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WO1996000079A1 - Produits d'addition copolymeres greffes de composes de platine (ii) - Google Patents

Produits d'addition copolymeres greffes de composes de platine (ii) Download PDF

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Publication number
WO1996000079A1
WO1996000079A1 PCT/US1995/007329 US9507329W WO9600079A1 WO 1996000079 A1 WO1996000079 A1 WO 1996000079A1 US 9507329 W US9507329 W US 9507329W WO 9600079 A1 WO9600079 A1 WO 9600079A1
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Prior art keywords
adduct
acid
polymer
group
poly
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PCT/US1995/007329
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English (en)
Inventor
Alexei Bogdanov
Ralph Weissleder
Thomas J. Brady
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The General Hospital Corporation
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Priority to AU27013/95A priority Critical patent/AU2701395A/en
Publication of WO1996000079A1 publication Critical patent/WO1996000079A1/fr

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    • 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
    • A61K47/62—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 a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A61K47/645—Polycationic or polyanionic oligopeptides, polypeptides or polyamino acids, e.g. polylysine, polyarginine, polyglutamic acid or peptide TAT
    • 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

Definitions

  • the present invention relates to graft co-polymer adducts which include a platinum (II) compound.
  • the compositions provide for lower toxicity, sustained release and stabilization of platinum (II) compounds, as well as selective delivery to a tumor site.
  • Contrast Agents for Magnetic Resonance Imaging Accurate detection of abnormalities in a patient's body is an essential prerequisite for diagnosing and adequately treating disease.
  • Visualization methods e.g., magnetic resonance imaging (MRI)
  • MRI magnetic resonance imaging
  • tissues of different origin such as normal and deviated, e.g., cancerous, tissues, may be differentiated on the basis of differences in relaxation times Tl, the spin-lattice or longitudinal relaxation time, or T2, the spin-spin or transverse relaxation time. Because of these differences, differential signal intensity is produced which gives various degrees of contrast in MR images.
  • MR imaging techniques employed to elucidate blood perfusion defects are based on the differentiation of flowing blood from stationary surrounding tissues, e.g., MR angiography (MRA) .
  • MRA MR angiography
  • Three dimensional angiographic techniques, e.g., “Time of Flight” (TOF) and “Phase Contrast” (PC) techniques provide detailed images of intracranial vessels.
  • Gadolinium (III) diethylenetriamine pentaacetic acid (Gd-DTPA) di eglumine is a widely used contrast agent which is relatively small (MW 538) and extravasates on the first pass through the capillaries.
  • Gd-DTPA Gadolinium diethylenetriamine pentaacetic acid
  • the use of Gd-DTPA for MR angiography in all organs except the brain is limited, since the blood half-life of Gd- DTPA is less than 20 minutes, and the biological life in man of GD-DTPA is about 90 minutes. The extravasation results in a rapid decrease in vessel/muscle signal ratio, which makes the accurate detection of abnormalities and disease difficult.
  • Gd-DTPA dimeglumine which is used in clinical practice, is immunogenic, which does not favor its repetitious administration to the same patient.
  • ferrioxamine-B as a contrast agent.
  • ferrioxamine-B causes a precipitous drop in blood pressure after its intravenous administration.
  • MRI contrast agents created using natural and synthetic macromolecules offer the advantage of high molecular relaxivity due to the multiple chelating groups coupled to a single polymer backbone. These groups can chelate paramagnetic cations, e.g., in Gd-DTPA-poly-1- lysine, or produce high relaxivity due to the presence of iron oxide, e.g., in iron-containing colloids.
  • iron oxide-based colloids have their own ligand- independent specific site of accumulation in the body, e.g., the liver, spleen, and lymphoid tissues.
  • Chelating groups may be attached to a variety of natural polymers, e.g., proteins and polysaccharides, and synthetic polymers. Chemical attachment, e.g., by conjugation, of DTPA to bovine serum albumin will result in a macromolecular contrast agent, which is suitable for some applications, e.g., NMR-angiography, but because of the efficient recognition of modified albumin by macrophages, and albumin-receptors on endothelial cells this contrast agent has a short blood half-life. It is also immunogenic and toxic to reticuloendothelial system organs. Therefore, use for MR imaging is limited.
  • albumin One way to diminish the antigenicity of albumin is to mask it with natural and synthetic polymers, e.g., spacer arms, by covalent attachment, but this leaves few reactive groups in the protein globule which are needed for binding the chelates and paramagnetic cations. Therefore, the use of such complexes in MR imaging is limited.
  • Synthetic polymers of 1-amino acids such as poly- 1-lysine (PL)
  • PL modified with DTPA can be used as a radionuclide carrier for antibody-mediated targeting in nuclear medicine.
  • Poly-1- lysine-DTPA i.e., poly-1-lysine with DTPA groups bonded to epsilon-amino groups of lysine residues has been suggested as a Gd complexone, .i.e., a compound which forms a complex with Gd, for use in MR angiography. It is also known that the toxicity of DTPA-poly-1-lysine is lower than that of DTPA-albu in.
  • DTPA-moieties on DTPA-polylysine are recognized by liver Kupffer cells and some kidneys cells, presumably glomerulonephral phagocytes, which cause elevated and relatively rapid removal of the contrast agent from the blood.
  • 90% of the intravenously injected agent e.g., poly-l-lysine-DTPA(Gd) (MW 48.7 kD)
  • Gd poly-l-lysine-DTPA(Gd)
  • synthesis of DTPA-poly-1-lysine can be carried out with a cross- linking reagent, e.g., cyclic anhydride of DTPA.
  • Nitrogen-containing polymers e.g., polethyleneimine
  • polethyleneimine have been modified with monofunctional derivatives of acetic acid to form a molecule where the backbone nitrogens and acetic acid residues are involved in complex formation with trivalent cations.
  • paramagnetic complexes of polyethyleneiminoacetic acid are not widely used in MRI.
  • Polymeric contrast agents e.g., starburst dendrimers, constitute a separate family of macromolecules with limited potential value as contrast agents. This family of agents has not been shown to be biocompatible and thus its value for in vivo imaging is limited.
  • PEG polyethylene glycol
  • MPEG monomethyl ester
  • Contrast agents containing MPEG or PEG as a component of paramagnetic mixtures or in cross-linked paramagnetic polymers also have been used.
  • Contrast agents targeted to the sites of interest help to increase the effectiveness of MR imaging methods.
  • diagnostic agents may include combinations of a ligand and a paramagnetic contrast agent coupled by strong interaction, e.g., a covalent chemical bond.
  • a contrast agent accumulates in the target site which is determined by ligand specificity.
  • the ligand which directs the contrast agent to the target site may be specific to receptors on either normal or transformed cells of a given organ or tissue. In the first case the contrast agent will be accumulated in normal tissue; in the second case, it will be accumulated in altered tissue.
  • Success in designing a targeted contrast agent is mainly determined by the following properties: 1. avidity to target site; 2. antigenicity, i.e., ability to pass through capillary endothelium; and 3. blood half- life of the ligand or targeting ("vector") molecule.
  • Coupling a contrast agent to a targeting ligand molecule e.g., an antibody or its fragments, which creates a targeted contrast agent, e.g., a chelated paramagnetic cation, paramagnetic colloid or combination of a chelate and a paramagnetic colloid conjugated to a targeting molecule, typically decreases its potential value for any of a number of reasons, e.g., decreased avidity to a target site, increased antigenicity, or decreased half- life.
  • a targeting ligand molecule e.g., an antibody or its fragments
  • a targeted contrast agent e.g., a chelated paramagnetic cation, paramagnetic colloid or combination of a chelate and a paramagnetic colloid conjugated to a targeting molecule
  • a small antibody fragment e.g., a Fab or Fv chi eric molecule
  • a large paramagnetic molecule e.g., DTPA-poly er
  • a superparamagnetic colloid e.g., iron oxide
  • the paramagnetic molecule or colloid itself may be recognized by the recipient organism's opsonizing proteins and the contrast agent may be trapped in reticuloendothelial system organs.
  • the contrast agent is removed from the circulation by the liver and spleen before any substantial concentration is achieved in the target site.
  • such a contrast agent may be recognized as a foreign antigen which may give rise to undesirable host antibodies.
  • Cis-diaminedichlorplatinum(II) is a platinum (II) compound which is used to treat bladder, lung, head, neck cervical, testicular and ovarian cancers (Sherman and Lippard, Chem. Rev. 87, 1153 (1987).
  • platinum (II) compounds of known or potential therapeutic value include cis-diamminediaquoplatinum (II) (i.e.
  • cDDP and carboplatin are also effective in combination with certain other chemotherapeutic drugs (e.g. doxorubicin, cyclophosphamide) in the treatment of cancer, for example, squamous cell carcinoma, metastatic melanoma, metastatic bladder carcinoma, basal cell carcinoma, and astrocytoma (Physician's Desk Reference pp. 754-757 (1993)).
  • chemotherapeutic drugs e.g. doxorubicin, cyclophosphamide
  • squamous cell carcinoma, metastatic melanoma, metastatic bladder carcinoma, basal cell carcinoma, and astrocytoma Physical target of cDDP is DNA, especially the DNA of rapidly dividing cells such as cancer cells.
  • platinum(II) compounds particularly cDDP and carboplatin
  • cDDP toxicity causes nephrotoxicity in 30% of the patients that receive the drug and other adverse reactions have been documented (Physician's Desk Reference, supra) .
  • cDDP exhibits a complex pattern of inactivation and elimination from the body, for example, approximately 10% is rapidly removed from the systemic system.
  • a large fraction of the remaining cDDP (>85%) is inactivated by binding with systemic proteins, for example, blood proteins. Therefore, a major clinical problem with the therapuetic administration of cDDP and other platinum (II) compounds is that a large fraction of the drug is rapidly inactivated and eliminated before contacting a tumor.
  • Another cDDP delivery system involves administering a non-crosslinked (i.e. linear or branched) homopolymer or co-polymer, combined with cDDP (Maeda, M. et al., Anti-Cancer Drugs 167 (1993); Yoshida M. et al., supra ; Schecter, B. et al., Cancer Chemother. Pharmacol . 24, 161 (1989); Schecter B. et al., Int . J. Cancer 39, 409 (1987); Schechter, B. et al., Cancer Biochem . Biophys . 8, 277 (1986); ibid, pg. 289). These systems result in a homopolymeric or co-polymeric adducts which are toxic and do not exhibit desirable solubility.
  • the invention features a biocompatible medical composition including a polymeric carrier, a protective chain linked to the polymeric carrier, and a reporter group linked to the carrier or to the carrier and the protective chain.
  • the polymeric carrier may be chosen from the group of polyamino acids, polyethyleneimines, natural saccharides, aminated polysaccharides, aminated oligosaccharides, polyamidoamine, polyacrylic acids, or polyalcohols.
  • the invention also features a composition having the formula:
  • R l R 2 groups can be linked in any order, e.g., the R ⁇ unit can be repeated several times in the chain before an R 2 unit occurs, and vice versa; wherein k is 100-560; R ⁇ . is (CH 2 ) 4 NHCO(CH 2 ) n COOCH 2 CH 2 A-B-OR 3 , where n is 2-6; A is [OCH 2 CH 2 ] ⁇ , where x is 15-220; B is [OCH 2 CH 2 ] ⁇ or [OCH(CH 3 )CH 2 ] , where y+x is 17-220; R 2 is a chelating group; and R 3 is H, (CH 2 ) y CH 3 or (CH 2 ) COOH, and p is 0-7.
  • the chelating group may be, e.g., diethylenetriamine pentaacetic acid,
  • the polyamino acid of the composition preferably has 20-560 amino acid units, a molecular weight of 1,000- 100,000 daltons, and is preferably non-proteinaceous.
  • the polyamino acid may be a polymer of a single species, or at least two different species of amino acid, or may be a block co-polymer.
  • the polyamino acid may include polyamino acid fragments linked by cleavable bonds, e.g., S-S bonds.
  • the polyamino acid may be, e.g., poly-1- lysine, poly-d-lysine, poly-alpha,beta-(2-aminoethyl) -D,L aspartamide, or poly-1-aspartic acid.
  • the protective chain of the composition may be, e.g., polyethylene glycol, methoxypolyethylene glycol, methoxypolypropylene glycol, a co-polymer of polyethylene glycol, methoxypolyethylene glycol, or methoxypolypropylene glycol, or derivatives thereof.
  • the protective chain may be a block co-polymer of polyethylene glycol and one of the group of polyamino acids, polysaccharides, polyamidoamines, polyethyleneamines, or polynucleotides.
  • the protective chain may also be a co-polymer of polyethylene glycol including a monoester of a dicarboxylic acid.
  • the protective chain preferably has a molecular weight of
  • the reporter group may be a complexone, e.g., a chelating group.
  • the chelating group may be, e.g., diethylenetriamine- pentaacetic acid, triethylenetetramine-hexaacetic acid, ethylenediamine- tetraacetic acid, 1,2-diaminocyclo- hexane-N,N,N' ,N'-tetra-acetic acid,
  • N,N' ,N' '-triacetic acid or 1,4,8, 11-tetraazacyclotetra- decane-N,N' ,N' ' ,N' ' '-tetra-acetic acid.
  • the composition may further include an alfa-, beta-, or gamma-emitting radionuclide linked to the complexone.
  • the radionuclide may be gallium 67, indium 111, technetium 99m, chromium 51, cobalt 57, molibdenium 99, or a molecule linked to an iodine isotope.
  • the reporter group may also include a diagnostic agent, e.g., a contrast agent, which may include a paramagnetic or superparamagnetic element, or a combination of a paramagnetic element and a radionuclide.
  • a diagnostic agent e.g., a contrast agent, which may include a paramagnetic or superparamagnetic element, or a combination of a paramagnetic element and a radionuclide.
  • the paramagnetic element may be chosen from the group of transitional metals or lanthanides having atomic numbers 21-29, 42, 44, or 57-71.
  • the paramagnetic element may be, e.g., gadolinium (III), dysprosium (III), holmium (III) , europium (III) , iron (III) , or manganese (II) .
  • the invention also features a composition in which the reporter group includes a therapeutic agent such as a cytostatic, antibiotic, hormonal, analgesic, psychotropic, anti-inflammatory, antiviral, or antifungal drug, or a lymphokine.
  • a therapeutic agent such as a cytostatic, antibiotic, hormonal, analgesic, psychotropic, anti-inflammatory, antiviral, or antifungal drug, or a lymphokine.
  • the composition may further include a targeting group linked to the polymeric carrier or the protective chain or both.
  • the targeting group may be an antibody, fragment of an antibody, chimeric antibody, enzyme, lectin, or saccharide ligand.
  • the composition may also include a reporter group which is a particle, colloidal particle, or a colloidal precipitate.
  • the colloidal precipitate may include an oxide, sulfide, or hydroxide of a transitional element, or lanthanide having atomic numbers 21-29, 42, 44, or 57- 71.
  • the reporter group may also be a silicon oxide colloid or polymer containing silicon, sulfur, or carbon, or a fluorine-containing molecule, e.g., a fluorocarbon.
  • the reporter group may also be a pyridiyldithioacyl group, e.g., a N-(2- pyridyldithio)propionyl group, N-hydroxysuccinimidyl, N- hydroxysulfosuccinimidyl, imidazolyl, benzotriazolyl, aminoalkyl, aldehyde, thioalkyls, thiolane, haloid acyl, haloid alkyl, or haloid phenyl, or a diazo- or hydrazo- group, e.g., a 4-hydrazionoxyethyl, 4-hydrazino- benzyl, diasirinyl, azidophenyl, or azidoalkyl group.
  • a pyridiyldithioacyl group e.g., a N-(2- pyridyldithio)propionyl group, N-hydroxy
  • the invention features a method of preparing the composition by linking the polymeric carrier with the protective chain to produce a protected carrier, and then linking the protected carrier with the reporter group. If the protective chain includes a methoxypolyethylene glycol analog, linking the polymeric carrier with the analog produces a semi-stable gel.
  • the method may further include linking a targeting group to the carrier, protective group, or both.
  • the invention also features a method of treating a disease in a patient by administering to the patient a therapeutically or diagnostically effective amount of the composition of the invention.
  • the method may further include scanning the patient using an imaging technique which can detect the reporter group to obtain a visible image of the distribution of the composition.
  • the administration may be by intravascular or intraperitoneal injection, and the imaging technique may be, e.g., magnetic resonance imaging, nuclear medicine imaging, position emission tomography, or single-photon-emission computed tomography.
  • the reporter group may include gadolinium supplied at a dosage of less than 0.05 mmol Gd/kg of body weight of the patient.
  • the dosage is about 0.02 to 0.04 mmol Gd/kg of body weight.
  • the invention also features a method of treating a patient by scanning a submillimeter vessel of the patent to obtain a visible image of the submillimeter vessel.
  • a submillimeter vessel is one that has an inner diameter of less than one millimeter.
  • the invention also features a biocompatible graft- co-polymer adduct which includes a polymeric carrier, a protective chain linked to the polymeric carrier, and a platinu (II) compound which is reversibly linked to the polymeric carrier or the protective chain or both the polymeric carrier and the protective chain.
  • the graft-co-polymer adduct includes a polymeric carrier, a protective chain linked to the polymeric carrier, a reporter group linked to the polymeric carrier or to the carrier and the protective chain and a platinum(II) compound which is reversibly linked to the polymeric carrier or the protective chain or both the polymeric carrier and the protective chain.
  • Graft-co-polymer adducts of the invention are therapeutic agents which provide dual pharmaceutical action: 1) systemic release of a platinum(II) compound from a graft co-polymer while the adduct circulates in the bloodstream and 2) targeted delivery of a bioactive platinum (II) compound to a tumor.
  • the graft-co-polymer adduct is capable of forming a circulating systemic depot for the sustained release of platinum(II) compounds; the adduct can also be targeted to a tumor.
  • the graft co-polymer adduct lowers the toxicity of platinum(II) compounds (i.e.
  • the co-polymer of a graft co-polymer is a negatively charged macromolecule which includes a backbone polymer covalently grafted with protective chains;
  • the backbone polymer of a graft co-polymer is preferably a polyacid, e.g. polyaspartic or polyglutamic acid, polylysine or carboxylated polylysine.
  • a negatively charged polymer is useful since it is capable of forming ionic bonds with aquated platinum (II) compounds.
  • the protective chain of a graft co-polymer is preferably a polymer of ethylene oxide
  • a protective chain is useful because: 1) it ensures the adduct solubility while maintaining a high drug payload. For example, with cDDP, approximately 30% by weight, or >l mol cisplatin/per mole carrier of carboxyl groups can be formed; 2) a protective chain assists in the formation of a steric barrier which prevents a platinum(II) compound from binding to molecules in the body, for example, plasma albumin; and 3) a protective chain provides a platinum(II) compound in a form which permits long circulation times (i.e creates a circulating depot).
  • the invention features an adduct which includes a polymeric carrier chosen from the group consisting of polyamino acids, preferably non- proteinaceous polyamino acids, polyethyleneimines, natural saccharides, aminated polysaccharides, aminated oligosaccharides, polyamidoamine, polyacrylic acids, polyalcohols, sulfonated polysaccharides, sulfonated oligosaccharides, carboxylated polysaccharides, carboxylated oligosaccharides, aminocarboxylated polysaccharides, aminocarboxylated oligosaccharides, carboxymethylated polysaccharides, and carboxymethylated oligosaccharides; where the polyamino acid has 20-560 amino acid residues; the polyamino acid has a
  • the adduct includes a protective chain which is polyethylene glycol, polypropylene glycol, a co-polymer of polyethylene glycol and polypropylene glycol; or a monoesterified derivative thereof, preferrably methoxypolyethylene glycol, methoxypolypropylene glycol, or a co-polymer of methoxypolyethylene glycol and methoxypolypropyleneglycol;
  • the protective chain is polyethylene glycol monoamine, methoxypolyethylene glycol monoamine, methoxy polyethylene glycol hydrazine, methoxy polyethylene glycol imidazolide or a polyethylene glycol diacid;
  • the protective chain is a block co-polymer of polyethylene glycol and one of the group of polyamino acids, polysaccharides, polyamidoamines, polyethyleneamines, or polynucleotides;
  • the protective chain is a co-polymer of polyethylene glycol comprising a monoester of a di
  • the adduct includes a reporter group.
  • the reporter group is a complexone, such as a chelating group, preferrably the chelating group is diethylenetriamine-pentaacetic acid, triethylenetetraminehexaacetic acid, ethylenediamine- tetraacetic acid, 1,2-diaminocyclo- hexane-N,N,N' ,N'-tetra-acetic acid, N,N'-Di(2-hydroxybenzy1)ethylenediamine, N-(2-hydroxyethyl)ethylenediaminetriacetic acid, nitrilotriacetic acid, ethylene-bis(oxyethylene- nitrilo)tetraacetic acid, 1,4,7,10,-tetraazacyclodo- decane-N,N' ,N' ' , N' ' '-tetraacetic acid,
  • the reporter group includes a diagnostic agent, such as a contrast agent, preferably the contrast agent is a paramagnetic element, preferably the paramagnetic element is chosen from the group of transitional metals or lanthanides having atomic numbers 21-29, 42, 44, or 57-71, preferably gadolinium (III), dysprosium (III) , holmium (III) , europium (III) , iron (III) , or manganese (II) .
  • the contrast agent can also include a superpara agnetic element.
  • the reporter group includes a complexone which includes an alpha-, beta-, or gamma-emitting radionuclide linked to the complexone, preferably the radionuclide is gallium 67, indium 111, technetium 99m, chromium 51, cobalt 57, molibdenum 99, or a molecule linked to an iodine isotope.
  • the reporter group includes a therapeutic agent, preferably a cytostatic, antibiotic, hormonal, analgesic, psychotropic, anti- inflammatory, antiviral, or antifungal drug, or a lymphokine;
  • the reporter group is a particle, colloidal particle, or a colloidal precipitate, preferably the colloidal precipitate includes includes an oxide, sulfide, or hydroxide of a transitional element or lanthanide having atomic numbers 21-29, 42, 44, or 57-71;
  • the reporter group is a silicon oxide colloid or polymer containing silicon, sulfur, or carbon;
  • the reporter group has the general formula -COOH or -(CH 2 ) p COOH, where p is between 1 and 7, inclusive; preferably the reporter group is -CH 2 CH 2 COOH.
  • the adduct includes a reversibly linked Pt(II) compound of the general formula:
  • each R a , R b , R c , R d independently is -OH 2 , Cl, Br, I, -NH 2 , or -N(R.) 2 , where each R e independently is H, lower alkyl, or lower cycloalkyl, with the proviso that both of R e are not H; and each R a , R b , R c , and R d is the same or different; or b) R a and R d are combined to form a linking group of the formula: -NH(CH 2 ) p2 NH-, where p2 is 1 or 2; -0-CO-C(CH 2 ) p3 -CO-0-, where p3 is between 4 and 6, inclusive; / ⁇ - ⁇ CUUH
  • R a and R d , R b and R c are each independently combined to form a linking group of the formula: -NH(CH 2 ) 2 NH ⁇ / where p2 is 1 or 2;
  • the Pt(II) compound is any one of cDDP, cis-aq, carboplatin, iproplatin, DACCP, malonatoplatinum, trans (+/-)-1,2-cylcohexanediammineplatinum (II), cis- DEP, or Pt(II) (NH 3 ) (RNH 2 )C1 2 , where R is H, methyl, ethyl, isopropyl, cyclopropyl, cyclobutyl, cyclopentyl, or cylcohexyl.
  • the invention features a method of preparing an adduct of the invention, the method including: a) linking a polymeric carrier with a protective chain to produce a protected carrier; and b) combining a Pt(II) compound with the graft co- polymer in order to form a graft co-polymer adduct, where the adduct includes a reversibly linked Pt(II) compound.
  • the invention features a method of preparing an adduct of the invention, the method including: a) linking a polymeric carrier with a protective chain to produce a protected carrier; b) linking the protected carrier with a reporter group sufficient to form a graft co-polymer; and c) combining a Pt(II) compound with the graft co- polymer in order to form a graft co-polymer adduct, where the adduct includes a reversibly linked Pt(II) compound.
  • the invention features a method of treating a disease, preferably cancer, in a patient including administering to the patient a therapeutically effective amount of an adduct of the invention.
  • the method further includes scanning the patient using an imaging technique which can detect a reporter group to obtain a visible image of the distribution of an adduct of the invention; the administration is by intravascular or intraperitoneal injection; the imaging technique is magnetic resonance imaging, nuclear medicine imaging, position emission tomography, or single-photon-emission computed tomography; the cancer is bladder, lung, head, neck, cervical, testicular or ovarian cancer in a human; and the method further includes administering a chemotherapeutic drug, preferably cDDP, carboplatin, doxorubicin or cyclophosphamide.
  • the invention features an adduct with a reporter group which includes gadolinium supplied at a dosage of less than 0.05 mmol Gd/kg of body weight of the patient.
  • the method further includes scanning a submillimeter vessel of the patient to obtain a visible image of the submillimeter vessel.
  • the invention features a method of selectively accumulating a Pt(II) compound, preferably cDDP or carboplatin in a mammalian tumor, preferably a human tumor, the method including administering an adduct of the invention to the mammal under conditions which allow the adduct to selectively accumulate in the tumor.
  • selective accumulation is meant that the adduct is preferentially concentrated in a tumor rather than surrounding tissues.
  • the invention features a method of providing a circulating depot of a bioactive Pt(II) compound in a mammal, preferably cDDP or carboplatin provided to a human, the method including administering an adduct of the invention to the mammal in an amount sufficient to provide a circulating depot of the bioactive Pt(II) compound.
  • the invention features an adduct which includes between 0.1% and 30% (w ⁇ w) , inclusive, of platinum, and exhibits a molecular weight of between 50 and 1500 kDa, inclusive.
  • the invention features an adduct which includes the graft co-polymer poly[([N- (methoxy poly(ethylene)glycol)-o-succinyl]-1-lysyl)n-(N- succinyl-l-lysyl)m]lysine and exhibits a molecular weight of between 1500 and 150,000 daltons, inclusive; where the succinate and the Pt(II) compound, preferably cDDP, are present in a molar ratio of between 1:1 and 1:20 (inclusive) , respectively.
  • the invention features an adduct where the linked polymeric carrier, protective chain and reporter group has the general formula: 0 0
  • R 2 groups can be linked in any order and k is 100-560; and a) R ⁇ is (CH 2 ) 4 NHCO(CH 2 ) n COOCH 2 CH 2 A-B-OR 3 , where n is 2-6; A is [OCH 2 CH 2 ] x , where x is 15-220; B is
  • R 2 is a chelating group; and R 3 is H, (CH 2 ) CH 3 or (CH 2 ) COOH, where y is 0-7; or b) R-L is -CH 2 (R g )NHCO(CH 2 ) nl COO((CH 2 ) n2 0) n3 CH 3 , where R g is -CH 2 CH 2 CH 2 -, -CO- or -CH 2 CO-, nl is 2 to 6, inclusive, n2 is 2 or 3, n3 is 10-200, inclusive; and R 2 is -CH 2 (R )NHCOR h , where R h is - COOH or -(CH 2 ) 2 COOH, where y2 is 1 to 7, inclusive.
  • the chelating group is diethylenetriamine pentaacetic acid, l,4,7,10,-tetraazacyclododecane-N,N' ,N",N" '-tetraacetic acid, l,4,7,10,-tetraazacyclododecane-N,N' ,N' ' ,-triacetic acid, ethylene-bis(oxyethylenenitrilo)tetraacetic acid, or ethylenediaminetetraacetic acid.
  • the invention features an adduct where the reporter group is a pyridiyldithioacyl group, or a diazo- or hydrazo-group, preferably the pyridyldithioacyl group is a N-(2-pyridyldithio)propionyl group, N-hydroxysuccinimidyl, N-hydroxysulfosuccinimidyl, imidazolyl, benzotriazolyl, aminoalkyl, aldehyde, thioalkyls, thiolane, haloid acyl, haloid alkyl, or haloid phenyl; the diazo- or hydrazo- group is 4- hydrazionoxyethyl, 4-hydrazinobenzyl, diasirinyl, azidophenyl, or azidoalkyl groups.
  • the invention features an adduct which includes a reporter group which is a fluorine-containing molecule.
  • the term "linked" means covalently or non-covalently bonded, e.g., by hydrogen, ionic, or Van-der-Waals interactions. Such bonds may be formed between at least two of the same or different atoms or ions as a result of a redistribution of electron densities of those atoms or ions.
  • reversibly linked means a non-covalent bond, e.g., hydrogen, ionic, or Van-der- Waals interactions which stabilizes a Pt(II) compound with a graft co-polymer and which is reversed or dissociated under human physiological conditions in vivo.
  • a “polymeric carrier” is a molecule comprised of several linked chemical moieties which may be the same or different, and serves as a site where a reporter group is linked and is shielded by protective chains.
  • a “protective chain” is a molecule(s) which protects a carrier molecule and a reporter group from contact with other macromolecules due to extensive linking of water to the chains.
  • a “complexone” is a molecule or several molecules or chemical radicals or moieties which constitute a favorable environment for linking an ion (a cation or an anion) . Dissociation of the ion from the environment is hindered due to kinetic or/and thermodynamic stability of linking.
  • a “chelating molecule” or “chelate” is a complexone which links cations.
  • reporter group as used herein is a non- traditional definition which includes an atom, ion, molecule, or complexone that may be linked to a polymeric carrier or protective chain and which can be detected by any methods disclosed herein.
  • a reporter group may be a therapeutic or diagnostic agent.
  • derivatives or analogs as used herein mean a compound whose core structure is the same as, or closely resembes that of, a parent compound, but which has a che cial or physical modifaction, such as a different or additional side groups; the term indues co- polymers of parent compunds that can be linked to other atoms or molecules.
  • ligand means any atom, ion, or molecule linked to a carrier and/or to a protective chain and/or to a reporter group to increase the accumulation of the composition in a target site of an organism to a greater degree through the targeting group were absent.
  • polyamino acid fragment means individual amino acid radicals or several linked amino acids which may be linked to form a polyamino acid.
  • a “semi-stable gel” is a gel which forms a liquid phase by standing, or when temperature, pH or other conditions are varied.
  • vessel mapping refers to obtaining an image of a vessel or vessels where spatial orientation and delineation of vessels may be elucidated.
  • aminoated describes molecules including linked amino groups.
  • a “diagnostically effective amount” of the composition is an amount that will provide an image of the composition in the patient.
  • a “therapeutically effective amount” of the composition is an amount that will provide a therapeutic benefit to the patient.
  • a “lower alkyl”, as used herein, is a branched or straight chain hydrocarbon of between 1 and 6 carbon atoms, inclusive.
  • a “lower cycloalkyl”, as used herein, is a cyclic hydrocarbon of between 4 and 6 carbon atoms, inclusive.
  • a bioactive Pt(II) compound is a Pt(II) compound, either free or reversibly linked with an adduct of the invention, which is capable of forming one or more covalent linkages with DNA under human physiological conditions in vivo.
  • compositions of this invention which make them surprisingly suitable for MR imaging include: 1) the ability to chelate paramagnetic cations to achieve a high molecular relaxivity, which is essential for its use as an NMR contrast agent 2) an extended blood half-life 3) low toxicity and 4) non-immunogenicity.
  • This invention also provides the advantages of only having to administer one dose of the contrast agent, along with enhanced signal/noise ratios in the diagnostic images obtained.
  • compositions of the invention 1) increased relaxivity of each paramagnetic cation compared to Gd-DTPA, 2) large numbers of chelating groups on each molecule, 3) enhanced blood pool concentration after intravenous injection, 4) enhanced sites of abnormal endothelial permeability, and 5) prolonged circulation time -compared to Gd-DTPA.
  • Fig. 1 is a diagram of three schemes for synthesizing the compositions of the invention.
  • Fig. 2 is a graph of the blood clearance of [ 1:L1 In]-labeled and Gd-saturated MPEG(MW 5 kD)-poly-1- lysine(MW 53.5 kD)-DTPA (squares) and MPEG(MW 2 kD)-poly- l-lysine(MW 41 kD)-DTPA (diamonds).
  • Fig. 3 is a graph of the biodistribution of
  • Fig. 4 is a graph of the response to Gd-DTPA of mice injected with Gd-DTPA-BSA (squares) and MPEG(MW 5 kD)-poly-l-lysine(MW 53.5 kD)-DTPA(Gd) (diamonds).
  • Fig. 5 is a graph of the effect of Gd-labelled MPEG(MW 2 kD)-poly-l-lysine(MW 41 kD)-DTPA (squares) or Gd-labelled MPEG(MW 5 kD) -poly-l-lysine(MW 25 kD)-DTPA (diamonds) on Tl values of blood at various concentrations.
  • Fig. 6 is a graph of the dose-dependent enhancement of vessels, with the vessel/muscle ratio determined by digitization of signal intensities of several large arteries, e.g., aorta, iliac, and femoral, and nearby muscle tissue.
  • Fig. 7 is a graph of the time-course of a contrast agent in large vessels in a comparative study.
  • Figs. 8a and 8b are MR images of the head of a rat in 3-D bright-pixel reconstruction showing the image before (Fig. 8a) and after (Fig. 8b) an intravenous injection of MPEG(MW 5 kD) -poly-l-lysine(MW 25 kD)- DTPA(Gd) .
  • Fig. 9 is an MR image of two rats in 3-D bright- pixel reconstruction after an intravenous injection of MPEG(MW 5kD)-poly-l-lysine(MW 25 kD)-DTPA(Gd) (left image) and gadopentate dimeglumine (right image) .
  • Figs. 8a and 8b are MR images of the head of a rat in 3-D bright-pixel reconstruction showing the image before (Fig. 8a) and after (Fig. 8b) an intravenous injection of MPEG(MW 5 kD) -poly-l-lysine(MW 25 kD)-DTPA(Gd)
  • FIGS. 10a and 10b are of MR images of a rabbit in 3-D bright-pixel reconstruction of the lateral (Fig. 10a) and cranio caudal projection (See Fig. 10b) after an intravenous injection of MPEG (MW 5 kD) -poly-l-lysine(MW 25 kD)-DTPA(Gd) .
  • Figs. 11a and lib are MR images of the left flank and thigh of a rat in 3-D bright-pixel reconstruction before (Fig. 11a) and after (Fig. lib) an intravenous injection of MPEG (MW 5 kD) -poly-1-lysine(MW 25 kD) - DTPA(Gd) . Images were taken two weeks after injection of R3230 mammary adenocarcinoma cells into the left flank of the rat.
  • Fig. 12 is a drawing outlining the chemical synthesis of the graft-co-polymer, poly[( [N-(methoxy poly(ethylene)glycol) -o-succinyl]-l-lysyl)n-(N-succinyl- l-lysyl)m]lysine (i.e., MPEG-Poly(L-lysine)succinate or MPEG-PL-succinate) .
  • Fig. 13 is a drawing showing a reversible linkage between a cis-aq molecule and a portion of the graft-co- polymer.
  • the reversible linkage is an ionic (i.e., electrostatic) interaction between the cis-aq molecule and the graft co-polymer.
  • Part I outlines the hydration of cDDP resulting in the formation of cis-aq.
  • Part 2 shows the electrostatic interaction between cis-aq and the graft co-polymer.
  • Fig. 14 is a graph showing the binding of cDDP to MPEG-PL-succinate as determined by HPLC quantitative analysis. The graph is presented in Scatchard coordinates.
  • Fig. 15 is a graph showing the time dependent release of cDDP from the adduct MPEG-Poly(L- Lys)succinate/cDDP in the presence of saline (triangles) or bovine serum albumin (i.e. BSA) (circles) .
  • Figs. 17A and 17C are pictorial and Figs. 17B and 17D are graphical representations of the biodistribution of the graft co-polymer adduct MPEG-Poly(L- lysine)succinate/cDDP in NF13762-adenocarcinoma-bearing Fisher rats.
  • the distribution of MPEG-Poly(L-Lys)- succinate/cDDP (Figs. 17A and 17B) and Poly(L-Lys)- succinate (Figs. 17C and 17D) in NF13762-adenocarcinoma- bearing Fisher rats is shown after 24hr (solid bars, Fig.
  • FIG. 17B shows gamma camera images.
  • compositions of this invention include a polymeric carrier, a protective chain linked to polymeric carrier, and, optionally, a reporter group.
  • a polymeric carrier e.g., polyethylene glycol, polypropylene glycol, polyethylene glycol, polypropylene glycol, polyethylene glycol, polypropylene glycol, polyethylene glycol, polypropylene glycol, polyethylene glycol, polypropylene glycol, polyethylene glycol, polypropylene glycol dimethacrylate, poly(ethylene glycol)-2-propylene glycol dimethacrylate, poly(ethylene glycol)-2-propylene glycol dimethacrylate, poly(ethylene glycol)-2-propylene glycol dimethacrylate, polystyrenethacrylate, polystyrenethacrylate, polystyrenethacrylate, polystyrenethacrylate, polystyrenethacrylate, polystyrenethacrylate, polystyrenethacrylate, polysty
  • R ⁇ is (CH 2 ) 4 NHCO(CH 2 ) n COOCH 2 CH 2 A-B-OR 3 , where n is 2-6; A is [OCH 2 CH 2 ] ⁇ , where x is 15-220; B is [OCH 2 CH 2 ] ⁇ or [OCH(CH 3 )CH 2 ] y , where y+x is 17-220; R 2 is a chelating group; and R 3 is H, (CH 2 ) y CH 3 or (CH 2 ) COOH, where y is 0-7; or b) R ⁇ is -CH 2 (R g )NHCO(CH 2 ) nl COO((CH 2 ) n2 0) n3 CH 3 , where R g is -CH 2 CH 2 CH 2 -, -CO- or -CH 2 CO-; nl is 2 to 6, inclusive; n2 is 2 or
  • the polymeric carrier may be chosen from synthetic, non-proteinaceious polyamino acids, e.g., a linear, linked or branched polymer of a single amino acid species or of different amino acid species, e.g., regular or statistic block-co-polymers of polyamino acids, e.g, preferably linear poly-1- or poly-d-lysine, carboxylated or carboxymethylated poly-alpha, beta-(2-aminoethyl)-d, 1-aspartamide, poly-1-aspartic acid, or poly-glutamic acid.
  • the molecular weight of the polyamino acid carrier is preferably between 1,000 and 100,000 Daltons.
  • Polyamino acids with narrow molecular weight (MW) distributions are preferred to those with broad MW distributions.
  • the polyamino acids are linked with peptide bonds or, when obtained by condensation of two or more polyamino acid fragments or individual amino acids with cleaveable bonds, e.g., S-S bonds, which may be cleaved in vivo.
  • Polyamino acids may be prepared by chemical synthesis or by recombinant techniques, such as genetic engineering.
  • the polymeric carrier also may include polyethyleneimines, e.g., branched amino-containing polymers or carboxylated polyethyleneimines, i.e., reacted with derivatives of carbonic acids; natural saccharides containing amino acids or carboxylic acids, e.g., galacturonic acid, glucuronic acid, mannuronic acid, hyaluronic acid, pectic acid, neuraminic acid, alginic acid, carrageenan; aminated, e.g.
  • polysaccharides or oligosaccharides linear or branched
  • carboxylated, carboxymethylated, sulfated or phosphorylated polysaccharides or oligosaccharides e.g., reacted with derivatives of carbonic, dicarbonic, sulf ric, aminosulf ric, or phosphoric acids with resultant linking of carboxylic, aminocarboxylic, carboxymethyl, sulfuric, amino or phosphate groups.
  • Such oligosaccharides may be obtained by chemical alteration of, e.g., dextran, mannan, xylan, pullulan, cellulose, chytosan, agarose, fucoidan, galactan, arabinan, fructan, fucan, chitin, pustulan, levan or pectin.
  • polysaccharides or oligosaccharides may be heteropolymers or homopolymers of monosaccharides, e.g., glucose, galactose, mannose, galactose, deoxyglucose, ribose, deoxyribose, arabinose, fucose, xylose, xylulose, or ribulose.
  • monosaccharides e.g., glucose, galactose, mannose, galactose, deoxyglucose, ribose, deoxyribose, arabinose, fucose, xylose, xylulose, or ribulose.
  • the polymeric carrier may be a linear, branched or dendrimeric polya idoamine; polyacrylic acid; polyalcohols, e.g., polyvinylalcohol and polyxylitol to which carboxylic or amino groups are chemically linked; or oligonucleotides.
  • the protective chain may be poly(ethylene glycol) (i.e. PEG), preferably the PEG is esterified by dicarboxylic acid to form a polyethylene glycol monoester; for example, methoxy poly(ethylene glycol) (i.e. MPEG) or a copyolymer of poly(ethylene glycol) and poly(propylene glycol) , preferably in a form of an ester with a dicarboxylic acid; methoxypolypropylene glycol; polyethylene glycol-diacid; polyethylene glycol monoamine; MPEG monoamine; MPEG hydrazide; or MPEG imidazolide, and derivatives of all of the above.
  • PEG poly(ethylene glycol)
  • MPEG methoxy poly(ethylene glycol)
  • MPEG a copyolymer of poly(ethylene glycol) and poly(propylene glycol)
  • the protective chain may be a block-co-polymer of PEG and another polymer, e.g., a polyamino acid, polysaccharide, polyamidoamine, polyethyleneamine or a polynucleotide (as described above under polymeric carriers) .
  • the blocks are preferably alternated to give a linear block-co-polymer.
  • the overall molecular weight of the protective chain is 500 to 10,000 daltons, inclusive.
  • the protective chain is preferably linked to the polymeric carrier by a single bond.
  • the reporter groups of the invention are preferably linked to a polymeric carrier but also may be linked to a protective chain.
  • the reporter groups include complexones, e.g., chelating molecules such as diethylenetriamine-pentaacetic acid (DTPA) , triethylenetetraminehexaacetic acid (TTHA) , ethylenediaminetetraacetic acid (EDTA) , l,2-diaminocyclohexane-N,N,N' ,N'-tetraacetic acid, N,N'-Di(2-hydroxybenzyl)ethylenediamine (HBED) , N-(2-hydroxyethyl)ethylenediaminetriacetic acid, nitrilotriacetic acid, ethylene-bis(oxyethylenenitrilo)tetraacetic acid (EGTA) , l,4,7,10,-tetraazacyclododecane-N,N' ,N' ' ,N' '-te
  • the paramagnetic elements include transitional metals or lanthanides, e.g. elements with atomic numbers 21-29, 42, 44, 57-71, preferably gadolinium (III) , dysprosium (III) , holmium (III) , europium (III) , iron (III) , or manganese (II) .
  • transitional metals or lanthanides e.g. elements with atomic numbers 21-29, 42, 44, 57-71, preferably gadolinium (III) , dysprosium (III) , holmium (III) , europium (III) , iron (III) , or manganese (II) .
  • the radionuclides include alfa-,beta- and gamma- emitters, preferably gallium 67, indium 111, technetium 99m, chromium 51, cobalt 57, molibdenium 99, molecules, e.g. , tyrosine and p-oxybenzoic acid, linked to isotopes of iodine, e.g., iodine 131.
  • the reporter group may also include fluorine-containing molecules, e.g., fluorocarbons.
  • the reporter group may- also include therapeutic agents, e.g., cytostatics, antibiotics, hormones, e.g., growth factor, analgesics, psychotropic, antiinflammatory, antiviral, antifungal drugs or ly phokines, e.g., interleukin 2.
  • the therapeutic agents are preferably linked to a carrier with detachable or semistable bonds.
  • the reporter group may also include a particle, or colloidal particle, or colloidal precipitate of oxides, sulfides and/or hydroxides of transitional elements and lanthanides with atomic numbers 21-29, 42, 44, 57-71, or silicon oxide colloids or polymers containing silicon or polymers of atoms of sulfur, carbon, or silicon.
  • the particle or particles may be contained as an integral part of, or may be surrounded by, a semi-permeable membrane.
  • the compositions may also include additional reporter groups which may be chosen from (CH2) COOH, where p is between 0 and 7, inclusive; pyridyldithioacyl groups, e.g., N-(2-pyridyldithio)propionyl groups; N-hydroxysuccinimidyl, N-hydroxysulfosuccinimidyl, imidazolyl, benzotriazolyl, aminoalkyl, aminoacyl, aldehyde, thioalkyls, thiolanes, haloid acyl, haloid alkyl, or haloid phenyl; diazo- and hydrazo-, e.g.
  • 4-hydrazinoxyethyl, 4-hydrazinobenzyl, diazirinyl, azidophenyl, or azidoalkyl groups are linked to the polymeric carrier and/or to the protective chains, and are needed for conjugating or linking other ligands, e.g., a targeting group, capable of interacting with cell surface receptors, proteoglycans, adhesion molecules, ion channels or enzymes, to the compositions of this invention.
  • the targeting group may include antibodies; fragments of antibodies; chimeric antibodies, where said antibodies are polyclonal or monoclonal; enzymes; quasi substrates of enzymes; lectins; or saccharide ligands of lectins detachably or nondetachably linked to the composition. Synthesis of the composition
  • compositions of this invention may be synthesized using any one of the following methods (See Fig. 1) .
  • An example of synthesis using poly-1-lysine as a polymeric carrier, MPEG as a protective chain, and a complexone as a reporter group is provided.
  • This synthetic composition is especially suitable as a macromolecular contrast agent.
  • compositions may be prepared in two stages by first reacting polyamino acid with activated MPEG analogs, and then reacting this reaction mixture with an activated chelating compound. This procedure is preferred when poly-1-lysine is used as the polymeric carrier (See Fig. 1) .
  • e-amino groups of poly-1-lysine were reacted with activated derivatives of carboxylated MPEG, e.g., acid chlorides, anhydrides, mixed anhydrides, nitrenes, isothiocyanates and imidazolides, and activated esters, e.g hydroxysuccinimide, hydroxysulfosuccinimide, p- nitrophenyl, benzotriazolide.
  • the chelating molecule is brought into reaction with the remaining amino groups, either in activated form, e.g., anhydride, mixed anhydride, or isothiocyanate, or in a non-activated form. If the chelating molecule is in the non-activated form, it is activated to obtain an activated ester in the presence of succinimide or sulfosuccinimide and carbodiimide and is then brought into reaction with the remaining amino groups.
  • the reaction may be preceded with an additional chemical modification of the polyamino acid backbone or MPEG chains which are not limited to reactions resulting in the formation or elimination of at least one chemical bond.
  • the sequence of chemically linking the protective chains and a reporter group to a polymeric carrier may be reversed, i.e., linking of a reporter group preceeds linking of protective chain(s) to the polymeric carrier, but preferably, the reporter group is used as a monofunctional activated analog, i.e., one molecule of activated reporter group forms only one covalent linkage with a polymeric carrier.
  • compositions also may be synthesized using standard peptide synthesis protocols with modified amino acid precursors such as MPEG-amino acid and complexone- amino acid.
  • modified amino acid precursors such as MPEG-amino acid and complexone- amino acid.
  • moieties of complexone and PEG may be alternated in a controllable manner.
  • Oligomers of PEG-polya ino acids may be conjugated with oligomers of complexone-amino acids to form a block- co-polymer.
  • carboxylated carriers such as carboxylated saccharides, or polyaminoacids with carboxy groups in their radicals, such as poly-1-aspartic acid
  • the polymeric carrier is preferably activated in the presence of carbodiimide and sulfosuccinimide, as described in Example 2 for DTPA, and then reacted with aminated protective chains, such as MPEG monoamine at pH 7-9.
  • aminated protective chains such as MPEG monoamine at pH 7-9.
  • the linking of complexone or chelate is then achieved preferably by carbodiimide condensation.
  • compositions of this invention preferably have a non-proteinaceous polyamino acid molecule serving as a carrier of covalently attached activated analogs of linear or branched chelating molecules, to which a MR reporter cation is linked, i.e., ionically chelated.
  • the carrier forms a single chemical entity with protective chains of MPEG.
  • the synthetic route of preparing the compositions of this invention includes covalent modification of the polyamino acid carrier.
  • Conjugation of 1,1'- carbonyldiimidazole-treated MPEG to aminogroups requires high excesses of the modifier, e.g., activated MPEG, which leads to the formation of semi-stable gels since the solubility of polyamino acids in the presence of MPEG is reduced.
  • the procedure for preparing N- hydroxysuccinimidyl MPEG-succinate described in Scheme 1 gives a product with a highly activated ester content, e.g., greater than 75%, which is advantageous for preparing the compositions of this invention.
  • Linking MPEG to the polymeric carrier e.g., polyamino acid
  • MPEG chains prevent the formation of by-products because they create a steric barrier for cross-linking the reagent. Therefore, the formation of high-molecular weight products can be controlled, which makes the synthetic steps predictable. As a result, a homogenous preparation is obtained with a narrow molecular weight distribution.
  • the polymeric carriers preferably contain peptide bonds. The same bonds are involved in conjugating a chelating molecule with reactive groups of the amino acid radicals.
  • compositions are potentially biodegradable by various animal non-specific peptidases.
  • elements of polymeric carrier or protective chains or reporter groups could be linked together by a semistable linkage, such as S-S bonds. Small amounts of trapped compositions may be removed from the body by degradation to smaller fragments.
  • activated PEG derivatives may be used for the preparation of the compositions thus making them either virtually undegradable or, on the contrary, labile.
  • labile compositions are undesirable, since detaching MPEG will result in more extensive accumulation of the contrast agent compositions in the reticuloendothelial system.
  • compositions of this invention require the presence of an MR reporter group, such as a paramagnetic cation, e.g., gadolinium (III).
  • a paramagnetic cation e.g., gadolinium (III).
  • the transchelation technique developed for this experiment is based on an embodiment of Harris et al., J. Polym . Sci . , 22:341-52, which is incorporated herein by reference.
  • Gd-citrate to prevent the contact of the contrast agent with gadolinium oxides, used previously by Griess et al., U.S. Pat. 4,647,447, or gadolinium chloride, used previously by Bardy et al. U.S. Pat. 4,804,529.
  • the gadolinium citrate easily forms contaminants such as colloidal hydroxides at pH values greater than 6.5, which is within the range of optimal pH values for the NMR contrast agents of this invention.
  • a special purification step e.g., an anion-exchange chromatography step, allows the separation of Gd-labeled MPEG-PL-DTPA (Gd) from possible anionic contaminants, e.g., MPEG-PL-DTPA(Gd) with a low degree of substitution of amino groups with MPEG or small amounts of PL-DTPA(Gd) .
  • the protective chains, e.g., MPEG, of this invention do not react with the C3 component of complement which is a distinct advantage over previously known agents, e.g. dextran-DTPA(Gd) , which are known to react with the C3 component of complement.
  • MPEG prevents the exposure of chelating groups and paramagnetic cations to receptor cells, e.g., glomerulonephral phagocytes, capable of recognizing them. MPEG also forms a steric barrier which prevents rechelation of Gd cations by serum proteins such as transferrin. The compositions of this invention also prevent possible delayed toxic effects of re-chelated gadolinium.
  • MPEG conjugation lowers the toxicity of the composition of this invention by preventing significant accumulation of the chelating polymer in the liver and spleen.
  • Acute toxicity studies of the compositions of this invention have indicated no apparent toxicity in mice at concentrations exceeding 10-35 times the optimal doses. Histological examination of tissues of these mice have shown no deviations from control animals.
  • the blood half-life of the compositions of the invention was determined in rats.
  • the radioactive and paramagnetic contrast agents were incorporated into the composition prepared according to Examples 1 and 3 in order to accurately determine its pharmacokinetic characteristics in vivo.
  • the rats were visualized at different time points using a gamma camera to follow the distribution of the composition. As indicated by the data presented in Fig.
  • the blood half-life of the disclosed contrast agent was equal to 24 hours for MPEG(MW 5 kD)-poly-l-lysine(MW 53.5 kD)-DTPA labelled with [ 11:L In] and saturated with gadolinium, while a smaller contrast agent MPEG(MW 2 kD) -poly-l-lysine(MW 25 kD)-DTPA labelled with [ l ⁇ :L In] and saturated with gadolinium, was removed from the blood at a faster rate with the t 1 2 being 6 hours.
  • ELISA enzyme-linked immunoadsorbent assay
  • compositions of this invention have demonstrated a surprisingly high capacity, e.g., up to 13% by weight, for gadolinium and exceptionally high Rl/Gd atom, e.g., 20 mM-1 sec-1.
  • Preliminary experiments showed that high-quality angiogra s could be obtained when Tl values of blood are decreased at least 5-fold as a result of the injection of the contrast agent.
  • the Gd concentration which allows a 5-fold decrease in Tl corresponds to ca. 300 nmol. Gd/ml of blood (See Fig. 5). In a typical human study this corresponds to an injection of ca. 20 ⁇ mol Gd/kg of total body weight, which is 5- fold lower than for Gd-DTPA dimeglumine, which is a frequently used MR contrast agent.
  • rats were injected with 20 ⁇ moles Gd/kg of MPEG(MW 5 kD)-poly-1-lysine(MW 25 kD)-DTPA(Gd) (MPEG squares) or with 50 ⁇ moles Gd/kg of polylysine(MW 25 kD)-DTPA(Gd) (PL-Gd-DTPA, diamonds (See Fig. 7).
  • MPEG MPEG squares
  • PL-Gd-DTPA polylysine(MW 25 kD)-DTPA(Gd)
  • the 3-D bright-pixel reconstructions of vessel maps provided a very high vessel/background signal ratio, eliminating the need for background subtraction. Contrary to known constrast agents, the compositions of the invention injected at 30 ⁇ moles Gd/kg total body weight surprisingly resulted in resolution of submillimeter vessels having an internal diameter of less than 1 mm.
  • one rat was intravenously injected with MPEG-poly-l-ly ⁇ ine-DTPA(Gd) (20 ⁇ mol Gd/Kg total body weight) (left image) and one rat was intravenously injected with gadopentate dimeglumine (100 ⁇ mol Gd/kg, from Magnevist®, Berlex Labs) (right image).
  • Gd-DTPA MPEG-poly-l-ly ⁇ ine-DTPA
  • the ratios were 2.0 for Gd-DTPA and 5.8 for the MPEG(MW 5kD)-poly-l-lysine(MW 25 kD)-DTPA(Gd) at a p-value less than 0.001 (See Fig 9).
  • Gd-DTPA initially yielded a small increase in vessel contrast.
  • contrast is lost.
  • the MPEG derivative compositions of the invention because of their unique vascular distribution, consistently resulted in high ratios.
  • the images were taken on a Signa using a 5 inch surface coil (See Fig. 9) .
  • Imaging experiments with rabbit and minipig (body weight 40 kg) thorax were performed demonstrating the feasibility of visualizing the pulmonary and coronary arteries using the compositions of this invention (See Figs. 10a and 10b) .
  • a rabbit was intravenously injected with MPEG(MW 2 kD)-poly-l- lysine(MW 41 kD)-DTPA(Gd) (20 ⁇ mol Gd/ml) .
  • the images were taken 20 minutes after injection on a Signa using a 5 inch surface coil.
  • compositions of this invention to reveal abnormalities of vessels in experimentally induced pathological conditions was tested in rabbits and rats.
  • TOF Time of Flight
  • MR angiography the narrowing of the femoral artery at the site of experimental stenosis could be reliably visualized.
  • rats with R3230 mammary adeno carcinoma were used for visualization of vessel abnormalities in tumor progression.
  • the MR images of the left flank and thigh of a rat are shown before (See Fig. lla) and 20 minutes after (See Fig.
  • compositions of this invention may be used for detection of both neoplasia and tumor neovascularity which is important in clinical practice for staging and surgical planning.
  • compositions of the invention were prepared from a wide range of drugs that can be used to investigate n vivo gamma imaging; biokinetics; immune response; and magnetic resonance imaging.
  • Sprague-Dawley rats (200-250 g) were injected into tail vein using a 26 gauge needle with 1-10 mg/0.5 ml of product I or III, labeled with [ 11:L In] and Gd, as described in Example 6. Images on a gamma-camera (from Ohio Nuclear) using parallel medium-energy collimator were obtained 30, 60, 120 minutes, and 24 and 70 hours after injection.
  • the Biokinetics of Gd-and [ 111 In] labeled product (III or I) was studied using Sprague-Dawley rats ranging from 230-250 g. The animals were injected in the tail vein with 1-10 mg of polymer (60-70 ⁇ Ci/kg, 2 ⁇ m/kg Gd) using a 26 gauge needle under ether anesthesia. Little variation in kinetics was detected within these dose limits.
  • the biodistribution of labeled product was determined in 16 organs, i.e., organ tissues, by measuring radioactivity at each time point indicated on graphs. Two rats were used for each point (See Fig. 3).
  • ELISA plates were coated with ovalbumin- DTPA(Gd) , ovalbumin-MPEG, BSA or acetylated poly-1- lysine (MW 70,000). Only wells of the plate coated with ovalbumin-DTPA(Gd) showed specific binding of mouse immunoglobulins.
  • compositions of this invention may be used in medical imaging, and administered intravascularly or by bolus-injection.
  • the vascular images are enhanced due to changes of blood relaxivity or radioactivity.
  • the contrast agents may be used for the improvement of vascular images of large vessels, e.g., arteries and veins, or to visualize small vessels, e.g., submillimeter capillaries. The resolution of the images is increased by providing more detailed information.
  • the contrast agents may be used for vascular anatomy mapping, determination of vessel stenosis, abnormal vascularity, e.g., neovascularity, normal perfusion, perfusion defects, or functional imaging of the brain.
  • compositions of this invention may also comprise a therapeutic agent, e.g., one or more species of cytostatics, analgesics, antiinflammatory, antiviral, antifungal or psychotropic drugs.
  • a therapeutic agent e.g., one or more species of cytostatics, analgesics, antiinflammatory, antiviral, antifungal or psychotropic drugs.
  • the compositions of this invention which include therapeutic agents are beneficial because the prolonged circulation of the composition in the blood substantially prolongs the therapeutic effect of the therapeutic agent.
  • the therapeutic agent should slowly detach or leave the polymeric carrier. This may be achieved by detachably linking or positioning a semi-permeable membrane around the carrier to form a vesicle, allowing the drug concentrated in the vicinity of polymeric carrier to slowly diffuse through the membrane into the intravascular space.
  • the compositions of this invention which include therapeutic agents may be administered intravascularly or by bolus-injection.
  • compositions of this invention are described in the following Examples and Experimental section which form embodiments of the present invention and should not be regarded as limiting the scope of invention.
  • a suspension of a cyclic anhydride of DTPA (0.5 g/ml in DMSO) was prepared by adding 200 ⁇ l portions (1.5 g of cDTPA total) to the solution of PL and MPEG, and the pH was adjusted to 8 with 5 N NaOH after each addition. The amount of titratable aminogroups was checked again and no free aminogroups were detected.
  • MPEG-poly-1-lysine-DTPA Dilute the reaction mixture of MPEG-poly-1-lysine- DTPA(MPEG-PL-DTPA) to 300 ml with 0.2 M sodium citrate (pH 6.), filter through .45 ⁇ nylon filter and dialyze in a flow-through cell using a membrane with cut-off of 100 kD (for globular proteins) . Concentrate to 30-50 ml and dilute to 300 ml with citrate. Repeat the procedure 2 times using water instead of citrate in the last stage. Concentrate the solution to 15 ml, and lyophilize. Alternatively, the sample may be filtered through sterile 0.2 ⁇ m membrane and stored at 4°C. A table of the theoretical and actual chemical analysis is presented below:
  • a suspension of cyclic anhydride of DTPA ( 0.5 g/ml in DMSO) by adding 200 ⁇ l portions (1.5 g of cDTPA total) to the solution of MPEG-PL and adjust the pH to 8 with 5 N NaOH after each addition.
  • the solution may be prepared by mixing of 2.5 mmol of DTPA, 0.5 mmol N-hydroxysulfosuccinimide (pH 4) and 0.5 mmol ethyl dia inopropylcarbodiimide in 50 ml of water. The solution is then mixed for 3 min and added to the mixture the solution of MPEG-PL (pH 8) . Check the amount of titratable aminogroups. (No titratable amino groups were detected) .
  • Example 4 Synthesis of MPEG-poly-1-lvsine (MW 53.5 kD)- DTPA, Product IV Prepare according to the procedures Examples 1 and 2, using poly-1-lysine with a mean MW of 87,400 and MPEG (MW 5000)succinyl succinate.
  • Example 5 Synthesis of MPEG-poly-1-lysine(69)- (dithio)propionylpolv-l-lvsine-DTPA, Product V Dissolve 50 mg of N-e-benzoyloxycarbonyl-poly-1- lysine in 3 ml of dimethylforma ide and treat with 10 mg of N-succinimidyl 3-(2-pyridyldithio)propionate in the presence of 20 ⁇ l of triethylamine. Incubate the product overnight and precipitate by the addition of 20 ml of water. Freeze-dry the precipitated product and divide into two equal parts.
  • Example 9 The purification of labeled products I. II, III or IV
  • gadolinium or [ 1:L1 In] and gadolinium labeled products was prepared at 50-100 mg of polymer/ml of 5 mM sodium citrate (pH 6) .
  • the Gd content was determined titrametrically, (as in Korbl, J. and Pribil, R. , Chemist-Analyst 45:101-103 (1956) , or by plasma emission spectroscopy (from Korbl, J. and Pribil, R. , Chemist-Analyst 45:101-103 (1956) , or by plasma emission spectroscopy (from Korbl, J. and Pribil, R. , Chemist-Analyst 45:101-103 (1956) , or by plasma emission spectroscopy (from
  • Graft-co-polymers of the invention include a central carrier chain, a protecting group, and, optionally, a reporter group. Each group is linked together and is capable of forming reversibly linkages with a platinum(II) compound.
  • a reversible linkage between the graft co-polymer and a platinum(II) compound includes, but is not limited to 1) the formation of hydrogen bonds, 2) the formation of bonds with aguated platinum(II) compounds, 3) the formation of coordination bonds with the platinum atom (charged or neutral) and 4) electrostatic interactions, particularly with chemical groups of the graft co-polymer which include a carbonyl group, for example carboxylic acid groups.
  • platinum (II) compounds The chemical bonds formed between platinum (II) compounds and amino acids have been investigated (Appleton and Hall J. Chem . Soc. Commun . 493 911 (1983) and references therein) .
  • the platinum(II) compound may be present as an electroneutral and/or positively charged (aquated) form.
  • a graft-co-polymer adduct from a polymeric carrier containing amino groups generally involves three synthetic stages: 1) covalent modification of a backbone carrier with protective chains; 2) modification of the product with negatively charged groups, for example, modification with succinic acid; and 3) incubating the co-polymer and the platinum(II) compound together to achieve formation of a graft co- polymer adduct (see Figs. 12 and 13).
  • Preparation of an adduct by starting with negatively charged polymeric carrier does not include modification with negatively charged groups and thus includes only the first and third stage.
  • a graft-co-polymer was prepared by obtaining a carboxylated derivative of methoxy poly(ethylene glycol) (MPEG) (I), and reacting it with sulfosuccinimide in the presence of carbodiimide (II) reacting polyamino acid with activated MPEG analogs (III) , and then reacting this mixture with an excess of dicarboxylic acid anhydride. This procedure was preferred when poly-1-lysine was used as the backbone.
  • MPEG methoxy poly(ethylene glycol)
  • the nucleophilic epsilon-amino groups of poly-1-lysine were also reacted with activated derivatives of carboxylated MPEG, e.g., acid chlorides, anhydrides, mixed anhydrides, nitrenes, isothiocyanates and imidazolides, activated esters, e.g hydroxysuccini ide, hydroxysulfosuccinimide, p-nitrophenyl, benzotriazolide (not shown) .
  • the dicarboxylic acid used can be in activated form, e.g., anhydride, mixed anhydride, isothiocyanate, succinimide or sulfosuccinimide.
  • the reaction may be preceded with additional chemical modification of the polyamino acid backbone.
  • the sequence of chemical linking of protective chains and an agent to a polymeric carrier may be reversed, i.e. linking of an acid preceeds linking of protective chain(s) to a polymeric carrier.
  • the first stage of synthesis resulted in the formation of a graft-co-polymer where approximately 15-30% of onomeric residues of the polymeric carrier here modified with protective chains.
  • the second and third stages yielded a graft co-polymer where generally all monomeric residues that were not linked to protective chains were modified with negatively charged moieties.
  • the fourth stage (Fig. 13) generally yielded a product having more than 0.1% of platinum by weight.
  • the adduct product had between 1% and 30% platinum by weight, inclusive, the majority of platinum (more than 50% of total content) being capable of dissociating from the graft co-polymer.
  • the graft co-polymer (i.e., without a reversibly bound platinum(II) compound) has a molecular weight between 50 and 1500 kDa.
  • the molecular weight of the adduct i.e., graft co-polymer and cDDP
  • the graft co-polymer adduct may be purified in a form which elutes as a single peak on a standard size-exclusion column.
  • Graft co-polymers of the invention may be synthesized using the following methods.
  • the synthesis of the graft co-polymer poly[([N-(methoxy poly(ethylene)glycol)-o-succinyl]-1-lysyl)n-(N-succinyl- l-lysyl)m]lysine includes poly-1-lysine, as an examplary polymeric carrier, methoxypolyethyleneglycol as an examplary protective chain, and succinate as an examplary reporter group.
  • Preparation of the graft co-polymer adduct proceeds by incubation of cDDP with the graft co- polymer in water or water/DMF mixtures. cDDP binds spontaneously to the graft co-polymer.
  • a graft co-polymer adduct is especially suitable as a macromolecular contrast agent.
  • the combination was mixed at 100°C for an additional 4 hrs under nitrogen.
  • the reaction mixture was cooled to 60°C and transferred to an apyrogenic 1-neck 1 L flask.
  • Dioxane was removed by using a rotary evaporator, mixing the residue with 200 ml of chloroform, filtering through glass fiber filters, and cooling on ice and filter again.
  • Chloroform was removed at 40°C on rotary evaporator, then, 300 ml of ethanol was added to the residue. 4 g of activated charcoal was added and the solution boiled with a reflux for 1 h.
  • the mixture was filtered, then 300 ml of ethyl acetate was added to the mixture; the mixture was then left at 4°C for 24 h.
  • the precipitate was dissolved in 800 ml of ethyl alcohol and mixed with lOOg of ethanol-washed AG50 W-X8 resin. The resin was filtered and concentrated by using a rotary evaporator. 400 ml of ethyl acetate was added and the material transfered into Erlenmeyer flasks and kept at -10°C for 4 hrs. The resulting material was filtered and the precipitate dried in a vacuum. The weight of the resulting product was 44g. The yield of purified MPEG- succinate was 57% of theoretical yield.
  • 0.1 M carbonate buffer was prepared by disolving 4.2 g of sodium bicarbonate in water, after which, 20 ⁇ l of 50% NaOH solution was added. The solution was filtered through a sterile 0.4 ⁇ m filter. A solution was prepared of lg of poly-l-lysine/175 ml of 0.1 M carbonate buffer and 50 ⁇ l was withdrawn for amino group determination.
  • Example 12 Synthesis of PL-succinate: A solution of lg of poly-l-lysine/175 ml of 0.1 M carbonate buffer was prepared and a 50 ⁇ l aliquot was withdrawn for amino group determination. 1 g of succinic anhydride was added and dissolved in 10 ml of dimethylsulfoxide, which was added to the reaction mixture dropwise. The pH was kept at 8 by addition of a 5 N NaOH solution. The reaction mixture was stirred for 4 hrs at room temperature. A 50 ⁇ l aliquot was removed for amino group determination. The amino groups were determined by trinitrobenzene sulfonic acid (TNBS) titration. The assay for amino groups gave 100% of amino group substitution in comparison to the initial poly-1-lysine.
  • TNBS trinitrobenzene sulfonic acid
  • the solution was filtered through a sterile apyrogenic membrane.
  • the solution was diluted with 100 ml of sterile apyrogenic water and transfered into a 300ml diafiltration cell equipped with a YM100 membrane.
  • the cell was pressurized by using a nitrogen source and concentrated to 30 ml at 25 psig.
  • the contents were diluted with sterile apyrogenic water to 300 ml and concentrated again.
  • the procedure i.e. concentration/dilution
  • the purity was analyzed by using size- exclusion HPLC.
  • the solution was transferred to an autoclaved lyophilization flask, frozen in liquid nitrogen and lyophylized.
  • Aqueous Solution A solution of MPEG-PL- succinate or PL-succinate was prepared in water at a concentration of 20 mg/ml. A suspension of 12 mg/ml cDDP was dissolved in water. 1 ml of the cDDP solution was combined with 1 ml of polymer solution and stirred overnight at 40°C. Any unsolubilized cPPP was removed by filtration. In order to purify the adduct, the mixture was loaded onto a spin-column filled with Sephadex G-25m (10x1 cm) . The eluate was collected after centrifuging at 800g for 5 min.
  • the non-bound cDDP was determined by a standard o-phenylenediamine assay Schechter et al., Cancer Immunol . Immunother 25, 225 (1987). The total amount of platinum in the adduct was determined by plasma adsorption spectroscopy.
  • ii) Water/Organic Solution A solution of MPEG-PL-succinate or PL-succinate was prepared in water at 100 mg/ml. A suspension of 16.5 mg/ml cDDP was prepared in dimethylformamide. 1 vol of the cDDP solution was combined with 3 vol of polymer solution and incubated overnight at 40°C. 2 vol of water was subsequently added.
  • the mixture was loaded onto a spin-column filled with Sephadex G-25m (10x1 cm) and the eluate collected after centrifuging at 800g for 5 min.
  • the amount of non- bound cisplatinum was determined by o-phenylenediamine assay.
  • the amount of total platinum was determined by plasma adsorption spectroscopy.
  • the calculated cDPP/ ⁇ uccinate ratio (8.5:1) in the purified adduct indicates that linkages between the protonated amino groups of cDDP (or cis-aq) and carboxylic groups of the graft co-polymer, as well as other non-covalent linkages, are present in the adduct.
  • the data indicate that the protective chain is involved in the stabilization of cDDP with the polymeric backbone.
  • Dissociation of the cDDP from the graft co-polymer was detected by dialysis against isotonic saline or against isotonic medium containing lOg/1 of serum albumin (Fig. 15) .
  • the latter experiment was designed to mimic the presence of plasma proteins in the bloodstream of a mammal. Plasma proteins, are capable of irreversible (i.e., covalent) binding of free (i.e., non-complexed) cDDP.
  • cDDP was released from the carrier with the half- time of 63h in saline.
  • BT-20 cells Human mammary adenocarcinoma cells (BT-20 cells) were plated in 96-well plates in medium (i.e. 10% FCS, DMEM) at a cell density of 350,000 cells/well. Free cDDP, a cDDP graft co-polymer adduct or cDDP linked to PL-succinate were each diluted serially with cell medium and incubated with the cells overnight at 37°C. Cytotoxicity was determined by a standard
  • [ 3 H]methylthymidine DNA incorporation assay For example, 10 ⁇ Ci of [ 3 H]methylthymidine were added per well and incubated with the cells for 3 hours. The cells were collected by harvesting on glass fiber membranes. The amount of bound radioactivity was determined on each membrane by standard scintillation counting.
  • MPEG-PL-succinate-cDDP adduct exhibited a long circulation time in the bloodstream, whereas PL- succinate-cDDP did not. 24h after i.v. injection of the DTPA-labeled co-polymer, 40% of it was found in the blood, whereas only 1% the MPEG-free adduct remained in blood. After 96h, most (>80%) of MPEG-PL-succinate-cDDP adduct had been removed from circulation. MPEG-free adducts accumulated in kidneys (15.0+1.2% dose/g) , whereas acumulation of MPEG-PL-succinate-cDDP was 5 fold lower (3.5+0.5% dose/g).
  • MPEG- PL-succinate/cDDP adduct has an advantageous pharmacological profile in terms of: 1) longer circulation in the blood stream; 2) lower accumulation in kidneys (lower chance of eliciting of nephrotoxicity) and; 3) higher accumulation in solid tumors.
  • Targeting of cDDP to the tumor could not be achieved with an adduct that included a polymer devoid of protective chains.
  • MPEG-PL-succinate was shown to be a high-capacity carrier for cDDP.
  • cDDP is a highly potent chemotherapeutic agent, one which nontheless exhibits significant systemic toxicity. Prolonged blood circulation of the adduct creates a circulating depot of reversibly bound cDDP.
  • the high cytotoxicity of the drug in vitro and in vivo indicates that graft co-polymer adducts which include a platinum(II) compound, particularly cDDP will be useful therapeutic agents in the treatment of human cancer.
  • graft co-polymer adducts which include a platinum(II) compound can be administered either alone or in combination with other chemotheraputic agents in order to treat human cancer.
  • an adduct consisting of a graft co-polymer and a platinum(II) compound other than cDDP can be made by following the above-described methods, except that the platinum(II) compound will be substituted for cDPP.
  • the amount of platinum(II) compound that can be combined with a graft co-polymer is within the range disclosed for cPDP.
  • the amount of platinum(II) compound associated with a graft-co-polymer can be assayed by plasma absorption spectroscopy and the adduct can be purified and tested by any of the in vitro or in vivo methods disclosed herein.
  • adducts provided herein can be administered either alone or formulated into pharmaceutical compositions by admixture with pharmaceutically acceptably nontoxic excipients and carriers.
  • An adduct of the invention may be prepared for use in parenteral administration, particularly in the form of liquid solutions or suspensions; for oral administration, particularly in the form of tablets or capsules; or intranasally, particularly in the form of powders, nasal drops, or aerosols.
  • An adduct of the invention may be conveniently administered in unit dosage form and may be prepared by any of the methods well known in the pharmaceutical art, or example, as described in Remington's Pharmaceutical Sciences (Mack Pub. Co. , Easton, PA, 1980) .
  • Formulations for parenteral administration of an adduct of the invention may include as common excipients sterile water or saline, polyalkylene glycols such as polyethylene glycol, oils of vegetable origin, hydrogenated naphthalenes and the like.
  • Formulations for parenteral administration may also include glycocholate for buccal administration, methoxysalicylate for rectal administration, or citric acid for vaginal administration.
  • An adduct of the invention can be employed as the sole active agent in a pharmaceutical or can be used in combination with other chemotherapeutic agents such as cDDP, carboplatin, doxorubicin, or cyclophosphamide.
  • the chemotherapeutic agent may be provided as a biocompatible, biodegradable lactide polymer, lactide/glycolide co-polymer, or polyoxyethylene- polyoxypropylene co-polymers, each of which may serve as useful adjuncts to therapy.
  • Other useful excipients to control the release of the chemotherapeutic agent include parenteral delivery systems such as ethylene-vinyl acetate co-polymer particles, osmotic pumps, implantable infusion systems, and liposomes.
  • an adduct of the invention described herein in a therapeutic composition will vary depending upon a number of factors, including the adduct to be administered, the chemical characteristics (e.g., hydrophobicity) of the adduct employed, and the route of administration.
  • an adduct of the invention may be provided in an aqueous physiological buffer solution containing about 0.1 to 10% w/v compound for parenteral administration. Typical dose ranges are from about 1 ⁇ g/kg to about 1 g/kg of body weight per day; a preferred dose range is from about 0.01 mg/kg to 100 mg/kg of body weight per day.
  • the preferred dosage of drug to be administered is likely to depend on such variables as the type and extent of progression of the cancer, the overall health status of the particular patient, the relative biological efficacy of the adduct selected, the formulation of the compound excipients, and its route of administration, and whether a chemotherapeutic drug is chosen for ad unctive therapy.
  • All publications and patent applications mentioned in the specification are indicative of the level of skill of those in the art to which this invention pertains. All publications and patent applications are incorporated herein by reference to the same extent as if each individual publication or patent application were specifically and individually stated to be incorporated by reference.

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Abstract

L'invention concerne un produit d'addition copolymère greffé biocompatible comprenant un véhicule polymère, une chaîne de protection liée au véhicule polymère, un groupe rapporteur lié au véhicule polymère ou au véhicule polymère et à la chaîne de protection, et un composé de platine (II) lié de manière réversible. L'invention concerne également un procédé de traitement d'une maladie chez un patient, en particulier le cancer, en lui administrant une quantité thérapeutiquement efficace du produit d'addition, et peut comprendre l'exploration de l'organisme du patient à l'aide d'une technique d'imagerie permettant d'obtenir une image visible de la distribution du produit d'addition.
PCT/US1995/007329 1994-06-27 1995-06-07 Produits d'addition copolymeres greffes de composes de platine (ii) WO1996000079A1 (fr)

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997023240A2 (fr) * 1995-12-21 1997-07-03 Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts Conjugue comprenant un principe actif, un polypeptide et un polyether
WO1998047496A2 (fr) * 1997-04-18 1998-10-29 Access Pharmaceuticals, Inc. Composes a base de polymere et de platine
WO1998057669A1 (fr) * 1997-06-15 1998-12-23 Yeda Research And Development Co. Ltd. Images radiologiques de tumeur a l'aide d'un support dextrane de composes de platine
WO2001034206A3 (fr) * 1999-11-09 2002-02-07 Cmic Co Ltd Complexe contenant un acide nucleique
WO2004014973A2 (fr) * 2002-08-13 2004-02-19 Sirus Pharmaceuticals Ltd Polymere biodegradable

Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4278660A (en) * 1977-05-02 1981-07-14 Research Corporation Complex or salt of a platinum (II) compound and a nitrogen containing polymer
US4793986A (en) * 1987-02-25 1988-12-27 Johnson Matthey, Inc. Macromolecular platinum antitumor compounds
US4921944A (en) * 1986-06-10 1990-05-01 Uniwersytet Warszawski Method of pharmaceutic production
US4946954A (en) * 1989-01-17 1990-08-07 Georgetown University Platinum pharmaceutical agents
US5024998A (en) * 1987-12-30 1991-06-18 University Of Florida Pharmaceutical formulations for parenteral use
US5091521A (en) * 1986-09-08 1992-02-25 Behringwerke Aktiengesellschaft Cis-platinum complexes, a process for the preparation thereof, and pharmaceuticals containing these compounds
US5169933A (en) * 1988-08-15 1992-12-08 Neorx Corporation Covalently-linked complexes and methods for enhanced cytotoxicity and imaging
US5252713A (en) * 1988-09-23 1993-10-12 Neorx Corporation Polymeric carriers for non-covalent drug conjugation
US5420105A (en) * 1988-09-23 1995-05-30 Gustavson; Linda M. Polymeric carriers for non-covalent drug conjugation

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4278660A (en) * 1977-05-02 1981-07-14 Research Corporation Complex or salt of a platinum (II) compound and a nitrogen containing polymer
US4921944A (en) * 1986-06-10 1990-05-01 Uniwersytet Warszawski Method of pharmaceutic production
US5091521A (en) * 1986-09-08 1992-02-25 Behringwerke Aktiengesellschaft Cis-platinum complexes, a process for the preparation thereof, and pharmaceuticals containing these compounds
US4793986A (en) * 1987-02-25 1988-12-27 Johnson Matthey, Inc. Macromolecular platinum antitumor compounds
US5024998A (en) * 1987-12-30 1991-06-18 University Of Florida Pharmaceutical formulations for parenteral use
US5169933A (en) * 1988-08-15 1992-12-08 Neorx Corporation Covalently-linked complexes and methods for enhanced cytotoxicity and imaging
US5252713A (en) * 1988-09-23 1993-10-12 Neorx Corporation Polymeric carriers for non-covalent drug conjugation
US5420105A (en) * 1988-09-23 1995-05-30 Gustavson; Linda M. Polymeric carriers for non-covalent drug conjugation
US4946954A (en) * 1989-01-17 1990-08-07 Georgetown University Platinum pharmaceutical agents

Non-Patent Citations (7)

* Cited by examiner, † Cited by third party
Title
ANTI-CANCER DRUGS, Volume 4, issued 1993, MAEDA M. et al., "Antitumor Activity of a New Series of Platinum Complexes: Trans(+,-)-a, 2-Cyclohexanediamineplatinum (II) Conjugated to Acid Polysaccharides", pages 167-171. *
BIOCONJUGATE CHEMISTRY, Volume 4, issued 1993, NATHAN, A. et al., "Copolymers of Lysine and Polyethylene Glycol: A New Family of Functionalized Drug Carriers", pages 54-62. *
BIOMATERIALS, Volume 13, Number 4, issued 1992, IKE O, et al., "Controlled Cisplatin Delivery System Using Poly(D, L-Lactic Acid)", pages 230-234. *
CANCER CHEMOTHERAPY PHARMACOLOGY, Volume 24, issued 1989, SCHECHTER B. et al., "Blood Levels and Serum Protein Binding of Cis-Platinum (II) Complexed to Carboxymethyl-Dextran", pages 161-166. *
DUNCAN R. et al., "Polymers in Medicine", Published 1984, pages 51-101. *
JOURNAL OF BIOLOGICAL CHEMISTRY, Volume 252, Number 11, issued 10 June 1977, ABUCHOWSKI A. et al., "Alteration of Immunological Properties of Bovine Serum Albumin by Covalent Attachment of Polyethylene Glycol", pages 3578-3591. *
MAKROMOLECULAR CHEMISTRY, Volume 190, issued 1990, YOKOYAMA M. et al., "Molecular Design for Missile Drug: Synthesis of Adriamycin Conjugated with Immunoglobulin G Using Poly(Ethylene Glycol)-Block-Poly(Aspartic Acid) as Intermediate Carrier", pages 2041-2054. *

Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997023240A2 (fr) * 1995-12-21 1997-07-03 Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts Conjugue comprenant un principe actif, un polypeptide et un polyether
WO1997023240A3 (fr) * 1995-12-21 1997-10-09 Deutsches Krebsforsch Conjugue comprenant un principe actif, un polypeptide et un polyether
US6395254B1 (en) 1995-12-21 2002-05-28 Deutsches Krebsforschungszentrum Stiftung Des Offentlichen Rechts Conjugate comprising an active agent, a polypeptide and a polyether
WO1998047496A2 (fr) * 1997-04-18 1998-10-29 Access Pharmaceuticals, Inc. Composes a base de polymere et de platine
WO1998047496A3 (fr) * 1997-04-18 1999-02-11 Access Pharma Inc Composes a base de polymere et de platine
US5985916A (en) * 1997-04-18 1999-11-16 Access Pharmaceuticals, Inc. Polymer-platinum compounds
WO1998057669A1 (fr) * 1997-06-15 1998-12-23 Yeda Research And Development Co. Ltd. Images radiologiques de tumeur a l'aide d'un support dextrane de composes de platine
US6694171B1 (en) 1997-06-15 2004-02-17 Yeda Research And Development X-ray imaging of tumors with dextran carrier of platinum compounds
WO2001034206A3 (fr) * 1999-11-09 2002-02-07 Cmic Co Ltd Complexe contenant un acide nucleique
CN100417418C (zh) * 1999-11-09 2008-09-10 盛英三 含核酸的复合物
WO2004014973A2 (fr) * 2002-08-13 2004-02-19 Sirus Pharmaceuticals Ltd Polymere biodegradable
WO2004014973A3 (fr) * 2002-08-13 2004-09-23 Sirus Pharmaceuticals Ltd Polymere biodegradable

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