WO2001083693A2 - Composes ayant une affinite pour le recepteur 2 du facteur de croissance de l'endothelium vasculaire (vegfr-2) et utilisations associees - Google Patents

Composes ayant une affinite pour le recepteur 2 du facteur de croissance de l'endothelium vasculaire (vegfr-2) et utilisations associees Download PDF

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WO2001083693A2
WO2001083693A2 PCT/US2001/013598 US0113598W WO0183693A2 WO 2001083693 A2 WO2001083693 A2 WO 2001083693A2 US 0113598 W US0113598 W US 0113598W WO 0183693 A2 WO0183693 A2 WO 0183693A2
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seq
compound
peptide
amino acids
vegfr
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PCT/US2001/013598
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WO2001083693A3 (fr
WO2001083693A8 (fr
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Peter Joseph Schatz
Min-Jia Chen
Sunila Piplani
Cecilia A. Mozsgai
Palani Balu
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Glaxo Group Limited
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Publication of WO2001083693A8 publication Critical patent/WO2001083693A8/fr
Publication of WO2001083693A3 publication Critical patent/WO2001083693A3/fr

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/59—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
    • A61K47/60—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
    • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
    • A61K51/04—Organic compounds
    • A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
    • A61K51/088—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins conjugates with carriers being peptides, polyamino acids or proteins
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/52—Cytokines; Lymphokines; Interferons
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates generally to novel compounds that have affinity for the vascular endothelial cell growth factor receptor-2 (VEGFR-2). More particularly, the invention relates to such compounds which act as modulators of vascular endothelial cell growth factor (VEGF) function.
  • the invention additionally relates to methods of using the novel compounds and pharmaceutical compositions containing a compound of the invention as the active agent.
  • the invention has application in the fields of biochemistry and medicinal chemistry and particularly provides inhibitors of VEGF function for use in the treatment of human disease.
  • Blood vessels serve an important role in the circulatory system of humans and many animals.
  • the internal surface of a blood vessel is comprised of endothelial cells. These cells are well-suited for imparting a smooth and "non-adhesive" or “non-tacky” quality to the internal surfaces of blood vessels.
  • This smooth and "non-adhesive" internal surface is critical for the free flow and transport of blood and related materials through the entire circulatory system. Without such a smooth internal surface, blood vessels would become obstructed as thrombi or other blockages form at "sticky" locations on the internal walls. Such obstructions can result in partial or even complete blockage of the blood flow necessary to maintain normal tissue and organ functioning.
  • endothelial cells serve not only an important structural component of blood vessels, but also provide blood vessels with a smooth and non-tacky internal surface. Blood vessel formation takes place in response to stimuli in the form of specialized growth factors. These growth factors induce mitosis in cells already present in blood vessels. The newly formed cells replace damaged cells or extend the length of the blood vessel. This process of growing blood vessels from preexisting endothelial cells is termed "angiogenesis.” As its name suggests, vascular endothelial cell growth factor (VEGF) is involved in endothelial cell proliferation.
  • VEGF vascular endothelial cell growth factor
  • VEGF is also referred to as "vascular permeability factor” or “vasculotropin.”
  • VEGF is a glycosylated dimer having an overall apparent molecular mass of about 46 kDa (the apparent molecular mass of each subunit is equal to 23 kDa). This important protein selectively induces mitosis of vascular endothelial cells resulting in endothelial cell proliferation and consequently, angiogenesis. Since blood vessels comprise, in part, endothelial cells, VEGF is required for the formation and maintenance of blood vessels.
  • VEGF acts on the extracellular portion of VEGFR-2 expressed on vascular endothelial cells. Thus, once VEGF binds to VEGFR-2, a signal is sent to the nucleus of the endothelial cell instructing it to divide.
  • angiogenesis is deleterious to an organism's overall health.
  • continuous angiogenesis can cause or exacerbate diseases such as psoriasis, rheumatoid arthritis and retinopathy.
  • angiogenesis makes possible continued tumor growth by vascularizing the tumor, thereby supplying the tumor, with blood and nutrients necessary to sustain the tumor's growth.
  • preventing or at least reducing angiogenesis is desirable.
  • One attempt at limiting angiogenesis is described in U.S. Patent No. 5,952,199.
  • the invention provides compounds comprising a peptide chain that binds to VEGFR-2.
  • the peptide chain is approximately 15 to 40 amino acids in length and contains a sequence of amino acids of formula (I) (I) X 1 CX 2 X 3 X 4 X 5 X 6 GX 7 X 8 X 9 CX 10 X 11 X 12 (SEQ ID NO: 1) wherein each amino acid is indicated by standard one-letter abbreviation, and wherein X, is S, T, Y, A, Q, V or E; X 2 is W, R, Y, G, Q or F; X 3 is P, L, T, S, A or E; X 4 is G, S, A or N; X 5 is E, D or A; X 6 is Y, F, N or A; X 7 is G or V; X 8 is V, E, F, L or M; X 9 is E, D or V; X 10 is W, Y or F; X
  • the peptide chain is approximately 8 to 40 amino acids in length and contains a sequence of amino acids of formula (II)
  • the peptide chain is approximately 8 to 40 amino acids in length and contains a sequence of amino acids of formula (III)
  • the compound comprises a single peptide chain. In other contexts, it is preferred that the compound is in the form of a dimer, i.e. a compound comprised of two peptide chains that may or may not be identical.
  • the compounds of the invention include a peptide having its N-terminus coupled to a polyethylene glycol molecule.
  • Peptides having an acetylated N-terminus and/or an amidated C-terminus are also preferred.
  • a pharmaceutical composition that comprises a therapeutically effective amount of a compound of the invention in combination with a pharmaceutically acceptable carrier.
  • a method is provided for treating a patient who would benefit from administration of a VEGFR-2 antagonist, the method comprising administering to the patient a therapeutically effective amount of an antagonist compound of the present invention.
  • a method for imaging and assessing neovascularization during angiogenesis comprising the steps of administering to a patient compound of the invention to a patient wherein the compound is coupled to a detectable label to form a labeled compound, allowing the labeled compound to bind to VEGFR-2, and detecting the detectable label.
  • Amino acid residues in peptides are abbreviated as follows: Phenylalanine is Phe or F; Leucine is Leu or L; Isoleucine is He or I; Methionine is Met or M; Valine is Val or V; Serine is Ser or S; Proline is Pro or P; Threonine is Thr or T; Alanine is Ala or A; Tyrosine is Tyr or Y; Histidine is His or H; Glutamine is Gin or Q; Asparagine is Asn or N; Lysine is Lys or K; Aspartic Acid is Asp or D; Glutamic Acid is Glu or E; Cysteine is Cys or C; Tryptophan is Trp or W; Arginine is Arg or R; and Glycine is Gly or G.
  • “1-Nal” is used to refer to 1-naphthylalanine
  • the "2-Nal” is used to refer to 2-naphthylalanine.
  • Stereoisomers e.g., D-amino acids of the twenty conventional amino acids, unnatural amino acids such as ⁇ , ⁇ -disubstituted amino acids, N-alkyl amino acids, lactic acid, and other unconventional amino acids may also be suitable components for compounds of the present invention.
  • unconventional amino acids include: ⁇ -alanine, 1-naphthylalanine, 2-naphthylalanine, 3-pyridylalanine, 4-hydroxyproline, O-phosphoserine, N-acetylserine, N-formylmethionine, 3-methylhistidine, 5-hydroxylysine, nor-leucine, and other similar amino acids and imino acids (e.g., 4-hydroxyproline) .
  • Peptide or “polypeptide” refers to a polymer in which the monomers are alpha amino acids joined together through amide bonds. Peptides are two or often more amino acid monomers long. One or more of the peptide chains disclosed herein may appear in the compounds of the present. It is also contemplated that the peptide chains disclosed herein represent only a portion of the overall peptide included in the compound.
  • dimer as in a peptide “dimer” refers to a compound in which two peptide chains are linked; generally, although not necessarily, the two peptide chains will be identical and are linked through a linking moiety covalently bound to the carboxyl terminus of each chain.
  • agonist is used herein to refer to a ligand that binds to a receptor and activates the receptor.
  • antagonist is used herein to refer to a ligand that binds to a receptor without activating the receptor. Antagonists are either competitive antagonists or noncompetitive antagonists. A “competitive antagonist” blocks the receptor site that is specific for the agonist. A “noncompetitive antagonist” inactivates the functioning of the receptor by interacting with a site other than the agonist binding site.
  • modulator refers to a compound that is either an agonist or an antagonist of VEGFR-2. "Pharmaceutically or therapeutically effective dose or amount” refers to a dose sufficient to induce a desired biological result.
  • the dose or amount is sufficient to inhibit or reduce the effects of VEGF and, thus, alleviate the symptoms associated with an undesired proliferation of endothelial cells in vivo.
  • treat as in “treat a disease” is intended to include any means of treating a disease in a mammal, including (1) preventing the disease, i.e., avoiding any clinical symptoms of the disease, (2) inhibiting the disease, that is, arresting the development or progression of clinical symptoms, and/or (3) relieving the disease, i.e., causing regression of clinical symptoms.
  • “Optional” or “optionally” means that the subsequently described circumstance may or may not occur, so that the description includes instances where the circumstance occurs and instances where it does not.
  • pharmaceutically acceptable carrier is meant a material which is not biologically or otherwise undesirable, i.e., the material may be administered to an individual along with the selected active agent without causing any undesirable biological effects or interacting in a deleterious manner with any of the other components of the pharmaceutical composition in which it is contained.
  • the invention provides compounds comprising a peptide chain that binds to VEGFR-2.
  • a compound comprising a peptide chain approximately 15 to 40 amino acids in length that binds to VEGFR-2 and contains a sequence of amino acids of formula (I)
  • X ! is S; X 2 is W; X 4 is G; X 5 is E; X 6 is Y; X 7 is G; X 8 is V; X 9 is E X ]0 is W; and X n is S.
  • Examples of particularly preferred sequences satisfying formula (I) include, but are not limited to, the following:
  • TCWSGEYGGVECWAR SEQ ID NO: 6
  • SCWPGDFGGVECWSL SEQ ID NO: 7
  • ACWTGEYGGEECWEL (SEQ ID NO: 10);
  • TCWPGEYGGVECWAV (SEQ ID NO: 11); TCWPGEYGGVECWGR (SEQ ID NO: 12);
  • TCWPGEYGGVECWSL (SEQ ID NO: 14);
  • ACWPSEYGGVECWSL (SEQ ID NO: 16); SCWPGEFGGVECWSV (SEQ ID NO: 17);
  • TCWPGEYGGEECWSL (SEQ ID NO: 19);
  • TCWPGEYGGVECYSV SEQ ID NO: 26
  • SCWPGEYGGEECWSL SEQ ID NO: 27
  • SC WPGEYGGVECYNV SEQ ID NO: 28
  • SCWPGEYGGVECWKL SEQ ID NO: 29
  • TCWPGEYGGEVCWAL (SEQ ID NO: 31);
  • SCWPGEYGGVDCWSV (SEQ ID NO: 32); SCWPGEYGGVECWTL (SEQ ID NO: 33);
  • SCWPGEYGGEVCWSL (SEQ ID NO: 37); VCWPGEYGGVECWNV (SEQ ID NO: 38);
  • VCWPGEYGGEDCWSL (SEQ ID NO: 40);
  • SCWEGEYGGVECWSI (SEQ ID NO: 42); TC(l-Nal)PGEYGGVECYSV (SEQ ID NO: 43);
  • ACWPGEYGGVECYSV (SEQ ID NO: 46);
  • TCAPGEYGGVECYSV SEQ ID NO: 47
  • TCWPGAYGGVECYSV SEQ ID NO: 48
  • TCWPGEAGGVECYSV (SEQ ID NO: 49);
  • TCWPGEYGGVACYSV (SEQ ID NO: 50);
  • TCWPGEYGGVECYAV (SEQ ID NO: 52).
  • ARVVSCWPGEYGGVECWSLNS (SEQ ID NO: 54);
  • PGVVSCWLGEYGGEECWSHNY (SEQ ID NO: 55); FPLVSCWPGEYGGVECWSLKT (SEQ ID NO: 56);
  • TGVVTCWSGEYGGVECWARNT SEQ ID NO: 57
  • HRVDSCWPGDFGGVECWSLSE SEQ ID NO: 58
  • VRVEYCWPGEYGGVDCWSPKP (SEQ ID NO: 59);
  • NRVEACWTGEYGGEECWELNL (SEQ ID NO: 61); AGQVTCWPGEYGGVECWAVKY (SEQ ID NO: 62);
  • VTCWPGEYGGVECWGRKG (SEQ ID NO: 63);
  • NRVESCWEGDNGGVECWWLKY (SEQ ID NO: 66); SGDVACWPSEYGGVECWSLHY (SEQ ID NO: 67);
  • HWEGSCWPGEFGGVECWSVNY (SEQ ID NO: 68);
  • LDVVSCWPGEFGGFDCWSMKY (SEQ ID NO: 71); NRVLVCRPGEYGGEECWSLDY (SEQ ID NO: 72);
  • GGVVSCWPAEYGGVECWSMDY (SEQ ID NO: 73);
  • PRVVTCWPGEYGGEECWSIKY (SEQ ID NO: 76); ERVTTCWPGEYGGVECYSVKY (SEQ ID NO: 77);
  • EWVVSCWPGEYGGEECWSLKY (SEQ ID NO: 78);
  • KRVVSCWPGEYGGVECYNVKY (SEQ ID NO: 79);
  • TQVESCWPGEYGGVECWKLRY (SEQ ID NO: 80);
  • DGVVSCWPSEYGGEDCFNLHY (SEQ ID NO: 81); RPSETCWPGEYGGEVCWALKY (SEQ ID NO: 82);
  • PRVVSCWPGEYGGVDCWSVKY SEQ ID NO: 84
  • MRVEYCWPAEYGGVECFSPRD (SEQ ID NO: 86); SRVLACGPGEYGGEECWFVQY (SEQ ID NO: 87);
  • ETVESCWTGEYGGVECWTLNY (SEQ ID NO: 88); VLEVSCWPGEYGGEVCWSLKM (SEQ ID NO: 89);
  • RVTTCWPGEYGGVECYSVKY (SEQ ID NO: 95);
  • VTTCWPGEYGGVECYSVKY (SEQ ID NO: 96);
  • TCWPGEYGGVECYSVKY (SEQ ID NO: 97); TCWPGEYGGVECYSVKY (SEQ ID NO: 98);
  • RVTTCWPGEYGGVECYSVK (SEQ ID NO: 99);
  • RVTTCWPGEYGGVECYSV (SEQ ID NO: 100);
  • RVTTCWPGEYGGVECYSVK(alloc)Y (SEQ ID NO: 102); ERVTTCWPGEYGGVEC YSVRY (SEQ ID NO: 103);
  • RVTTCWPGEYGGVECYSVRY (SEQ ID NO: 104);
  • RVTTCWPGEYGGVECYSVAY (SEQ ID NO: 106);
  • RVTTC(l-Nal)PGEYGGVECYSVKY (SEQ ID NO: 107); RVTTC(2-Nal)PGEYGGVECYSVKY (SEQ ID NO: 108);
  • RVTTCFPGEYGGVECYSVAY (SEQ ID NO: 109);
  • HVTTCWPGEYGGVECYSVRY (SEQ ID NO: 110);
  • ERVTTC(l-Nal)PGEYGGVECYSVAY SEQ ID NO: 114
  • RVATCWPGEYGGVECYSVAY (SEQ ID NO: 115);
  • RVTACWPGEYGGVECYSVAY (SEQ ID NO: 116);
  • RVTTCAPGEYGGVECYSVAY SEQ ID NO: 117
  • RVTTCWPGAYGGVECYSVAY SEQ ID NO: 118
  • RVTTCWPGEAGGVECYSVAY (SEQ ID NO: 119); RVTTCWPGEYGGVACYSVAY(SEQ IDNO: 120);
  • RVTTCWPGEYGGVECASVAY (SEQ IDNO: 121);
  • RVTTCWPGEYGGVECYAVAY (SEQ IDNO: 122).
  • RVTTCWPGEYGGVECYSVRA (SEQ IDNO: 123).
  • compounds comprising a peptide chain approximately 8 to 40 amino acids in length that binds to VEGFR-2 and contains a sequence of amino acids of formula (II) (II) CX , 1 GX , 2 X , 3 X , 4 CW (SEQ ID NO: 2) wherein each amino acid is indicated by standard one-letter abbreviation, and wherein X' j is W or G; X' 2 is P or A; X' 3 is E or D; X* 4 is G or R.
  • X' j is W; X 2 is P; X' 3 is E; and X' 4 is G.
  • Examples of particularly preferred sequences satisfying Formula (II) include, but are not limited to, the following:
  • CWGPDGCW (SEQ ID NO: 125);
  • YNTVENCWGPDGCWLD (SEQ ID NO: 129);
  • MSLVDKCWGPEGCWLE (SEQ ID NO: 130); TITVGSCWGPEGCWLDSRY (SEQ ID NO: 131);
  • IFLGENCWGPDGCWLE SEQ ID NO: 132
  • IREGDMCWGPEGCWVD (SEQ ID NO: 133);
  • LTLVDNCWGPDGCWLE (SEQ ID NO: 134);
  • ESRVDDCWGPDGCWLDPIT (SEQ ID NO: 135); VEYVANCGGAERCWLGTNM (SEQ ID NO: 136);
  • QDCWGPEGCWLQEQG (SEQ ID NO: 137); VLDADNCWGPEGCWLE (SEQ ID NO: 138); MSEVEDCWGPEGCWLE (SEQ ID NO: 139); NCWGPEGCWLE (SEQ ID NO: 140); SHRVDDCWGPDGCWLE (SEQ ID NO: 141); IIEVGNCWGPEGCWLE (SEQ ID NO: 142); and
  • VDNCWGPEGCWLE (SEQ ID NO: 143).
  • the invention provides compounds comprising a peptide chain approximately 8 to 40 amino acids in length that binds to VEGFR-2 and contains a sequence of amino acids of formula (III)
  • GWLGVGVH (SEQ ID NO: 144);
  • GWLGAGEHN (SEQ IDNO: 145); GWLGVGEH (SEQ IDNO: 146);
  • GWLGLGEH (SEQ IDNO: 147);
  • GWRGIGEH (SEQ ID NO: 149).
  • NASRISSGWLGVGVHNLSA (SEQ ID NO: 150);
  • VIGRTWSGWLGVGVHNLSN (SEQ ID NO: 152);
  • QCGRVSSGWLGVGVHNLPF (SEQ ID NO: 153); TNQRRSSGWLGVGVHTLSP (SEQ ID NO: 154);
  • GHKRASSGWLGVGVHKLSH (SEQ ID NO: 155); HRRRMSSGWLGVGEHKLP (SEQ ID NO: 156); SSGWLGVGVHYLSD (SEQ ID NO: 157); QRRRSSSGWLGLGEHRL (SEQ ID NO: 158); PQRRSSSGWIGLGVHDLFN (SEQ ID NO: 159); SSRRASSGWRGIGEHNLYN (SEQ ID NO: 160); and
  • RCSSGWLGVGVHNLS (SEQ ID NO: 161).
  • Standard solid phase peptide synthesis techniques are preferred for synthesis of the peptides of the present invention. Such techniques are described, for example, by Merrifield (1963) J Am. Chem. Soc. 85:2149. As is well known in the art, solid phase synthesis using the Merrifield method involves successive coupling of Ct-amino protected amino acids to a growing support-bound peptide chain. After the initial coupling of a protected amino acid to a resin support (e.g., a polystyrene resin, a chloromethylated resin, a hydroxymethyl resin, a benzhydrylamine resin, or the like, depending on the chemistry used), the ⁇ -amino protecting group is removed by a choice of reagents, depending on the specific protecting group.
  • a resin support e.g., a polystyrene resin, a chloromethylated resin, a hydroxymethyl resin, a benzhydrylamine resin, or the like, depending on the chemistry used
  • Suitable CC-amino protecting groups are those known to be useful in the art of stepwise synthesis of peptides. Included are acyl type protecting groups (e.g., formyl, trifluoroacetyl, acetyl), aromatic urethane type protecting groups (e.g., benzyloxycarbonyl (Cbz) and substituted Cbz), aliphatic urethane protecting groups (e.g., t-butyloxycarbonyl (Boc), isopropyloxycarbonyl, cyclohexyloxycarbonyl), alkyl type protecting groups (e.g., benzyl, triphenylmethyl), fluorenylmethyl oxycarbonyl (Fmoc), Alloc and Dde.
  • acyl type protecting groups e.g., formyl, trifluoroacetyl, acetyl
  • aromatic urethane type protecting groups e.g., benzyloxycarbonyl (
  • the side chain protecting groups (typically ethers, esters, trityl, and the like) remain intact during coupling; however, the side chain protecting group must be removable upon completion of the synthesis of the final peptide.
  • Preferred side chain protecting groups will depend on the particular amino acid that is being protected as well as the overall chemistry used. After removal of the (X-amino protecting group, the remaining protected amino acids are coupled stepwise in the desired order. Each protected amino acid is generally reacted in about a 3 -fold excess using an appropriate carboxyl group activator such as
  • HBTU 2-(lH-benzotriazol-l-yl)-l,l,3,3 tetramethyluronium hexafluorophosphate
  • DCC dicyclohexylcarbodiimide
  • the compound is cleaved from the solid support by treatment with a reagent such as trifluoroacetic acid, preferably in combination with a scavenger such as ethanedithiol, ⁇ -mercaptoethanol or thioanisole.
  • a reagent such as trifluoroacetic acid
  • a scavenger such as ethanedithiol, ⁇ -mercaptoethanol or thioanisole.
  • naphthylalanine can be substituted for tiyptophan, with 1 -naphthylalanine (1 -Nal) or 2-naphthylalanine (2-Nal).
  • Other synthetic amino acids that can be substituted into the peptides of the present invention include, but are not limited to, pyroglutamic acid (pyr), nor-leucine and 3-pyridylalanine.
  • the compounds of the present invention may possess agonist or antagonist activity.
  • the compounds exhibit antagonist activity toward VEGFR-2 when they are in the form of a single peptide chain.
  • the compounds of the invention exhibit agonist activity toward VEGFR-2 when they are in the form of a dimer, i.e., a compound comprised of two peptide chains that may or may not be identical.
  • the compounds ofthe present invention may be in the form of a dimer. As indicated above, the dimer forms ofthe present invention possess agonist activity toward VEGFR-2. Preferably, the dimer compounds ofthe invention have the structure of formula (IV)
  • R 1 , R 2 , nl, n2, n3, n4, x, y and L ⁇ are defined as follows.
  • R 1 is a peptide chain that binds to VEGFR-2 and contains a sequence of amino acids of formula (I), (II) or (III) as defined above.
  • R 2 is also a peptide chain that binds to VEGFR-2 and contains a sequence of amino acids of formula (I), (II), or (III).
  • R 1 and R 2 can be the same or different. It is preferred, however, that R 1 and R 2 are the same.
  • ⁇ A is a ⁇ -alanine residue and may or may not be present, meaning that nl, n2, n3 and n4 are independently zero or 1.
  • Lk is a terminal linking moiety.
  • Each dimer contains only one linking moiety, meaning that one of x and y is zero and the other is 1.
  • the terminal linking moiety Lk can be any moiety recognized by those skilled in the art as suitable for joining the peptides of R 1 and R 2 .
  • Lk is preferably although not necessarily selected from the group consisting of a disulfide bond, a carbonyl moiety and a C,_i 2 linking moiety optionally terminated with one or two -NH- linkages and optionally substituted at one or more available carbon atoms with a lower alkyl substituent.
  • the linking moiety comprises -NH-R 3 -NH- wherein R 3 is lower (C,_ 6 ) alkylene substituted with a functional group such as a carboxyl group or an amino group that enables binding to another molecular moiety (e.g., as may be present on the surface of a solid support), and is optionally substituted with a lower alkyl group.
  • R 3 is lower (C,_ 6 ) alkylene substituted with a functional group such as a carboxyl group or an amino group that enables binding to another molecular moiety (e.g., as may be present on the surface of a solid support), and is optionally substituted with a lower alkyl group.
  • the linking moiety is a lysine residue or lysine amide, i.e., a lysine residue wherein the carboxyl group has been converted to an amide moiety -CONH 2 .
  • the pair form a disulfide bond linking these residues.
  • the disulfide bond may be present within a single peptide chain forming an intramolecular disulfide bond.
  • the disulfide bond may connect the two chains.
  • more than one disulfide bond may be present.
  • Amino terminal modifications include methylation (i.e., conversion of a free amino group to an -NHCH 3 or -N(CH 3 ) 2 moiety), acetylation (with either acetic acid per se, or with a halogenated derivative thereof such as ⁇ -chloroacetic acid, cc-bromoacetic acid, or cc-iodoacetic acid), addition of a benzyloxycarbonyl group, or blocking with a blocking group containing a carboxylate functionality RCOO- or a sulfonyl functionality R-SO 2 -, where R is selected from the group consisting of alkyl, aryl, heteroaryl, alkyl aryl, and the like, and similar groups.
  • amino terminal modifications herein include conjugation to a polyethylene glycol molecule ("PEGylation") and acetylation.
  • PEGylation polyethylene glycol molecule
  • acetylation it is also preferred that the peptide chains contain no amino groups other than at the N-terminus (and thus no lysine residues), so that N-terminal-specific modification may be carried out.
  • the peptides and compounds ofthe invention can advantageously be modified with or covalently coupled to one or more of a variety of hydrophilic polymers. It has been found that when the peptide compounds are derivatized with a hydrophilic polymer, their solubility and circulation half-lives are increased and their immunogenicity is masked. Quite surprisingly, the foregoing can be accomplished with little, if any, di inishment in binding activity.
  • Nonproteinaceous polymers suitable for use in accordance with the present invention include, but are not limited to, polyalkylethers as exemplified by polyethylene glycol and polypropylene glycol, polylactic acid, polyglycolic acid, polyoxyalkenes, polyvinylalcohol, polyvinylpyrrolidone, cellulose and cellulose derivatives, dextran and dextran derivatives, etc.
  • hydrophilic polymers have an average molecular weight ranging from about 500 to about 100,000 daltons, more preferably from about 2,000 to about 60,000 daltons and, even more preferably, from about 5,000 to about 50,000 daltons. In preferred embodiments, such hydrophilic polymers have average molecular weights of about 5,000 daltons, 10,000 daltons 20,000 daltons and 40,000 daltons.
  • the peptide compounds ofthe invention can be derivatized with or coupled to such polymers using any ofthe methods set forth in Zallipsky (1995) Bioconjugate Chem. 6:150-165; Monfardini et al. (1995) Bioconjugate Chem. 6:62-69; U.S. Patent No. 4,640,835; U.S. Patent No. 4,496,689; U.S. Patent No. 4,301,144; U.S. Patent No. 4,670,417; U.S. Patent No. 4,791,192; U.S. Patent No. 4,179,337 or WO 95/34326.
  • the N-terminus of a peptide ofthe invention is coupled to a polyethylene glycol molecule.
  • the polymer is selected from the group consisting of polyethylene glycol, polypropylene glycol, polylactic acid, polyglycolic acid and derivatives thereof.
  • the polymer is polyethylene glycol (PEG), in which case the peptide is referred to as "PEGylated.”
  • PEG is a linear, water-soluble polymer of ethylene oxide repeating units with two terminal hydroxyl groups. PEGs are classified by their molecular weights which typically range from about 500 daltons to about 40,000 daltons.
  • the PEGs employed have an average molecular weight of from about 500 to about 80,000 daltons. It is particularly preferred that the polymer has an average molecular weight of between about 5,000 to 40,000 daltons.
  • Preferred PEGylated compounds include, by way of example, the following:
  • the PEG coupled to the peptide compounds ofthe invention can be either branched or unbranched.
  • PEG is commercially available from Shearwater Polymers, Inc. (Huntsville, Alabama), Sigma Chemical Co. and other companies.
  • Suitable PEGs include, but are not limited to, monomethoxypolyethylene glycol (MePEG-OH), monomethoxypolyethylene glycol- succinate (MePEG-S), monomethoxypolyethylene glycol-succinimidyl succinate (MePEG- S-NHS), monomethoxypolyethylene glycol-amine (MePEG-NH 2 ), monomethoxypolyethylene glycol-tresylate (MePEG-TRES) and monomethoxypolyethylene glycol-imidazolyl-carbonyl (MePEG-IM).
  • MePEG-OH monomethoxypolyethylene glycol
  • MePEG-S monomethoxypolyethylene glycol- succinate
  • MePEG- S-NHS monomethoxypolyethylene glycol-succinimidyl succinate
  • MePEG-NH 2 monomethoxypolyethylene glycol-amine
  • MePEG-TRES monomethoxypolyethylene glycol-
  • the hydrophilic polymer that is employed e.g., PEG
  • an unreactive group such as a methoxy or ethoxy group.
  • the polymer is activated at the other terminus by reaction with a suitable activating agent, such as a cyanuric halide (e.g., cyanuric chloride, bromide or fluoride), diimidazole, an anhydride reagent (e.g., a dihalosuccinic anhydride, such as dibromosuccinic anhydride), acyl azide, />-diazoniumbenzyl ether, 3-(p-diazoniumphenoxy)-2-hydroxypropylether, or the like.
  • a suitable activating agent such as a cyanuric halide (e.g., cyanuric chloride, bromide or fluoride), diimidazole, an anhydride reagent (e.g., a dihalosuccinic anhydride, such as
  • the activated polymer is then reacted with a peptide compound of the invention to produce a polymer-derivatized peptide compound.
  • a functional group in the peptide compounds ofthe invention can be activated for reaction with the polymer, or two groups can be joined in a concerted coupling reaction using known coupling methods. It will be readily appreciated that the peptide compounds ofthe invention can be derivatized with PEG using a myriad of other reaction schemes known to those of skill in the art.
  • the N-terminus ofthe peptide is acetylated.
  • a preferred acetylated peptide is as follows:
  • the peptides and compounds ofthe invention can be modified with an acetyl moiety (Ac) using standard techniques known to those skilled in the art. On such technique includes combining the peptide with an acetylating reagent (e.g., acetyl chloride, acetic anhydride) in a suitable solvent to form the acetylated product. To the extent that other acetylated products are formed during the reaction, the N-terminus derivative can be isolated using conventional separation techniques.
  • an acetylating reagent e.g., acetyl chloride, acetic anhydride
  • the peptides and compounds ofthe invention can advantageously be modified to include an amide functionality at the carboxyl terminus ofthe peptide.
  • Preferred amidated peptides include:
  • RVTTCWPGEYGGVECYSVKY(NH 2 ) (SEQ ID NO: 95); and ERVTTCWPGEYGGVECYSVAY(NH 2 ) (SEQ ID NO: 105).
  • a benzhydrylamine resin is preferably used as the solid support for peptide synthesis.
  • a hydrogen fluoride treatment is employed to release the peptide from the support, directly resulting in the free peptide amide (i.e., the C-terminus is -C(O)NH 2 ).
  • proline analogues in which the ring size ofthe proline residue is changed from 5 members to 4, 6, or 7 members can be employed.
  • the peptides of the invention also serve as structural models for non-peptidic compounds with similar biological activity.
  • the peptide backbones may be replaced with a backbone composed of phosphonates, amidates, carbamates, sulfonamides, secondary amines, and N-methylamino acids.
  • the compounds ofthe invention are useful in vitro as unique tools for understanding the biological role of vascular endothelial cell growth factor, including the evaluation ofthe many factors thought to influence, and be influenced by, the production of vascular endothelial cell growth factor and the binding of vascular endothelial cell growth factor to the VEGFR-2 (e.g., the mechanism of endothelial cell growth factor signal transduction/receptor activation).
  • the present compounds are also useful in the development of other compounds that bind to VEGFR-2, because the compounds provide important structure-activity relationship (SAR) information that facilitates that development.
  • SAR structure-activity relationship
  • a compound ofthe invention can be used as a reagent for detecting a VEGFR-2 receptor or related receptor on living cells, fixed cells, in biological fluids, in tissue homogenates, in purified, natural biological materials, etc. For example, by labeling a compound ofthe invention, one can identify a cell expressing VEGFR-2 on its surface.
  • a compound ofthe invention can be used in in situ staining, FACS (fluorescence-activated cell sorting), Western blotting, ELISA (enzyme-linked immunoadsorptive assay), etc. or in receptor purification or in purifying cells expressing VEGFR-2 on the cell surface (or inside permeabilized cells).
  • a compound ofthe invention can also be utilized as a commercial research reagent for various medical research and diagnostic uses. Such uses include but are not limited to: (1) use as a calibration standard for quantitating the activities of candidate VEGFR-2 antagonists in a variety of functional assays; (2) use as a blocking reagent in random peptide screening, i.e., in searching for new families of VEGFR-2 peptide ligands; (3) use in the co-crystallization with VEGFR-2, i.e., a compound ofthe invention will allow formation of crystals bound to VEGFR-2, enabling the determination of receptor/peptide structure x-ray crystallography; (4) use in inhibiting or decreasing the proliferation and growth of vascular endothelial cell growth factor-dependent cell lines, such as human umbilical vein endothelial cells (HUVEC); and (5) other research and diagnostic applications wherein the VEGFR-2 is antagonized and such antagonization is conveniently calibrated against a known quantity of a vascular endotheli
  • a particularly preferred application for the compounds ofthe present invention is in imaging and assessing neovascularization during angiogenesis.
  • the expression ofthe VEGFR-2 in endothelial tissue and its reported concentration increase during angiogenesis may allow the compounds ofthe invention to be useful in imaging, and thereby assessing, neovascularization associated with several disorders including cancer.
  • the compounds ofthe invention may serve as sensitive and specific imaging agents for angiogenesis.
  • a compound ofthe invention can be labeled with a radionuclide such as 99m Tc, In or 123 I and injected intravenously into the patient.
  • Either 3-D single photon emission computed tomography (SPECT) imaging or 2-D planar gamma scintigraphy can be applied to localize tracer concentration that positively correlates with neovascularization.
  • SPECT computed tomography
  • There are numerous labeling mechanisms for the metals which would generally include either the use of bifunctional chelators or amino acid derived chelators.
  • labels such as 123 I can be attached via a covalent bond to the compound.
  • a compound ofthe invention can be labeled with a positron-emitting isotope such as ⁇ C, 18 F, 68 Ga, 76 Br, 61 Cu, or 64 Cu, injected into the patient and imaged using positron-emitting tomography (PET).
  • PET positron-emitting tomography
  • PET provides improved sensitivity and resolution compared with SPECT, although the availability of PET isotopes is lower as compared to the radionuclides used for SPECT.
  • a compound ofthe invention can be PEGylated and labeled with one ofthe above positron-emitting nuclei or gamma-emitting radionuclides to extend blood half life and reduce extravasation.
  • a compound ofthe invention can be attached to a liposome or amorphous polymer to reduce extravasation. The carrier (i.e. liposome or polymer) is then labeled with one or more ofthe above positron-emitting atoms or gamma-emitting radionuclides for detection with PET or SPECT.
  • the longer-lived nuclei such as ⁇ In or 76 Br are most appropriate as there could be significant delay between tracer administration and imaging while waiting for the unbound compound to be eliminated from the bloodstream.
  • the carrier can also be labeled with multiple copies of paramagnetic nuclei such as Gd, Fe or Mn for detection with magnetic resonance imaging (MRI). Although MRI sensitivity of tracer detection is relatively low (particularly when compared to SPECT or PET), spatial resolution is improved. In addition, MRI techniques have the added benefit of avoiding the use of ionizing radiation.
  • a compound ofthe invention can be attached to a monocrystalline iron oxide (MION) particle and used for detection via MRI.
  • a compound ofthe can be labeled with a fluorophore that preferentially emits in the near infrared region (700-1 OOOnm) and used for the detection of angiogenesis in tissue relatively close to the surface ofthe skin.
  • the invention provides a method for imaging and assessing neovascularization during angiogenesis comprising the steps of administering a compound ofthe present invention to a patient wherein the compound is coupled to a detectable label to form a labeled compound, allowing the labeled compound to bind to VEGFR-2, and detecting the detectable label.
  • the detectable label is selected from the group consisting of 99m Tc, m In, 123 I, ⁇ C, 18 F, 68 Ga, 76 Br, 61 Cu, 64 Cu, Gd, Fe, Mn and a fluorophore.
  • the detecting step is performed using SPECT, 2-D planar gamma scintigraphy, PET, MRI, infrared detection, or a combination thereof.
  • An antagonist compound ofthe invention can also be administered to a warm blooded animal, including a human, to treat a disease, condition or disorder that is responsive to a compound that antagonizes the effects of vascular endothelial cell growth factor in vivo.
  • the present invention encompasses methods for treating a patient who would benefit from administration of a VEGFR-2 antagonist, comprising administering to the patient a therapeutically effective amount of a VEGFR-2 antagonist and thus alleviate the symptoms associated with angiogenesis in vivo.
  • an antagonist compound of this invention will find use in the treatment of diseases such as psoriasis, rheumatoid arthritis, retinopathy and cancer.
  • an agonist compound ofthe invention can be administered to a warm blooded animal, including a human, to treat a disease, condition or disorder that is responsive to a compound that agonizes the effects of vascular endothelial cell growth factor in vivo.
  • the present invention encompasses methods for treating a patient who would benefit from administration of a VEGFR-2 agonist and thus alleviate the symptoms associated with decreased angiogenesis in vivo.
  • an agonist compound of this invention will find use in the treatment of diseases such as coronary artery disease wherein the formation of new blood vessels serving cardiac tissue at least partially remedies the decrease in blood flow from coronary vessels that are obstructed.
  • the invention includes pharmaceutical compositions comprising, as an active ingredient, at least one ofthe compounds ofthe invention in association with a pharmaceutical carrier or diluent.
  • the composition can be administered by oral, parenteral (intramuscular, intraperitoneal, intravenous (IV) or subcutaneous) injection, transdermal (either passively or using iontophoresis or electroporation), or transmucosal (nasal, vaginal, rectal, or sublingual) routes of administration, or using bioerodible inserts, and can be formulated in dosage forms appropriate for each route of administration.
  • Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules.
  • the active compound is admixed with at least one inert pharmaceutically acceptable carrier such as sucrose, lactose, or starch.
  • Such dosage forms can also comprise, as is normal practice, an additional substance other than an inert diluent, e.g., a lubricating agent such as magnesium stearate.
  • the dosage forms may also comprise a buffering agent. Tablets and pills can additionally be prepared with enteric coatings.
  • Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions and syrups, with the elixirs containing an inert diluent commonly used in the art, such as water. These compositions can also include one or more adjuvants, such as a wetting agent, an emulsifying agent, a suspending agent, a sweetening agent, a flavoring agent or a perfuming agent. Preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions, and emulsions.
  • non-aqueous solvents or vehicles examples include propylene glycol, polyethylene glycol, vegetable oils, such as olive oil and corn oil, gelatin, and injectable organic esters such as ethyl oleate.
  • Such dosage forms may also contain one or more adjuvants such as a preserving agent, a wetting agent, an emulsifying agent and a dispersing agent.
  • the dosage forms may be sterilized by, for example, filtration through a bacteria-retaining filter, by incorporating sterilizing agents into the compositions, by irradiating the compositions, or by heating the compositions. They can also be manufactured using sterile water, or some other sterile injectable medium, prior to use.
  • compositions for rectal or vaginal administration are preferably suppositories which may contain, in addition to the active substance, an excipient such as cocoa butter or a suppository wax.
  • Compositions for nasal or sublingual administration are also prepared with one or more standard excipients well known in the art.
  • the dosage of active ingredient in the compositions of this invention may be varied; however, it is necessary that the amount ofthe active ingredient is such that a suitable dosage form is obtained.
  • the selected dosage depends upon the desired therapeutic effect, the route of administration, the duration ofthe treatment desired, and other factors well known to those skilled in the art. Generally, dosage levels of between 0.001 to 10 mg/kg of body weight daily are administered to mammals.
  • the peptides of Table 1 were synthesized chemically using standard solid phase techniques as described above. To evaluate binding affinity, the synthesized peptides were tested against labeled VEGF by competing for binding to VEGFR-2 in a standard scintillation proximity assay (STND SPA). Briefly stated, the binding portion of human VEGFR-2 is replicated and thereafter attached to a suitable substrate using conventional techniques. A known amount of peptide is introduced together with a standard amount of 12 T labeled VEGF ( I25 I-VEGF-165 available from Amersham Pharmacia Biotech, Piscataway, New Jersey). After a sufficient period of time to allow for binding, the substrate is washed and a scintillation detector is used to detect I25 I labeled VEGF.
  • STND SPA scintillation proximity assay
  • the quantity of peptide used is not sufficient to displace the 125 I labeled VEGF. Alternatively, if no scintillation is detected, the quantity of peptide used is sufficient to remove the I25 I labeled VEGF.
  • the procedure is then repeated for a different peptide. In this way, it is possible to determine the relative binding affinities for each peptide.
  • Some peptides were also tested in a mouse VEGFR-2 based SPA (Mouse SPA). The testing procedure is substantially the same as that described immediately above with the exception that mouse-derived VEGFR- 2 is substituted for human VEGFR-2.
  • VEGF human umbilical vein endothelial cells
  • human umbilical vein endothelial cells By way of general procedure, human umbilical vein endothelial cells are placed on a suitable culture medium. A standard amount of VEGF along with a known amount of peptide is added to the cells. After a sufficient period of time, the cultures are inspected for detection of cell proliferation. If cell proliferation is detected, the quantity of peptide used is not sufficient to displace VEGF and consequently inhibit cell growth. Alternatively if no proliferation is detected, the quantity of peptide used is sufficient to inhibit VEGF-induced proliferation of cells.
  • the peptides, along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA), HUVEC and Mouse SPA, are shown in Table 1.
  • the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • each peptide is the result of effectively deleting the amino acid located on the N-terminus ofthe peptide in a sequential fashion.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA) and HUVEC are shown in Table 2.
  • the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • RVTTCWPGEYGGVECYSVKY (SEQ ID NO: 95), were synthesized chemically using standard solid phase techniques as described previously. As seen in Table 3, each peptide is the result of effectively deleting the amino acid located on the C-terminus ofthe peptide in a sequential fashion. The peptides were tested as described above for Examples 1-8. The peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA) are shown below. Again, the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • the peptides of Table 4 were synthesized chemically using standard solid phase techniques as described previously. The amino acid lysine, however, was substituted for other amino acids.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA), HUVEC and Mouse SPA are shown below. Again, the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • the peptides of Table 5 based were synthesized chemically using standard solid phase techniques as described previously.
  • the C-terminus of each peptide was amidated using techniques well known to those skilled in the art.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA) and HUVEC are shown below. As before, the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • the following peptide was synthesized chemically using standard solid phase techniques as described previously.
  • the N-terminus ofthe peptide was acetylated using techniques well known to those skilled in the art.
  • the peptide was tested as described above for Examples 1-8.
  • the peptide along with its corresponding IC 50 value (in nM) for standard SPA (STND SPA) is shown below.
  • the results ofthe assay reveals important information about the structure-activity relationship for VEGF to its receptor.
  • the peptides of Table 7 were synthesized chemically using standard solid phase techniques as described previously. The amino acid tryptophan, however, was substituted for other amino acids.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA), HUVEC and Mouse SPA are shown in Table 7. Again, the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • the peptides of Table 8 were synthesized chemically using standard solid phase techniques as described previously. The amino acid arginine, however, was substituted for other amino acids.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA) and HUVEC are shown below.
  • the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA) and HUVEC are shown below.
  • the results of these assays reveal important information about the structure-activity relationship for VEGF to its receptor.
  • the peptides were tested as described above for Examples 1-8.
  • the peptides along with their corresponding IC 50 values (in nM) for standard SPA (STND SPA) are shown below.
  • the results of these assays reveal important information about the structure- activity relationship for VEGF to its receptor.
  • These PEGylated compounds are also expected to exhibit improved pharmacokinetic values e.g., serum half-life, bioavailablity, etc.
  • the compounds ofthe invention are thus ligands for VEGFR-2.
  • the compounds ofthe invention demonstrate effective antagonism for the VEGFR-2.
  • 123 I is covalently attached to a compound ofthe invention using a conventional technique.
  • the labeled compound is administered intravenously to a cancer patient in an amount sufficient to detect neovascularization in the patient's tumor.
  • the tumor-affected area is imaged using three-dimensional single photon emission computed tomography.
  • a concentrated area of labeled-compound is detected indicating the presence of new blood vessels in the patient's tumor.
  • the patient's chemotherapy regimen is adjusted accordingly.

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Abstract

Nouveaux composés qui se lient à VEGFR-2. Ces nouveaux composés possèdent une chaîne peptidique d'une longueur appoximative de 8 à 40 acides aminés qui se lie à VEGFR-2, ou bien sont des dimères de telles chaînes peptidiques. Lesdits composés sont utiles en tant que sondes pour le criblage d'affinité et en tant qu'agents d'imagerie de l'angiogenèse. En outre, ces composés qui sont des antagonistes de VEGFR-2 sont utiles pour traiter, entre autres, des maladies dont le cancer, la rétinopathie, l'arthrite rhumatoïde. Des compositions pharmaceutiques et des méthodes d'utilisation desdites compositions sont également décrites.
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US9056138B2 (en) 2002-03-01 2015-06-16 Bracco Suisse Sa Multivalent constructs for therapeutic and diagnostic applications
US8642010B2 (en) 2002-03-01 2014-02-04 Dyax Corp. KDR and VEGF/KDR binding peptides and their use in diagnosis and therapy
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EP2559698A3 (fr) * 2007-08-24 2013-03-06 Julius-Maximilians-Universität Würzburg Peptides de récepteurs cycliques double mutants inhibant les anticorps bêta 1-adrénergiques
WO2009027063A3 (fr) * 2007-08-24 2009-07-23 Univ Wuerzburg J Maximilians Récepteurs peptidiques cyclisés double mutants inhibiteurs d'anticorps anti-récepteurs β1-adrénergiques
US9062095B2 (en) 2007-08-24 2015-06-23 Julius-Maximilians-Universtitat-Wurzburg Mutant double cyclized receptor peptides inhibiting β1-adrenoceptor antibodies
WO2012000862A1 (fr) * 2010-06-30 2012-01-05 Siemens Aktiengesellschaft Peptide marqué 11c pour la détection d'un tissu malade
WO2012000866A1 (fr) * 2010-06-30 2012-01-05 Siemens Aktiengesellschaft Peptide marqué 11c pour la détection d'un tissu malade
WO2012000865A1 (fr) * 2010-06-30 2012-01-05 Siemens Aktiengesellschaft Peptide marqué au 11c servant à détecter une tumeur exprimant un récepteur her2/neu
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WO2001083693A3 (fr) 2003-04-10
WO2001083693A8 (fr) 2002-01-31

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