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WO2008101469A2 - Préparation pharmaceutique pour lutter contre des métastases - Google Patents

Préparation pharmaceutique pour lutter contre des métastases Download PDF

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Publication number
WO2008101469A2
WO2008101469A2 PCT/DE2008/000264 DE2008000264W WO2008101469A2 WO 2008101469 A2 WO2008101469 A2 WO 2008101469A2 DE 2008000264 W DE2008000264 W DE 2008000264W WO 2008101469 A2 WO2008101469 A2 WO 2008101469A2
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Prior art keywords
pharmaceutical preparation
phosphatidylcholine
liposomes
preparation according
peg
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German (de)
English (en)
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WO2008101469A3 (fr
Inventor
Reinhardt Zeisig
Iduna Fichtner
Jane Wenzel
Christian Nowak
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Epo Experimentelle Pharmakologie & Onkologie Berlin-Buch GmbH
Max Delbrueck Centrum fuer Molekulare in der Helmholtz Gemeinschaft
Original Assignee
Epo Experimentelle Pharmakologie & Onkologie Berlin-Buch GmbH
Max Delbrueck Centrum fuer Molekulare in der Helmholtz Gemeinschaft
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Publication of WO2008101469A3 publication Critical patent/WO2008101469A3/fr
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/66—Phosphorus compounds
    • A61K31/683—Diesters of a phosphorus acid with two hydroxy compounds, e.g. phosphatidylinositols
    • A61K31/685—Diesters of a phosphorus acid with two hydroxy compounds, e.g. phosphatidylinositols one of the hydroxy compounds having nitrogen atoms, e.g. phosphatidylserine, lecithin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes or liposomes coated or grafted with polymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes or liposomes coated or grafted with polymers
    • A61K9/1272—Non-conventional liposomes, e.g. PEGylated liposomes or liposomes coated or grafted with polymers comprising non-phosphatidyl surfactants as bilayer-forming substances, e.g. cationic lipids or non-phosphatidyl liposomes coated or grafted with polymers

Definitions

  • the invention relates to a pharmaceutical preparation for combating metastases based on liposomes. Areas of application of the invention are the medicine and the pharmaceutical industry.
  • the success of tumor therapy thus depends essentially on a comprehensive prevention of metastasis, which is triggered by circulating tumor cells. Metastasis can occur via two processes. On the one hand, a single circulating tumor cell undergoes a complex cascade of different, causally dependent adhesion processes before it can be fixed in a distal tissue 1 .
  • the initial step of adhesion is mediated by the endothelial receptor E-selectin, which recognizes the carbohydrate ligands sialyl Lewis "(sLe x ) and sialyl Lewis a (sLe a ) found on various metastatic cancer cells, through ligand-receptor interactions weak affinity binding of the tumor cell to the endothelium as a prerequisite for the subsequent steps ("rolling along” the membrane, release of further mediators, firm adhesion and invasion into the surrounding tissue). Finally, the tumor cell nests in the secondary tissue, initiates the formation of a neovascular blood system and proliferates 2 .
  • the spontaneous formation of tumor cell microthrombi is relevant for the metastasis process 3 .
  • these complexes can contribute to a local accumulation of the tumor cells, which in turn leads to metastasis through individual cells 4 .
  • activated platelets in particular attach to tumor cells and thus contribute to the shielding of immune reactions and the formation of microthrombi 5 .
  • targets that play an essential role in this process include the adhesion molecules on the endothelial surface and / or the ligands that are expressed by the tumor cell.
  • targets can be switched off 7 by competitive binding of suitable inhibitors (antibodies, carbohydrates) 6 or by inhibition of its expression with antisense or gene knockout strategies.
  • Other approaches are aimed at inhibiting angiogenesis. This also essential process of formation of new blood vessels to supply the newly forming metastases follows the adhesion cascade.
  • the heart of the invention is a pharmaceutical preparation comprising liposomal formulations containing at least one substance with antitumor effect and at least one substance with anticoagulant activity and optionally other substances.
  • An important starting point of the invention is the elimination of the formation of aggregates of tumor cells with each other or from tumor cells and platelets, which is considered an essential prerequisite of tumor cell engraftment. This is to be achieved by surface-modified, drug-containing liposomes. In addition, under certain conditions, the adhesion of the tumor cells to the blood vessel wall can be prevented by liposomes as an alternative step of a secondary tumor cell colonization.
  • the substances with antitumor action used in the context of the invention are in particular those substances which negatively influence or inhibit the formation, growth and / or spread of tumors.
  • alkyl phospholipids such as tetradecylphosphatidylcholine (TPC), hexadecylphosphatidylcholine (HPC), heptadecylphosphatidylcholine (HpPC), octadecylphosphatidylcholine (OPC), hexadecyl (1, 1-dimethyl-piperidin-4-yl) -phosphate (HPP), octadecyl (1, 1-dimethyl-piperidin-4-yl) -phosphate (OPP), hexadecylphosphatidylserine (HPS), hexadecylphosphatidylethanolamine (HPE) or edelfo
  • TPC tetradecylphosphatidylcholine
  • HPC hexadecylphosphatidylcholine
  • HpPC heptadecylphosphatidylcholine
  • OPC
  • substances with anticoagulant effect are used: substances from the drug group of aggregation inhibitors, such as dipyridamole, acetylsalicylic acid, ticlopidine, resveratrol, anticoagulants, such as heparin, low molecular weight heparin, heparinoids, clopidogrel, the vitamin K antagonists, such as warfarin or phenprocoumone.
  • substances from the drug group of aggregation inhibitors such as dipyridamole, acetylsalicylic acid, ticlopidine, resveratrol
  • anticoagulants such as heparin, low molecular weight heparin, heparinoids, clopidogrel
  • the vitamin K antagonists such as warfarin or phenprocoumone.
  • Particularly suitable substances with anticoagulant activity in the context of the invention have proven to be the active substances dipyridamole and acetylsalicylic acid.
  • the liposomal formulation used to implement the invention is constructed from known ingredients. It consists of base lipids and additionally preferably contains helper lipids, membrane stabilizers and charge carriers, whereby a particularly favorable stability of the preparation is achieved.
  • base lipids in the context of the invention are in particular soya phosphatidylcholine, egg phosphatidylcholine (EggPC), hydrogenated phosphatidylcholine (HePC), dimyristoyl-phosphatidylcholine (DMPC), (dipalmitoyl-phosphatidylcholine) (DPPC), distearyl-phosphatidylcholine (DSPC), stearyl palmitoyl-phosphatidylcholine (SPPC), palmitoyl-stearyl-phosphatidylcholine (PSPC), dioleyl-phosphatidylcholine (DOPC), palmitoyl-oleyl-phosphatidylcholine (POPC) or sphingomyelin.
  • EggPC egg phosphatidylcholine
  • HePC hydrogenated phosphatidylcholine
  • DMPC dimyristoyl-phosphatidylcholine
  • DPPC dimyristoyl-phosphati
  • Helper lipids in the context of the invention are preferably dioleyl phosphatidylethanolamine (DOPE) and dioleyl phosphatidylcholine (DOPC).
  • DOPE dioleyl phosphatidylethanolamine
  • DOPC dioleyl phosphatidylcholine
  • cholesterol is used as a membrane stabilizer for the realization of the invention.
  • the charge carriers used are preferably negative charge carriers, such as e.g. Dicetyl phosphate (DCP), stearylamine (SA), phosphatidic acid (PA), phosphatidylglycerol (PG) or monosialoganglioside.
  • DCP Dicetyl phosphate
  • SA stearylamine
  • PA phosphatidic acid
  • PG phosphatidylglycerol
  • the liposomal formulation can be adapted to the requirements of a targeting.
  • the substance used for modifying the surface of the liposomal formulation is preferably polyethylene glycol-phosphatidylethanolamine (PEG-PE).
  • PEG-PE polyethylene glycol-phosphatidylethanolamine
  • PEG sterols, sialyl Lewis x -Peg-PE, sialyl Lewis A -PEG-PE, sialyl Lewis x -Peg sterols or sialyl Lewis A -PEG sterols are also effective. These compounds increase the hydrophilicity of the liposomal surface, thereby sterically shielding the vesicles from the monocyte / phagocyte system and preventing the vesicles from being recognized as foreign bodies and eliminated by the immune system.
  • such a surface increases the affinity for platelets, whereby a nonspecific targeting effect is achieved.
  • specific ligands such as Lewis x or A specific targeting to the site of action (platelets, endothelial membrane) is further increased.
  • liposomal formulations in which the liposomes have an average diameter smaller than 400 nm and the size distribution has a polydispersity index smaller than 0.4 to reduce the elimination by the monocyte / phagocytic system, thereby causing accumulation in the aggregates Platelets and tumor cells is promoted.
  • the liposomes are preferably prepared by classical methods, ie via lipid film hydration to produce the multilamellar vesicles, then extrusion to obtain small vesicles of defined size.
  • the stock solutions of the individual substances are mixed together and a lipid film is prepared by evaporation or other suitable methods. From this one obtains by the application of known methods, e.g. Rehydration, multilamellar vesicles, which are then subsequently converted by suitable methods, preferably extrusion, into a homogeneous suspension of small vesicles having a diameter of about 100-150 nm.
  • known methods e.g. Rehydration
  • the pharmaceutical preparation is suitable for the manufacture of a medicament for the treatment of diseases such as thromboembolic disorders and coagulation disorders, preferably when metastatic tumor diseases occur in this context.
  • the pharmaceutical preparation for the treatment of metastatic liposomal-based tumors according to the invention is completely novel.
  • the combination liposomes contained in the pharmaceutical preparation are stable over a sufficient period of time, inhibit aggregate formation, inhibit the adhesion of tumor cells to the endothelial surface and endothelial components, influence the distribution of metastases in animal experiments, and restrict metastasis and thus the metastasis number and the metastasis Reduce total tumor weight.
  • the liposomes used have significant advantages over other drugs.
  • an accumulation in tumor cell platelet aggregates is achieved, whereby a targeted Release of the trapped drugs in close local proximity to the targets is achieved.
  • a second important advantage is the possibility of the simultaneous inclusion of several active ingredients with different active principles in the liposomes, whereby a targeted and simultaneous enrichment of all drugs (even with different pharmacokinetics of the individual components) is achieved in tumor cells and this can be attacked efficiently.
  • Novel and original to the combination liposomes according to the invention is that they have a modified surface and have included a cytostatic and an anticoagulant. This synergistically prevents both the binding to the endothelium (metastasis cascade) and the formation of tumor cell aggregates (embolic model). Despite the complexity of preparation of these vesicles, these liposomes can be prepared and modified in their properties (charge, stability and binding of ligands) and adapted to the requirements of a specific targeting.
  • This new drug system is that it can be used on a variety of tumors, thereby reducing the high metastatic mortality of tumors by synergistically inhibiting the process of metastasis through various mechanisms:
  • the pharmaceutical preparation according to the invention represents a completely novel approach to the inhibition of metastasis, which promises extraordinarily high efficacy through the complex and local approach.
  • the invention will be explained in more detail by exemplary embodiments.
  • Examples 1-6 It is shown in Examples 1-6 how various vesicles are prepared which have one or more active agents included in effective amount. This is followed by the characterization of physicochemical properties, storage stability and in vitro biological activity.
  • Example 1 Liposomes (vesicles with a diameter ⁇ 200 nm and uniform size distribution) are prepared by known methods.
  • a cytostatic, z.Bsp. included the alkylphospholipid octadecyl-1, 1-dimethylpiperidin-4-yl-phosphate (OPP), doxorubicin or mitoxantrone to specifically damage tumor cells.
  • OPP alkylphospholipid octadecyl-1, 1-dimethylpiperidin-4-yl-phosphate
  • AK anticoagulant
  • Size and size distribution quadsi-elastic light scattering
  • HPTLC current drug concentration
  • the following components are pipetted as stock solutions into a 50 ml round-bottomed flask: 31.2 ⁇ mol phosphatidylcholine (PC), 12 ⁇ mol octadecyl- (1.1-dimethylpiperidin-4-yl) -phosphate (OPP), 12 ⁇ mol cholesterol (CH), 2 , 8 ⁇ mol polyethylene glycol 2 ooo-phosphatidylethanolamine (PEG-PE), 6 ⁇ mol dicycetic phosphate (DCP) and 15.9 ⁇ mol dipyridamole (DIP).
  • PC phosphatidylcholine
  • OPP octadecyl- (1.1-dimethylpiperidin-4-yl) -phosphate
  • CH omol cholesterol
  • PEG-PE polyethylene glycol 2 ooo-phosphatidylethanolamine
  • DCP ⁇ mol dicycetic phosphate
  • DIP 15.9 ⁇ mol dipyridamole
  • the lipid film is prepared by evaporation of the solvent at 42 0 C in vacuo and then resuspended with 2 ml of PBS, pH 7.4 again.
  • the suspension becomes yellowish turbid and consists of MLV, which are shaken overnight.
  • the MLVs are then repeatedly forced through a mini extruder containing two pooled polycarbonate filters of 400 nm pore diameter. After 19-fold extrusion, completely large unilamellar vesicles (LUVs) with a mean diameter of 131 ⁇ 28 nm are present.
  • the suspension has a polydispersity index (PI) of 0.14 ⁇ 0.07.
  • the non-included DIP is separated by size exclusion chromatography on a Sephadex column (G50 fine).
  • the content of PC is determined by high performance thin-layer chromatography (HPTLC) and after column separation is 16.4 ⁇ 0.1 ⁇ mol / ml, while about 150 ⁇ 24 ⁇ g DIP /
  • the liposomes can be stored in the refrigerator for at least 8 weeks without their properties changing significantly (maximum size increase of 6% and a rise in the polydispersity index PI by a maximum of 30%).
  • liposomes having a total lipid content of 42.4 ⁇ mol / ml are prepared by a known method.
  • PC 6 .mu.mol PC
  • 12 .mu.mol CH 6 .mu.mol DCP
  • 2.8 .mu.mol PEG 8 .mu.mol dipyridamole
  • 4 mg mitoxantrone in the solvent mixture dichloromethane / methanol (7: 3, v / v) dissolved the solvent removed in vacuo to obtain a thin lipid film.
  • This is resuspended in PBS (without Ca, Mg), the suspension shaken overnight at room temperature to obtain multilamellar vesicles.
  • the liposomes are extruded 19 times by means of mini extruders equipped with two polycarbonate membranes with a pore diameter of 400 nm to obtain a uniform size distribution of the liposomes. Subsequently, the liposomes are passed through a Sephadex G50fine column to separate by size exclusion chromatography the unencapsulated DIP and mitoxantrone.
  • the size and polydispersity index (PI) of the liposomes is determined by PCS.
  • the mean size of the liposomes is 123 nm, the PI for these liposomes being 0.134.
  • the trapped MTX is determined by HPTLC. It is up to 850 ⁇ g / ml liposomal suspension for these liposomes. This corresponds to an inclusion rate of 20%.
  • liposomes having a total lipid content of 32 ⁇ mol / ml are prepared by a known method. For this purpose, first 15.6 mM PC, 6 mM CH, 3 mM DCP, 1.4 mM PEG and 8 mM dipyridamole are dissolved in a round bottom flask in an organic solvent and then the solvent is removed in vacuo to obtain a thin lipid film. This is in citrate buffer pH 3.8 resuspended, the suspension shaken overnight at room temperature
  • MLV multilamellar vesicles
  • the MLV are extruded 19 times by means of mini extruders equipped with two polycarbonate membranes with a pore diameter of 400 nm, to obtain small vesicles with a uniform size distribution.
  • Doxorubicin is encapsulated via the well-known pH-jumping technique.
  • 800 .mu.l of the liposome suspension are brought to a pH value of 7.6 by means of NaOH solution and subsequently heated in a water bath at 60 0 C. Then be
  • the size and polydispersity index (PI) of the liposomes is determined by PCS.
  • the mean size of the liposomes is 182 nm, with these liposomes having a value of 0.263.
  • the trapped doxorubicin is determined by HPTLC. It is up to 950 ⁇ g / ml liposomal suspension for these liposomes.
  • the effectiveness of the liposomes is characterized in vitro in various test systems.
  • the influence of liposomes on the adhesion of tumor cells and platelets is determined under static conditions in the binding assay.
  • the influence of liposomes on the aggregation behavior of tumor cells in the absence and presence of platelets, and the complex formation between tumor cells and platelets as an essential step of metastasis is used as test systems in vitro.
  • 100 ⁇ l of platelet-rich plasma (PRP) with a platelet count of 2.5 ⁇ 10 8 / ml are added per well to a 96-well plate and activated with 0.05 ⁇ thrombin / well. In some wells, the activation is omitted in order to be able to carry unactivated platelets as a control.
  • PRP platelet-rich plasma
  • Inhibition of aggregation by the combination liposomes can be achieved by 40%, while no inhibition is observed by control liposomes (see Figure 1).
  • DlP as a free drug inhibits platelet aggregation by 30% at a concentration of 10 nmol / ⁇ l (data not shown).
  • Example 5.2 Inhibition of the adhesion of platelets to immobilized tumor cells
  • the tumor cells and platelets are incubated with liposomes (100 nmol total lipid / well) or 10 ⁇ l of a 1 M EDTA solution / well as control.
  • the seeding and serial dilution of fluorescently labeled platelets into a black Maxi-Sorp plate (Nunc) and their activation by 0.05 ⁇ thrombin are performed to obtain a standard series for the quantification of bound platelets.
  • Lysis buffer Add 2.5 g sodium dodecyl sulfate (SDS) with 1 ml 10 N NaOH and make up to 50 ml with PBS, filter. Result: The activation-related increase in the adhesion of platelets to HT29 colon carcinoma cells ( Figure 2A) and to MT-3 breast cancer cells ( Figure 2B) can be completely prevented by combination liposomes.
  • SDS sodium dodecyl sulfate
  • Tumor cells (MT3 and MT1 breast cancer cells, and Lewis lung lung carcinoma cells) are fluorescently labeled as described in 2.2.1, but the marker used is 282 nmol Hoechst 33258/75 cm 2 culture flask. The cells thus labeled are taken up in PBS (Ca, Mg) and adjusted to 1, 2 ⁇ 10 6 cells / ml.
  • the inhibition of the adhesion of the tumor cells by liposomes is then carried out as follows.
  • the quantification of the bound tumor cells then takes place via the mean fluorescence intensity from the standard series for each tumor cell line.
  • Figure 3 shows the results for the inhibition of the adhesion of MT3, MT1 and Lewis lung and tumor cells.
  • the combined liposomes inhibit the adhesion of MT3 to collagen and to fibronectin by 65 and 52%, respectively.
  • inhibition levels of 34% and 6% were achieved.
  • Example 5.4 Inhibition of complex formation from tumor cells and platelets by combination liposomes in vitro.
  • Tumor cells in the exponential growth phase are washed twice with Ca 2+ / Mg 2+ -PBS and incubated with 94 nmol of the Hoechst dye 33258 in 2 ml of Ca 2-1 VMg 2+ - PBS for 30 min at 37 0 C. After removal of the dye, the cells are washed with Ca 2-1 VMg 2+ PBS and harvested by trypsinization, taken up in 2 ml of medium, centrifuged at 200 g '1 for 2 min and then dissolved in 2 ml Ca 2 -VMg 2+ . PBS resuspended. The cell suspension is then adjusted to a defined cell number.
  • 1x10 4 MT3 tumor cells labeled with Hoechst 33258, 2.5 ⁇ 10 6 platelets and 10 nmol liposomes were taken up in 100 ⁇ l Thyrode buffer (TH buffer), activated with 0.01 ⁇ l thrombin and for 20 min under room temperature with uniform motion incubated.
  • the mixture is then diluted to 2 ml with TH buffer and applied to slides with the cytospin centrifuge at 1500 rpm for 20 min. Thereafter, the samples are covered with a drop of mounting medium (Fluorescent Mounting Medium) and a coverslip bubble-free.
  • THR platelets
  • aPt activated platelets
  • Kon-L blank control
  • DIP-L and DIP / OPP-L liposomes with encapsulated dipyridamole, OPP or both active ingredients
  • Combination liposomes completely prevent the aggregation (complex formation) of the tumor cells, whereas they increase to 172% without the addition of liposomes when the platelets are activated.
  • Example 7 Therapeutic efficacy of the liposomal formulations in the MT3 breast cancer model
  • the antimetastasis effect is tested in the mouse model in different tumor models with different potential for metastasis (eg in the human HT29 colon carcinoma and MT3 and the MT1 breast cancer model).
  • Example 7.1 Influence of the time of liposome application on the inhibition of metastasis development in MT3 mouse breast cancer
  • a group at the time - 6h (based on the tumor cell injection), another group at the times -6, -4 and -2 h, and a group at the times -6, - 2, +2 and +4 h each treated with 100 nmol total lipid liposomes per 20 g mouse iv.
  • the animals are killed, autopsied and the weights of the tumors and lungs determined.
  • Example 7.2 Influence of the dose of liposomes on the inhibition of metastasis development in the mouse
  • mice receive 50, 100 or 200 nmol total lipid combination liposomes per 20 g of mouse.
  • a single treatment with 100 nmol combination liposomes / mouse can significantly reduce metastasis.
  • lung metastases no longer occur. While no other organs are affected, muscle metastases occur in some mice.
  • Example 7.3 Inhibition of metastasis development in MT3 mouse breast cancer
  • mice of the control group receive physiological saline once, while the mice in the other groups receive 100 nmol of total lipid of the specified liposomes or corresponding amounts of active ingredients iv injected.
  • the animals are killed, an autopsy performed and the weights of the tumors and the metastatic number in the lung are determined.
  • mice of the control group receive a single saline solution while the mice in the second group receive combination liposomes injected with 100 nmol of total lipid iv.
  • the animals are killed, an autopsy and determined the metastatic number in the lung and outside the lungs.
  • Example 7.5 Inhibition of metastasis development in HT29 colon carcinoma in the mouse
  • the control group receives saline while the mice in the second group receive liposomes injected with 100 nmol of total lipid iv.
  • the animals are killed, autopsied and the weights of the tumors and the number of metastases determined.
  • Platelets activated platelets without liposomes as controls.
  • Comb-Lip aggregation in the presence of DIP-OPP-liposomes (combination liposomes).
  • Each value represents the mean +/- S.D. for a quadruple determination of at least 8 independently performed experiments.
  • Platelets Platelets without liposomes as controls
  • Kon Lip Adhesion in the presence of control liposomes.
  • Comb Lip Adhesion in the presence of DIP-OPP liposomes.
  • Collagen or fibronectin were immobilized in a 96-well cell culture plate. Subsequently, 6 ⁇ 10 4 tumor cells / well were added and incubated with 100 nmol TL liposomes for 1 hour at 37 ° C.
  • Tumor cells quantified by fluorescence measurement at: ⁇ Ex : 355 nm and XEM: 460 nm. Given the percent inhibition of adhesion in the presence of liposomes compared to the control. Each value represents the mean +/-
  • MT1 Human breast cancer cells
  • the control group received physiological saline once, while the mice in the other groups at the indicated times (based on the tumor cell injection) in each case 100 nmol
  • Fig. 4 Number of visible metastases
  • Fig. 5 Weight of extrapulmonary metastases
  • the control group received a single saline solution while the mice in the other groups received the indicated amounts (total lipid) of liposomes injected iv.
  • the animals were killed, an autopsy made and the weights of the tumors as well as the metastasis! in the
  • Fig. 7 Weight of extrapulmonary metastases
  • Liposomes DIP / OPP-L: combination liposomes with dipyridamole and OPP
  • DIP-L liposomes with dipyridamole
  • OPP-L liposomes with OPP
  • OPP free octadecyl- (1, 1-dimethyl-piperidin-4-yl) -phosphate
  • Scores [indicated is the diameter (in mmy.core): 1: 1; 1-2: 8; 2-3: 27; 3-4: 64; 4-5: 125; 5-8: 512.
  • Figure 10 Inhibition of metastasis by combination liposomes in the MT1 breast cancer model in vivo
  • mice were treated with physiological NaCl solution (control group) or combination liposomes with 100 nmol total lipid iv.
  • the animals were killed, autopsied and metastatic numbers determined in the lung and outside the lungs.
  • mice were injected with 2.5x10 6 HT29 colon carcinoma cells at time t ⁇ O. Six and two hours before, the mice were given physiological saline (control) or liposomes with 100 nmol of total lipid iv. 8 mice were used per group. After 36 days, the animals were killed, an autopsy performed and the weights of the tumors and the metastatic count were determined.
  • DOPC dioleyl phosphatidylcholine

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Abstract

L'invention concerne de nouvelles préparations pharmaceutiques et leur utilisation pour produire des médicaments destinés à lutter contre des métastases, ces médicaments contenant des liposomes. Ces préparations sont des formulations liposomales. Selon l'invention, ces formulations contiennent, outre des substances additives éventuelles, au moins une substance à action antitumorale et au moins une substance à action anticoagulante. Elles se composent en particulier de lipides de base et éventuellement d'autres constituants, tels que des lipides auxiliaires, des stabilisateurs de membranes et des porteurs de charges, et peuvent présenter une surface modifiée améliorant leur efficacité.
PCT/DE2008/000264 2007-02-19 2008-02-14 Préparation pharmaceutique pour lutter contre des métastases Ceased WO2008101469A2 (fr)

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DE102007008484.8 2007-02-19
DE102007008484A DE102007008484A1 (de) 2007-02-19 2007-02-19 Pharmazeutische Zubereitung zur Bekämpfung von Metastasen

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WO2008101469A2 true WO2008101469A2 (fr) 2008-08-28
WO2008101469A3 WO2008101469A3 (fr) 2009-01-29

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JP2018505186A (ja) * 2015-01-28 2018-02-22 リアルイン ライフ サイエンス リミテッド ＰＰＡＲγ発現および核転座を促進する化合物およびそれの治療的使用
EP3250201A4 (fr) * 2015-01-28 2018-08-01 Realinn Life Science Limited Composés pour augmenter l'expression et la translocation nucléaire de ppar et utilisation thérapeutique correspondante
AU2016212527B2 (en) * 2015-01-28 2019-03-07 Realinn Life Science Limited Compounds for enhancing PPARgamma expression and nuclear translocation and therapeutic use thereof
CN115282152A (zh) * 2015-01-28 2022-11-04 瑞采生技有限公司 用于增强PPARγ表现及核转位之化合物及其医疗用途
US12178817B2 (en) 2015-01-28 2024-12-31 Realinn Life Science Limited Compounds for enhancing PPARγ expression and nuclear translocation and therapeutic use thereof

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