, 0.1 ⁇ camptothecin or the combination of a DHODH inhibitor and PF477736 for up to 24 hours.
  • Cells were washed twice with PBS, incubated in 4% paraformaldehyde in PBS for 10 minutes, washed in PBS for 5 minutes, blocked with PBS containing 2% bovine serum albumin and 0.5% triton X-100, and incubated overnight at 4°C with anti-phospho-histone (Serl39) H2AX (1/500, Merck Millipore) antibody.
  • tumour-bearing mice Five tumour-bearing mice were monitored in each group on days 2 and 3 of the protocol for pharmacodynamic assessment of the aforementioned regimens by immunohistochemistry. Tumours were harvested 24 or 48 hours after the beginning of each treatment and were fixed with formalin, all sections were counterstained with hematoxylin and eosin. Samples were also probed for Ki67 and cleaved caspase-3 levels. Quantitative analysis of Ki67 section staining was performed using ImageJ software and caspase-3 cleavage was quantitated using Aperio ImageScope software.
  • TFN DHODH inhibitor teriflunomide
  • Nucleotide depletion is a major cause of replication stress and DNA damage.
  • Cells cope with these insults by triggering a protective signalling pathway that involves Chkl kinase activation by phosphorylation.
  • antimetabolites such as gemcitabine and Chkl inhibitors
  • ⁇ 2 ⁇ staining H2AX phosphorylation on serine 139
  • ATR-dependent phosphorylation of Chkl on serine 345 Gamma H2AX staining and Chkl phosphorylation were indeed reported as pharmacodynamic markers of chemopotentiation and Chkl inhibition in combinations of genotoxic drugs and Chkl inhibitors [37].
  • Chk2 phosphorylation [38] as exemplified here by exposure of transformed MEFs to camptothecin (CPT) used as a positive control (data not shown). Accordingly Chk2 phosphorylation was detected in lysates prepared from cells exposed to DHODH + Chkl inhibitors.
  • annexin V/7-AAD staining was performed in cell populations that were collected 48 hours after the beginning of the time course. Dot plots (data not shown) showed a larger population of annexin V positive/7-AAD positive cells upon exposure to the combination as compared with matched controls or each individual inhibitor. These annexin V/7-AAD profiles were indicative of both apoptosis and necrosis.
  • TNBC triple negative breast cancer
  • TNBC cells were exposed to increasing concentrations of TFN with or without a fixed concentration of PF477736.
  • the chemopotentiation of TFN effect by PF477736 was dependent on the value of the fixed concentration of Chkl inhibitor (data not shown).
  • SUM159 cells were exposed to either 0.1 ⁇ , 0.5 ⁇ or 2.5 ⁇ PF477736 alone or in combination with IC70 TFN (or 40 ⁇ gemcitabine as a positive control).
  • Phospho-Ser296 Chkl levels were reduced in a concentration-dependent manner, with the lowest level observed at 2.5 ⁇ PF477736, which confirmed optimal kinase inhibition at that concentration.
  • a concentration-dependent increase in phospho-Ser345 Chkl levels was concurrently observed when Chkl was inhibited (data not shown).
  • Kirschbaum BJ Mechanism of action for leflunomide in rheumatoid arthritis. Clin Immunol Orlando Fla. 1999; 93: 198-208. doi: 10.1006/clim.1999.4777.
  • Meuth M The molecular basis of mutations induced by deoxyribonucleoside triphosphate pool imbalances in mammalian cells. Exp Cell Res. 1989; 181 : 305-16.

Abstract

La présente invention concerne le domaine de la cancérothérapie. Dans cette étude, les inventeurs ont cherché à savoir si l'effet antiprolifératif d'inhibiteurs de DHODH pouvait être amélioré par association avec l'inhibition de la Chk1 kinase. Ils montrent que l'effet du tériflunomide inhibiteur de DHODH est amplifié lorsque les cellules sont ensuite exposées à un inhibiteur de Chk1, PF477736. Des analyses par cytométrie en flux ont révélé des accumulations notables de cellules aux phases S et G2/M, suivies d'une cytotoxicité accrue qui est caractérisée par une induction dépendante de la caspase 3 de la mort cellulaire. L'association de PF477736 avec le tériflunomide sensibilise considérablement les lignées cellulaires de cancer du sein triple négatif humain SUM159 et HCC1937 à une inhibition de la dihydroorotate déshydrogénase. De plus, l'association de l'inhibiteur de DHODH avec l'inhibiteur de Chk1 à une dose considérablement moindre permet de réduire au minimum les effets hors cible de l'inhibiteur de Chk1. Tous ces résultats suggèrent que l'association d'inhibitions de DHODH et de Chk1 peut être une stratégie à prendre en compte en tant qu'alternative possible aux chimiothérapies classiques. En particulier, la présente invention concerne un inhibiteur de DHODH et un inhibiteur de Chk1 destinés à être utilisés dans le traitement d'un cancer chez un sujet le nécessitant.
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111329865A (zh) * 2020-03-30 2020-06-26 华东理工大学 奋乃静在制备治疗子宫内膜癌的药物中的应用
WO2020198510A1 (fr) * 2019-03-28 2020-10-01 Sierra Oncology, Inc. Procédés de traitement du cancer avec des inhibiteurs de chk1
WO2021262874A1 (fr) * 2020-06-24 2021-12-30 Servier Pharmaceuticals , Llc Utilisation d'un composé inhibiteur de dhodh en polythérapie anticancéreuse
CN116457457A (zh) * 2021-04-28 2023-07-18 山东大学 改善早期胚胎发育的方法
US11717512B2 (en) 2018-02-20 2023-08-08 Servier Pharmaceuticals Llc Methods of use for trisubstituted benzotriazole derivatives

Citations (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4816397A (en) 1983-03-25 1989-03-28 Celltech, Limited Multichain polypeptides or proteins and processes for their production
US4946778A (en) 1987-09-21 1990-08-07 Genex Corporation Single polypeptide chain binding molecules
US5225539A (en) 1986-03-27 1993-07-06 Medical Research Council Recombinant altered antibodies and methods of making altered antibodies
US5800988A (en) 1992-08-21 1998-09-01 Vrije Universiteit Brussel Immunoglobulins devoid of light chains
WO1999032619A1 (fr) 1997-12-23 1999-07-01 The Carnegie Institution Of Washington Inhibition genetique par de l'arn double brin
WO2001036646A1 (fr) 1999-11-19 2001-05-25 Cancer Research Ventures Limited Inhibition d"expression genique a l"aide d"arn bicatenaire
WO2001068836A2 (fr) 2000-03-16 2001-09-20 Genetica, Inc. Procedes et compositions d'interference d'arn
US6573099B2 (en) 1998-03-20 2003-06-03 Benitec Australia, Ltd. Genetic constructs for delaying or repressing the expression of a target gene
US6765087B1 (en) 1992-08-21 2004-07-20 Vrije Universiteit Brussel Immunoglobulins devoid of light chains
US6838254B1 (en) 1993-04-29 2005-01-04 Conopco, Inc. Production of antibodies or (functionalized) fragments thereof derived from heavy chain immunoglobulins of camelidae
WO2008132500A2 (fr) 2007-04-27 2008-11-06 Astrazeneca Ab Procédés de traitement de malignités hématologiques
WO2009153043A1 (fr) 2008-06-20 2009-12-23 Almirall, S.A. Combinaisons renfermant du méthotrexate et des inhibiteurs de dhodh
WO2017037292A1 (fr) 2015-09-04 2017-03-09 Aslan Pharmaceuticals Pte Limited Association d'un inhibiteur de la dhodh et d'un inhibiteur des her destinée à une utilisation dans le traitement du cancer

Patent Citations (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4816397A (en) 1983-03-25 1989-03-28 Celltech, Limited Multichain polypeptides or proteins and processes for their production
US5225539A (en) 1986-03-27 1993-07-06 Medical Research Council Recombinant altered antibodies and methods of making altered antibodies
US4946778A (en) 1987-09-21 1990-08-07 Genex Corporation Single polypeptide chain binding molecules
US6765087B1 (en) 1992-08-21 2004-07-20 Vrije Universiteit Brussel Immunoglobulins devoid of light chains
US5800988A (en) 1992-08-21 1998-09-01 Vrije Universiteit Brussel Immunoglobulins devoid of light chains
US5874541A (en) 1992-08-21 1999-02-23 Vrije Universiteit Immunoglobulins devoid of light chains
US6015695A (en) 1992-08-21 2000-01-18 Vrije Universiteit Brussel Immunoglobulins devoid of light chains
US6838254B1 (en) 1993-04-29 2005-01-04 Conopco, Inc. Production of antibodies or (functionalized) fragments thereof derived from heavy chain immunoglobulins of camelidae
US6506559B1 (en) 1997-12-23 2003-01-14 Carnegie Institute Of Washington Genetic inhibition by double-stranded RNA
WO1999032619A1 (fr) 1997-12-23 1999-07-01 The Carnegie Institution Of Washington Inhibition genetique par de l'arn double brin
US6573099B2 (en) 1998-03-20 2003-06-03 Benitec Australia, Ltd. Genetic constructs for delaying or repressing the expression of a target gene
WO2001036646A1 (fr) 1999-11-19 2001-05-25 Cancer Research Ventures Limited Inhibition d"expression genique a l"aide d"arn bicatenaire
WO2001068836A2 (fr) 2000-03-16 2001-09-20 Genetica, Inc. Procedes et compositions d'interference d'arn
WO2008132500A2 (fr) 2007-04-27 2008-11-06 Astrazeneca Ab Procédés de traitement de malignités hématologiques
WO2009153043A1 (fr) 2008-06-20 2009-12-23 Almirall, S.A. Combinaisons renfermant du méthotrexate et des inhibiteurs de dhodh
WO2017037292A1 (fr) 2015-09-04 2017-03-09 Aslan Pharmaceuticals Pte Limited Association d'un inhibiteur de la dhodh et d'un inhibiteur des her destinée à une utilisation dans le traitement du cancer

Non-Patent Citations (58)

* Cited by examiner, † Cited by third party
Title
ALBIGES L; GOUBAR A; SCOTT V; VICIER C; LEFEBVRE C; ALSAFADI S; COMMO F; SAGHATCHIAN M; LAZAR V; DESSEN P: "Chkl as a new therapeutic target in triple-negative breast cancer", BREAST EDINB SCOTL., vol. 23, 2014, pages 250 - 8
AL-EJEH F; PAJIC M; SHI W; KALIMUTHO M; MIRANDA M; NAGRIAL AM; CHOU A; BIANKIN AV; GRIMMOND SM: "Australian Pancreatic Cancer Genome Initiative, Brown MP, Khanna KK. Gemcitabine and CHK1 inhibition potentiate EGFR-directed radio immunotherapy against pancreatic ductal adenocarcinoma", CLIN CANCER RES OFF J AM ASSOC CANCER RES., vol. 20, 2014, pages 3187 - 97
ANGLANA M; APIOU F; BENSIMON A; DEBATISSE M: "Dynamics of DNA replication in mammalian somatic cells: nucleotide pool modulates origin choice and interorigin spacing", CELL, vol. 114, 2003, pages 385 - 94, XP002460010, DOI: doi:10.1016/S0092-8674(03)00569-5
BAUMANN P; MANDL-WEBER S; VOLKL A; ADAM C; BUMEDER I; ODUNCU F; SCHMIDMAIER R: "Dihydroorotate dehydrogenase inhibitor A771726 (leflunomide) induces apoptosis and diminishes proliferation of multiple myeloma cells", MOL CANCER THER., vol. 8, 2009, pages 366 - 75
BESTER AC; RONIGER M; OREN YS; IM MM; SARNI D; CHAOAT M; BENSIMON A; ZAMIR G; SHEWACH DS; KEREM B: "Nucleotide deficiency promotes genomic instability in early stages of cancer development", CELL, vol. 145, 2011, pages 435 - 46, XP028201066, DOI: doi:10.1016/j.cell.2011.03.044
BLASINA A; HALLIN J; CHEN E; ARANGO ME; KRAYNOV E; REGISTER J; GRANT S; NINKOVIC S; CHEN P; NICHOLS T: "Breaching the DNA damage checkpoint via PF-00477736, a novel small-molecule inhibitor of checkpoint kinase 1", MOL CANCER THER., vol. 7, 2008, pages 2394 - 404, XP055034300, DOI: doi:10.1158/1535-7163.MCT-07-2391
BODDY MN; FURNARI B; MONDESERT O; RUSSELL P: "Replication checkpoint enforced by kinases Cdsl and Chkl", SCIENCE, vol. 280, 1998, pages 909 - 12
BONAVIA A; FRANTI M; PUSATERI KEANEY E; KUHEN K; SEEPERSAUD M; RADETICH B; SHAO J; HONDA A; DEWHURST J; BALABANIS K: "Identification of broad-spectrum antiviral compounds and assessment of the druggability of their target for efficacy against respiratory syncytial virus (RSV", PROC NATL ACAD SCI USA, vol. 108, 2011, pages 6739 - 44
BOOKER ML; BASTOS CM; KRAMER ML; BARKER RH; SKERLJ R; SIDHU AB; DENG X; CELATKA C; CORTESE JF; GUERRERO BRAVO JE: "Novel inhibitors of Plasmodium falciparum dihydroorotate dehydrogenase with anti-malarial activity in the mouse model", J BIOL CHEM., vol. 285, 2010, pages 33054 - 64
BURHANS WC; WEINBERGER M: "DNA replication stress, genome instability and aging", NUCLEIC ACIDS RES., vol. 35, 2007, pages 7545 - 56
CANCER RES., vol. 70, no. 12, 2010, pages 4972
CARR AM; MOUDJOU M; BENTLEY NJ; HAGAN IM: "The chkl pathway is required to prevent mitosis following cell-cycle arrest at ''start.", CURR BIOL CB, vol. 5, 1995, pages 1179 - 90, XP004829744, DOI: doi:10.1016/S0960-9822(95)00234-X
CELL CYCLE, vol. 10, no. 13, 2011, pages 2121 - 2128
CLIN. CANCER RES., vol. 16, no. 7, 2010, pages 2076 - 2084
COOK MR; PINCHOT SN; JASKULA-SZTUL R; LUO J; KUNNIMALAIYAAN M; CHEN H: "Identification of a novel Raf-1 pathway activator that inhibits gastrointestinal carcinoid cell growth", MOL CANCER THER., vol. 9, 2010, pages 429 - 37
COTERON JM; MARCO M; ESQUIVIAS J; DENG X; WHITE KL; WHITE J; KOLTUN M; EL MAZOUNI F; KOKKONDA S; KATNENI K: "Structure-guided lead optimization of triazolopyrimidine-ring substituents identifies potent Plasmodium falciparum dihydroorotate dehydrogenase inhibitors with clinical candidate potential", J MED CHEM., vol. 54, 2011, pages 5540 - 61, XP055191680, DOI: doi:10.1021/jm200592f
DAVIES M; HEIKKILA T; MCCONKEY GA; FISHWICK CWG; PARSONS MR; JOHNSON AP: "Structure-based design, synthesis, and characterization of inhibitors of human and Plasmodium falciparum dihydroorotate dehydrogenases", J MED CHEM., vol. 52, 2009, pages 2683 - 93
DEEPA SAMPATH; JORGE CORTES; ZEEV ESTROV; MIN DU; ZHENG SHI; MICHAEL ANDREEFF; VARSHA GANDHI; WILLIAM PLUNKETT, PHARMACODYNAMICS OF CYTARABINE ALONE AND IN COMBINATION WITH 7-HYDROXYSTAUROSPORINE (UCN-01) IN AML BLASTS IN VITRO AND DURING A CLINICAL TRIAL, vol. 107, 2006, pages 2517 - 2524
DOBBELSTEIN M; SORENSEN CS: "Exploiting replicative stress to treat cancer", NAT REV DRUG DISCOV., vol. 14, 2015, pages 405 - 23
EXPERT OPIN. THER. PATENTS, vol. 21, no. 8, 2011, pages 1191 - 1210
FEIJOO C; HALL-JACKSON C; WU R; JENKINS D; LEITCH J; GILBERT DM; SMYTHE C: "Activation of mammalian Chkl during DNA replication arrest: a role for Chkl in the intra-S phase checkpoint monitoring replication origin firing", J CELL BIOL., vol. 154, 2001, pages 913 - 23
FOX RI; HERRMANN ML; FRANGOU CG; WAHL GM; MORRIS RE; STRAND V; KIRSCHBAUM BJ: "Mechanism of action for leflunomide in rheumatoid arthritis", CLIN IMMUNOL ORLANDO FLA., vol. 93, 1999, pages 198 - 208, XP000985931
GOLDENBERG MM: "Leflunomide, a novel immunomodulator for the treatment of active rheumatoid arthritis", CLIN THER., vol. 21, 1999, pages 1837 - 1852
HERRMANN ML; SCHLEYERBACH R; KIRSCHBAUM BJ: "Leflunomide: an immunomodulatory drug for the treatment of rheumatoid arthritis and other autoimmune diseases", IMMUNOPHARMACOLOGY, vol. 47, 2000, pages 273 - 89, XP002611357, DOI: doi:10.1016/S0162-3109(00)00191-0
HOFFMANN H-H; KUNZ A; SIMON VA; PALESE P; SHAW ML: "Broad-spectrum antiviral that interferes with de novo pyrimidine biosynthesis", PROC NATL ACAD SCI USA, vol. 108, 2011, pages 5777 - 82
JONES ME: "Pyrimidine nucleotide biosynthesis in animals: genes, enzymes, and regulation of UMP biosynthesis", ANNU REV BIOCHEM., vol. 49, 1980, pages 253 - 79
KHUTORNENKO AA; DALINA AA; CHERNYAK BV; CHUMAKOV PM; EVSTAFIEVA AG: "The Role of Dihydroorotate Dehydrogenase in Apoptosis Induction in Response to Inhibition of the Mitochondrial Respiratory Chain Complex III", ACTA NATURAE, vol. 6, 2014, pages 69 - 75
KHUTORNENKO AA; ROUDKO VV; CHERNYAK BV; VARTAPETIAN AB; CHUMAKOV PM; EVSTAFIEVA AG: "Pyrimidine biosynthesis links mitochondrial respiration to the p53 pathway", PROC NATL ACAD SCIU S A, vol. 107, 2010, pages 12828 - 33
KINSELLA AR; SMITH D; PICKARD M: "Resistance to chemotherapeutic antimetabolites: a function of salvage pathway involvement and cellular response to DNA damage", BR J CANCER, vol. 75, 1997, pages 935 - 45
KOH S-B; COURTIN A; BOYCE RJ; BOYLE RG; RICHARDS FM; JODRELL DI: "CHK1 Inhibition Synergizes with Gemcitabine Initially by Destabilizing the DNA Replication Apparatus", CANCER RES., vol. 75, 2015, pages 3583 - 95
KUNZ BA; KOHALMI SE; KUNKEL TA; MATHEWS CK; MCINTOSH EM; REIDY JA.: "International Commission for Protection Against Environmental Mutagens and Carcinogens. Deoxyribonucleoside triphosphate levels: a critical factor in the maintenance of genetic stability", MUTAT RES., vol. 318, 1994, pages 1 - 64
LI EK; TAM L-S; TOMLINSON B: "Leflunomide in the treatment of rheumatoid arthritis", CLIN THER., vol. 26, 2004, pages 447 - 59
LUCAS-HOURANI M; MUNIER-LEHMANN H; EL MAZOUNI F; MALMQUIST NA; HARPON J; COUTANT EP; GUILLOU S; HELYNCK O; NOEL A; SCHERF A: "Original 2-(3-Alkoxy-lH-pyrazol-l-yl)azines Inhibitors of Human Dihydroorotate Dehydrogenase (DHODH", J MED CHEM., vol. 58, 2015, pages 5579 - 98
MA CX; JANETKA JW; PIWNICA-WORMS H: "Death by releasing the breaks: CHK1 inhibitors as cancer therapeutics", TRENDS MOL MED., vol. 17, 2011, pages 88 - 96, XP028361762, DOI: doi:10.1016/j.molmed.2010.10.009
MCNEELY S; CONTI C; SHEIKH T; PATEL H; ZABLUDOFF S; POMMIER Y; SCHWARTZ G; TSE A: "Chkl inhibition after replicative stress activates a double strand break response mediated by ATM and DNA-dependent protein kinase", CELL CYCLE GEORGET TEX., vol. 9, 2010, pages 995 - 1004
MEUTH M: "The molecular basis of mutations induced by deoxyribonucleoside triphosphate pool imbalances in mammalian cells", EXP CELL RES., vol. 181, 1989, pages 305 - 16, XP024859275, DOI: doi:10.1016/0014-4827(89)90090-6
MUNIER-LEHMANN H; LUCAS-HOURANI M; GUILLOU S; HELYNCK O; ZANGHI G; NOEL A; TANGY F; VIDALAIN P-O; JANIN YL: "Original 2-(3-alkoxy-lH-pyrazol-l-yl)pyrimidine derivatives as inhibitors of human dihydroorotate dehydrogenase (DHODH", J MED CHEM., vol. 58, 2015, pages 860 - 77
MUNIER-LEHMANN H; VIDALAIN P-O; TANGY F; JANIN YL: "On dihydroorotate dehydrogenases and their inhibitors and uses", J MED CHEM., vol. 56, 2013, pages 3148 - 67, XP055148746, DOI: doi:10.1021/jm301848w
O'CONNOR P; COMI G; FREEDMAN MS; MILLER AE; KAPPOS L; BOUCHARD J-P; LEBRUN-FRENAY C; MARES J; BENAMOR M; THANGAVELU K: "Long-term safety and efficacy of teriflunomide: Nine-year follow-up of the randomized TEMSO study", NEUROLOGY, vol. 86, 2016, pages 920 - 30
O'CONNOR P; WOLINSKY JS; CONFAVREUX C; COMI G; KAPPOS L; OLSSON TP; BENZERDJEB H; TRUFFINET P; WANG L; MILLER A: "TEMSO Trial Group. Randomized trial of oral teriflunomide for relapsing multiple sclerosis", N ENGL J MED., vol. 365, 2011, pages 1293 - 303
OLIVER JD; SIBLEY GEM; BECKMANN N; DOBB KS; SLATER MJ; MCENTEE L; DU PRE S; LIVERMORE J; BROMLEY MJ; WIEDERHOLD NP: "F901318 represents a novel class of antifungal drug that inhibits dihydroorotate dehydrogenase", PROC NATL ACAD SCI U S A, 2016
PRUDHOMME, RECENT PATENTS ON ANTI-CANCER DRUG DISCOVERY, vol. 1, 2006, pages 55 - 68
RAWLINSON R; MASSEY AJ: "yH2AX and Chkl phosphorylation as predictive pharmacodynamic biomarkers of Chkl inhibitor-chemotherapy combination treatments", BMC CANCER, vol. 14, 2014, pages 483
REYES P; RATHOD PK; SANCHEZ DJ; MREMA JE; RIECKMANN KH; HEIDRICH HG: "Enzymes of purine and pyrimidine metabolism from the human malaria parasite, Plasmodium falciparum", MOL BIOCHEM PARASITOL., vol. 5, 1982, pages 275 - 90, XP023699893, DOI: doi:10.1016/0166-6851(82)90035-4
RODIER G; KIRSH O; BARAIBAR M; HOULES T; LACROIX M; DELPECH H; HATCHI E; ARNOULD S; SEVERAC D; DUBOIS E: "The transcription factor E4F1 coordinates CHKl-dependent checkpoint and mitochondrial functions", CELL REP., vol. 11, 2015, pages 220 - 33
SHIBATA ET AL., CANCER SCI., 2011
SKEHAN P; STORENG R; SCUDIERO D; MONKS A; MCMAHON J; VISTICA D; WARREN JT; BOKESCH H; KENNEY S; BOYD MR: "New colorimetric cytotoxicity assay for anticancer-drug screening", J NATL CANCER INST., vol. 82, 1990, pages 1107 - 12, XP001022779, DOI: doi:10.1093/jnci/82.13.1107
SKERLJ RT; BASTOS CM; BOOKER ML; KRAMER ML; BARKER RH; CELATKA CA; O'SHEA TJ; MUNOZ B; SIDHU AB; CORTESE JF: "Optimization of Potent Inhibitors of P. falciparum Dihydroorotate Dehydrogenase for the Treatment of Malaria", ACS MED CHEM LETT., vol. 2, 2011, pages 708 - 13, XP055149839, DOI: doi:10.1021/ml200143c
STÉPHANIE ARNOULD ET AL: "Checkpoint kinase 1 inhibition sensitises transformed cells to dihydroorotate dehydrogenase inhibition", ONCOTARGET, vol. 8, no. 56, 12 July 2017 (2017-07-12), pages 95206 - 95222, XP055445067, DOI: 10.18632/oncotarget.19199 *
TALLANTYRE E; EVANGELOU N; CONSTANTINESCU CS: "Spotlight on teriflunomide", INT MS J., vol. 15, 2008, pages 62 - 8, XP002556505
WARNKE C; STUVE O; KIESEIER BC: "Teriflunomide for the treatment of multiple sclerosis", CLIN NEUROL NEUROSURG., vol. 115, no. 1, 2013, pages 90 - 94
WEBER G: "Biochemical strategy of cancer cells and the design of chemotherapy: G. H. A. Clowes Memorial Lecture", CANCER RES., vol. 43, 1983, pages 3466 - 92
WHITE RM; CECH J; RATANASIRINTRAWOOT S; LIN CY; RAHL PB; BURKE CJ; LANGDON E; TOMLINSON ML; MOSHER J; KAUFMAN C: "DHODH modulates transcriptional elongation in the neural crest and melanoma", NATURE, vol. 471, 2011, pages 518 - 22, XP002734018, DOI: doi:10.1038/nature09882
XIAO Y; RAMISCAL J; KOWANETZ K; DEL NAGRO C; MALEK S; EVANGELISTA M; BLACKWOOD E; JACKSON PK; O'BRIEN T: "Identification of preferred chemotherapeutics for combining with a CHK1 inhibitor", MOL CANCER THER., vol. 12, 2013, pages 2285 - 95
YIN S ET AL., SCI REP., vol. 7, 13 January 2017 (2017-01-13), pages 40670
ZANNINI L; DELIA D; BUSCEMI G: "CHK2 kinase in the DNA damage response and beyond", J MOL CELL BIOL., vol. 6, 2014, pages 442 - 57
ZHAO H; PIWNICA-WORMS H: "ATR-mediated checkpoint pathways regulate phosphorylation and activation of human Chkl", MOL CELL BIOL., vol. 21, 2001, pages 4129 - 39
ZHU S; YAN X; XIANG Z; DING H-F; CUI H: "Leflunomide reduces proliferation and induces apoptosis in neuroblastoma cells in vitro and in vivo", PLOS ONE, vol. 8, 2013, pages e71555, XP055467963, DOI: doi:10.1371/journal.pone.0071555

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