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WO2008031113A2 - Procédé amélioré d'inhibition de la néoformation de vaisseaux sanguins - Google Patents

Procédé amélioré d'inhibition de la néoformation de vaisseaux sanguins Download PDF

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WO2008031113A2
WO2008031113A2 PCT/US2007/078080 US2007078080W WO2008031113A2 WO 2008031113 A2 WO2008031113 A2 WO 2008031113A2 US 2007078080 W US2007078080 W US 2007078080W WO 2008031113 A2 WO2008031113 A2 WO 2008031113A2
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squalamine
cells
inhibition
growth
nhe
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WO2008031113A3 (fr
WO2008031113A8 (fr
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Michael Mclane
Andrew V. Albright
Hsiao-Ling Hung
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Genaera Corp
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Genaera Corp
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/13—Amines

Definitions

  • This application is directed to the use of the aminosterol compound squalamine for the inhibition of neovascularization as an improved method for the treatment of macular degeneration, cancer and other diseases associated with inappropriate neovascularization.
  • Each of the body's cells must maintain its acid-base balance or, more specifically, its hydrogen ion or proton concentration. Only slight changes in hydrogen ion concentration cause marked alterations in the rates of chemical reactions in the cells-some being depressed and others accelerated.
  • hydrogen ion concentration when a person has a high concentration of hydrogen ions (acidosis), that person is likely to die in a coma, and when a person has a low concentration of hydrogen ions (alkalosis), he or she may die of tetany or convulsions. In between these extremes is a tremendous range of diseases and conditions that depend on the cells involved and level of hydrogen ion concentration experienced. Thus, the regulation of hydrogen ion concentration is one of the most important aspects of homeostasis.
  • the normal cell pH is 7.4, but a person can only live a few hours with a pH of less than 7.0 or more than 7.7. Thus, the maintenance of pH is critical for survival.
  • NHE sodium/proton exchanger-the "NHE,” which is also called an "antiporter.”
  • Na + / H + sodium/proton exchanger-the "NHE”
  • the body has developed a family of NHEs, and recent work has elucidated a family of NHE "isoforms" that are localized in certain tissues and associated with various functions.
  • the NHE isoforms listed below are most likely to be significant.
  • NHEl is a housekeeping exchanger and is believed to be unregulated in hypertension. It is thought to play a role in intracellular pH conduct. Also, it is believed that control of this exchanger will protect a patient from ischemic injury.
  • NHEl is associated genetically with diabetes and, thus, inhibition might alter evolution of diabetes through effects on beta cells in the pancreas.
  • vascular smooth muscle proliferation responsive to glucose, is associated with increased expression of NHEIa.
  • NHE l ⁇ is present on nucleated erythrocytes. It is inhibited by high concentrations of amiloride. This NHE isoform is regulated by adrenergic agents in a cAMP-dependent fashion.
  • NHE2 is associated with numerous cells of the GI tract and skeletal muscle. Inhibition could alter growth of hyperplastic states or hypertrophic states, such as vascular smooth muscle hypertrophy or cardiac hypertrophy. Cancers of muscle origin such as rhabdomyosarcoma and leiomyoma are reasonable therapeutic targets.
  • NHE3 is associated with the colon.
  • the work described below shows it to be associated with endothelial cells. Inhibition would affect functions such as water exchange in the colon (increase bowel fluid flux, which is the basis of, e.g., constipation), colonic cancer, etc. On endothelial cells, normal growth would be inhibited through inhibition of the exchanger.
  • NHE4 is associated with certain cells of the kidney. It appears to play a role in cellular volume regulation. Specific inhibitors might affect kidney function, and hence provide therapeutic benefit in hypertension.
  • NHE5 is associated with lymphoid tissue and cells of the brain. Inhibition of NHE5 should cause inhibition of proliferative disorders involving these cells. NHE5 is a likely candidate for the proliferation of glial cells in response to HIV and other viral infections.
  • the NHE functions to assist the body
  • the inhibition of NHE function should provide tremendous therapeutic advantages.
  • the NHE normally operates only when intracellular pH drops below a certain level of acidity
  • the NHEs upon growth factor stimulation the cell's NHEs are turned on even though the cell is poised at a "normal" resting pH.
  • the NHEs begin to pump protons from the cell at a pH at which they would normally be inactive.
  • the cell undergoes a progressive loss of protons, increasing its net buffering capacity or, in some cases, actually alkalinizing.
  • the growth stimulus does not result in a cellular effect.
  • inhibitors of the NHE family are likely to exert growth-inhibitory effects.
  • NHE3 isoforms found in the colon are believed to play a role in regulating the fluid content of the colonic lumen. This pump is inhibited in cases of diarrhea.
  • the NHE3 isoform present on the proximal tubules of the kidney is believed to play a similar role with respect to renal salt and acid exchange. Accordingly, inhibitors of the NHE family have been regarded as therapeutic modalities for the treatment of hypertension.
  • NHE inhibitors In view of the expected value of the inhibition of NHE action, scientists have sought out NHE inhibitors.
  • the most widely studied inhibitor of NHE is amiloride, a guanidine- modified pyrazine used clinically as a diuretic.
  • a number of derivatives have been generated, incorporating various alkyl substitutions. These derivatives have been studied with the several isoforms of NHE that are known and described above, except for NHE5, for which there is no known inhibitor.
  • NHE inhibitors described by Counillon et al. exhibit specificity for NHEl . They therefore serve a therapeutic value in the treatment of conditions where inhibition of this isoform is beneficial. However, these inhibitors do not target the other known NHE isoforms- e.g., NHE3 is unaffected.
  • NHE3 is expressed on endothelial cells, and its inhibition results in anti-angiogenic effects.
  • the spectrum of NHE isoforms inhibited by squalamine in accordance with the invention are different from those inhibited by the amiloride or the Counillon et al. compounds, and have different, distinct pharmacological effects.
  • NHE inhibitors that exhibit selective action against a single, specific NHE. Such inhibitors would permit more precise inhibition of a tissue by perturbing the effect of the NHE on its growth.
  • NHE-specific inhibitors would allow for the development of new therapies for a whole host of diseases or conditions, including: treating arrhythmias; treating and preventing cardiac infarction; treating and preventing angina pectoris and ischemic disorders of the heart; treating and preventing ischemic disorders of the peripheral and central nervous system; treating and preventing ischemic disorders of peripheral organs and limbs; treating shock; providing anti- arteriosclerotic agents; treating diabetic complications; treating cancers; treating fibrotic diseases, including fibroses of lung, liver and kidney; and treating prostatic hyperplasia.
  • Other therapeutic targets include: treatment of viral disease, such as HIV, HPV and HSV; prevention of malignancies; prevention of diabetes (i.e., islet cell injury); prevention of vascular complications of diabetes; treatment of disorders of abnormal neovascularization, e.g., macular degeneration (such as age-related, both wet and dry forms), rheumatoid arthritis, psoriasis, cancer, malignant hemangiomas; prevention of vascular retenosis; prevention of hypertension-associated vascular damage; immunosuppression; and treatment of collagen vascular disorders.
  • viral disease such as HIV, HPV and HSV
  • prevention of malignancies i.e., islet cell injury
  • prevention of vascular complications of diabetes e.g., macular degeneration (such as age-related, both wet and dry forms), rheumatoid arthritis, psoriasis, cancer, malignant hemangiomas
  • prevention of vascular retenosis prevention of hypertension-associated vascular damage
  • NHEs of bacteria fungi and protozoa would also be valuable as specific antimicrobials. It is known that all living cells use an NHE of one form or another to maintain intracellular Na + and pH homeostasis. NHEs have been cloned from numerous bacteria and fungi, and bear some sequence homology to the mammalian isoforms. Using a highly specific bacterial or fungal NHE as a target, it should be possible to develop a highly specific inhibitor of such an exchanger, one that is particularly advantageous or that lacks activity against the mammalian isoforms. Such compounds would be useful as antibiotics of a different mechanism.
  • Squalamine the structure of which is illustrated in FIG. 5, is an aminosterol which has been isolated from the liver of the dogfish shark, Squalus acanthias.
  • Methods for synthesizing squalamine have been devised, such as the methods described in U.S. Patents 6,262,283, 6,933,383 and 6,610,866.
  • U.S. Patent 5,721,226 describes the use of squalamine as an antiangiogenic agent and U.S. Patents 6,147,060, 6,596,712 and 6,962,909 describe its use to treat cancer and neovascularization in the eye.
  • squalamine e.g., as a sodium/proton exchanger (isoform 3), or NHE3, inhibiting agent and as an agent for inhibiting the growth of endothelial cells
  • squalamine synthesis techniques are disclosed in U.S. Patent No. 5,792,635.
  • VEGF Vascular endothelial growth factor
  • eNOS endothelial nitric oxide synthetase
  • NO nitric oxide
  • AvastinTM is used as a systemic anti-angiogenesis therapy and causes hypertension, presumably through the reduction in the production or stimulation of eNOS, with the resulting reduction of NO causing an increase in vascular tone.
  • Squalamine an anti-angiogenic agent, does not inhibit VEGF-induced eNOS and the linked NO production, as monitored by cGMP production, and therefore should not induce hypertension.
  • an inhibitor of angiogenesis that does not inhibit the induction of eNOS is provided.
  • An aspect of the invention is a method for treating an individual with a disorder associated with neovascularization, comprising administering to the individual an effective amount of squalamine, or a pharmaceutically acceptable salt thereof, to inhibit the neovascularization, wherein the individual is diagnosed with hypertension.
  • FIGS. IA and IB show the inhibition of rabbit sodium/proton exchanger isoform 3 (NHE3) by squalamine.
  • FIG. IA is a plot of the rate of pH recovery (y-axis) as a function of restored extracellular sodium ion concentration (x-axis) for cells acid-preloaded by exposure to 40 mM NH 4 CI, with the curve marked by "+” being for control (no drug) and the curve marked by " ⁇ ” being for squalamine.
  • FIG. IB shows the actual internal pH (y-axis) as a function of time (x-axis) following addition of 5 .mu.g/ml of squalamine for cells not acid-preloaded.
  • FIG. 2 A shows the lack of inhibition of rabbit sodium/proton exchanger isoform 1 (NHEl) by squalamine.
  • FIG. 2B shows the lack of inhibition of human NHEl by squalamine. In these plots of internal pH vs. time, the curve marked by "0" is for squalamine and that marked by "+” is the control (cells incubated in the absence of squalamine).
  • FIGS. 3 A, 3B and 3C show the suppression of the growth of murine melanoma, respectively through the subcutaneous, intraperitoneal and oral administration of squalamine.
  • FIG. 5 shows the structure of squalamine.
  • FIG. 6 indicates that squalamine treatment does not affect VEGF-induced eNOS activity.
  • squalamine The steroid known as squalamine is the subject of U.S. Pat. No. 5,192,756 to Zasloff et al, the disclosure of which is herein incorporated by reference. This compound is a broad- spectrum antibiotic, killing bacteria, fungi and protozoa.
  • the absolute stereochemistry for squalamine, compound 1256, is shown in FIG. 5. The total chemical synthesis of squalamine was reported in 1994.
  • Squalamine has been recovered from extracts of dogfish shark liver.
  • shark liver was extracted in methanokacetic acid.
  • the aqueous extract was adsorbed to Cl 8 silica and eluted with 70% acetonitrile, and the eluate was adsorbed to SP- sephadex and eluted with 1.5 M NaCl.
  • the eluate was adjusted to 5M NaCl, and the steroids salted out.
  • the precipitate was filtered over Celite and eluted with hot water, followed by methanol.
  • the eluate was reduced in volume and applied to a 1-inch Cl 8 column, and subjected to chromatography utilizing an increasing gradient in acetonitrile.
  • Aminosterol compounds such as squalamine have been discovered to be effective inhibitors of NHE.
  • squalamine has been found to advantageously inhibit a specific NHE isoform-the compound inhibits NHE3, but not NHEl.
  • squalamine has been determined to inhibit the exchanger through a special mechanism. The special and advantageous effects and utilities of squalamine and other aminosterols are further evident from the results of the experimental tests discussed below.
  • squalamine was assayed against a cell line expressing either human NHEl or human NHE3 following procedures outlined in Tse et al., J. Biol. Chem. 268, 1993, 11917-11924. Internal pH was measured either following acid loading or in the absence of an acid-loading ' challenge, with the results shown in FIGS. IA and IB.
  • PS 120 fibroblasts transfected with rabbit NHE3 were grown in supplemented Dulbecco's-modified Eagle's medium as described by Levine et al., J. Biol. Chem. 268, 1993, 25527-25535.
  • Transfected cells grown on glass coverslips were then assayed for internal pH changes following treatment with 5 ⁇ g/ml squalamine using the fluorescent dye BCECF-AM (2 l ,7'-bis(carboxyethyl)-5(6)-carboxyfiuorescein-acetoxymethyl ester) as a pH indicator as described by Levine et al.
  • the aminosterol squalamine not only reduces the absolute number of protons that can be secreted by the cell (the V max effect), but also forces the cell to fall to a lower pH in the presence of this inhibitor (the K n , effect).
  • the sodium/proton exchanger is more profoundly inactivated by squalamine than by amiloride.
  • squalamine exhibited no inhibitory activity against human NHEl or rabbit NHEl as shown in FIGS. 2A and 2B.
  • PS 120 fibroblasts transfected with rabbit or human NHEl were grown as described above.
  • Transfected cells expressing rabbit NHEl (FIG. 2A) or human NHEl (FIG. 2B) grown on glass coverslips were then assayed for internal pH changes following treatment with 5 .mu.g/ml squalamine using the fluorescent dye BCECF-AM with cells acid-preloaded by exposure to 40 mM NH4CI. The rate of pH recovery as a function of restored extracellular sodium ion concentration was monitored.
  • squalamine has been discovered to be a distinct inhibitor with specificity for NHE3 over NHE 1. Moreover, squalamine has been identified as an inhibitor that causes a cell to drop to a lower pH before the pump is activated. The results shown in FIGS. IA, IB, 2A and 2B demonstrate that squalamine exhibits a unique NHE specificity.
  • Endothelial Cells When non-transformed human cells are grown in the presence of increasing concentrations of squalamine, endothelial cells exhibit a particular sensitivity to squalamine, as shown by the following experiment. Bovine pulmonary endothelial cells, human epithelial cell line MCF 1 OA, and human foreskin fibroblasts were incubated in the presence of 12 different memb ⁇ ane-active agents, including peptides and squalamine.
  • LPS-Induced Neutrophil Adherence to Human Umbilical Venous Endothelial Cells When endothelial cells are exposed to certain stimuli, including lipopolysaccharide (LPS) and certain cytokines, specific adhesion molecules are induced on the plasma membrane that enhance the binding of leukocytes. These leukocyte-endothelial cell interactions are believed to be necessary to localize leukocytes to sites of bacterial invasion and to facilitate extravasation of the leukocytes from the capillary into the surrounding tissue space.
  • Leukocyte-adhesion molecules include the Selectins and ICAM-I .
  • the cells were grown in serum-free media overnight. For induction, either TNF- ⁇ (40 ng/ml) was added to endothelial cells for 6 hours prior to adding neutrophils or LPS (100 ng/ml) was added for 4-6 hours. It was found that the LPS response was increased by adding 1% FBS to the wells to provide a source of LPS-binding protein. After activation of the endothelial cells, approximately 5OxIO 6 neutrophils were added per well. The plates were gently rocked for 30 minutes at room temperature, followed by removal of the media and washing in serum-free media three times and then photographing of each well.
  • TNF- ⁇ 40 ng/ml
  • LPS 100 ng/ml
  • squalamine is an inhibitor of capillary growth.
  • the growing capillaries within the chorioallantoic membrane model have been used as a system in which to evaluate the effect of agents on their potential to inhibit new vessel growth.
  • Neovascularization occurs most aggressively over the first week of embryonic development. Thereafter capillary growth is characterized by principally "elongation” rather than "de novo" formation.
  • agents are applied locally to a region of the embryo over which neovascularization will occur. Agents are assessed by their ability to inhibit this process, as evaluated by visual examination about 7 days after application. Agents which disrupt vascular growth during the period of de novo capillary formation, but do not interfere with subsequent capillary growth, are generally regarded as "specific" inhibitors of neovascularization, as distinguished from less specific toxic substances.
  • the assay utilized is described in detail in Auerbach et al., Pharm. Ther. 51, 1991, 1-11. Results are tabulated below.
  • squalamine exhibited potent but specific inhibitory activity, equal in potency to the most active compounds described to date in the literature.
  • the effect is compatible with suppression of neovascularization rather than toxic inhibition of capillary growth.
  • Squalamine was found to disrupt vitelline capillaries in 3- to 5-day chick embryos.
  • the 3-day chick embryo consists of an embryonic disc from which numerous vessels emerge and return, forming a "figure 8"-shaped structure-the embryo in the center with vascular loops extending outward over both poles.
  • Application of squalamine onto the embryonic structure (0.1 ml in 15% Ficol in PBS) resulted in progressive "beading up" of the vitelline vessels, with the finest capillaries being the first to exhibit these changes. Following a lag period of around 15 minutes, the constriction of continuity between capillary and secondary vessels, generally on the "venous" side, was observed.
  • a newly developed assay employing tadpoles preferably Xenopus laevis Stages 59-60, were employed to study the effect of a compound by monitoring capillary occlusion in the tadpole's tail. Animals at these stages were used because they represent the period of transition through metamorphosis at which time the animals possess both embryonic and adult stage tissues.
  • the compounds of the invention affect the shape, viability and integrity of the embryonic tissues while not affecting the adult tissues, providing a powerful, highly specific screen. For example, substances that destroy all of the animal's epithelium, both adult and embryonic, could be regarded as toxic. Substances that destroy only the embryonic tissues exhibit a very unique specificity.
  • tadpoles are introduced into Petri dishes containing a solution of the test compound in distilled water, preferably about 100 ml.
  • the preferred concentration of the test compound is from about 1 ⁇ g/ml to about 10 ⁇ g/ml.
  • the volume of liquid is sufficient for the animal to swim freely and drink from the solution. Thus, the effect observed results from oral absorption and subsequent systemic distribution of the agent. If the volume of liquid is not sufficient to permit oral intake, the effects that are observed would result from absorption through the surface epithelium.
  • this simple assay can identify if an compound has characteristics of oral availability.
  • a solution of a compound in water can be injected directly into the abdomen of the animal using standard techniques. Concentrations of the compound from about 0.05 mg/ml to about 0.5 mg/ml in about 0.05 ml of water are preferred.
  • B16 melanoma cells Using the growth of B16 melanoma cells in C57B mice, a recognized model for the evaluation of inhibitors of angiogenesis on the growth of cancers, the effects of subcutaneous, intraperitoneal and oral administration of squalamine were evaluated.
  • An inoculum of B16 melanoma cells was implanted subcutaneously on the dorsum of the C57B mouse, which resulted in the progressive growth of melanoma lesions over 30-40 days as shown in FIGS. 3A-3C.
  • melanoma 1205Lu develops aggressively in RAG-I mice after implantation. Squalamine has been found to suppress the growth of melanoma 1205Lu in RAG-I mice in a dose-dependent fashion. Squalamine was administered after tumors had reached about 0.1 ml, and clear suppression of tumor growth in a dose-dependent fashion was found as evidenced by FIG. 4. After cessation of treatment, tumor growth continued at a rate similar to untreated controls, suggesting that the impact of squalamine in this setting is reversible.
  • squalamine provides a potent inhibitor of NHE3. Squalamine therefore should provide invaluable therapeutic intervention wherever new blood vessel formation in vivo is implicated.
  • squalamine has therapeutic utility in the treatment of diseases or disorders dependent on continued neovascularization where interruption of neovascularization diminishes the intensity of the pathological process.
  • squalamine has utility for treating disorders including solid tumor growth and metastasis, rheumatoid arthritis, psoriasis, diabetic retinopathy, macular degeneration, neovascular glaucoma, papilloma, retrolental fibroplasia, and organ rejection.
  • HAVECs Human Umbilical Vein Endothelial Cells
  • EMM-2 Endothelial Growth Media-2
  • Squalamine was added at a final concentration of 3 ⁇ M for 23 hrs as indicated, after which cells were washed in unsupplemented media to remove serum and growth factors.
  • the cells were incubated in HBSS at 37 0 C for 30 minutes, and then recombinant VEGF-A (165) was added at a final concentration of 12.5 ng/mL as indicated.
  • IBMX was added to all dishes at a final concentration of 0.5 mM to inhibit the non-specific phosphodiesterases from lowering guanosine 3',5' cyclic monophosphate (cGMP) levels after cell lysis. After 15 minutes of VEGF and IBMX incubation, the supernatant was discarded and cells scraped off the plates in 500 ⁇ L of 0.1 N HCl. Cell lysates were assayed for cGMP levels using a cGMP competitive enzyme-linked immunoassay (Cayman Chemical). Protein levels were measured with the colormetric microBCA Protein Assay (BioRad) to insure normalization and plot pmol cGMP per mg of protein in the cell lysates.
  • cGMP guanosine 3',5' cyclic monophosphate
  • cGMP is a surrogate marker for the activity of eNOS and therefore also an indicator of an increase in NO production.
  • the results of this assay, shown in FIG. 6, indicate that squalamine does not affect the VEGF-induced induction of eNOS. Therefore, unlike anti-VEGF agents, squalamine should not cause a hypertensive side effect when used for the inhibition of neovascularization.
  • the patient to be treated can be any animal, and is preferably a mammal. More preferably, the patient is a human.
  • compositions for use in vitro or in vivo in accordance with the present invention thus may be formulated in a conventional manner using one or more physiologically acceptable carriers comprising excipients and auxiliaries that facilitate processing of the active compounds into preparations which can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen.
  • carriers or excipients include but are not limited to calcium carbonate, calcium phosphate, various sugars, starches, cellulose derivatives, phospholipids, liposomal carriers, gelatin and polymers such as polyethylene glycols.
  • a pharmaceutical carrier for the squalamine is a cosolvent system comprising benzyl alcohol, a nonpolar surfactant, a water-miscible organic polymer, and an aqueous phase.
  • the proportions of a co-solvent system may be varied considerably without destroying its solubility and toxicity characteristics.
  • identity of the co-solvent components may be varied: for example, other low-toxicity nonpolar surfactants may be used instead of polysorbate 80; the fraction size of polyethylene glycol may be varied; other biocompatible polymers may replace polyethylene glycol, e.g. polyvinyl pyrrolidone; and sugars or polysaccharides, such as dextrose.
  • compositions of the invention may also include stabilizers and preservatives.
  • stabilizers and preservatives for an exemplary listing of typical carriers, stabilizers and adjuvants known to those of skill in the art, see Remington: The Science and Practice of Pharmacy, 21 st ed. (2005).
  • Pharmaceutically acceptable salts of the squalamine include the conventional non-toxic salts or the quaternary ammonium salts which are formed from inorganic or organic acids or bases.
  • acid addition salts include acetate, adipate, benzoate, benzenesulfonate, citrate, camphorate, dodecylsulfate, hydrochloride, hydrobromide, lactate, maleate, methanesulfonate, nitrate, oxalate, pivalate, propionate, succinate, sulfate and tartrate.
  • Base salts include ammonium salts, alkali metal salts such as sodium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases such as dicyclohexylamine salts and salts with amino acids such as arginine. Also, the basic nitrogen-containing groups may be quatemized with, for example, alkyl halides.
  • compositions of the invention may also include stabilizers and preservatives.
  • stabilizers and preservatives for examples of typical carriers, stabilizers and adjuvants known to those of skill in the art, see Remington: The Science and Practice of Pharmacy, 21 st ed. (2005).
  • the squalamine may be administered alone or preferably as a pharmaceutical formulation comprising the squalamine together with at least one pharmaceutically acceptable carrier.
  • other therapies known to those of skill in the art may be combined with the administration of the squalamine.
  • a “therapeutically effective amount” is an amount of squalamine which inhibits, totally or partially, the progression of the condition or alleviates, at least partially, one or more symptoms of the condition.
  • a therapeutically effective amount can also be an amount that is prophylactically effective. The amount that is therapeutically effective will depend upon the patient's size and gender, the condition to be treated, the severity of the condition and the result sought. For a given patient, a therapeutically effective amount can be determined by methods known to those of skill in the art.
  • In vivo administration of the squalamine can be effected in one dose, multiple doses, continuously or intermittently throughout the course of treatment. Doses range from about 0.05 mg/kg to about 5 mg/kg, preferably between about 0.5 mg/kg to about 1 mg/kg, in single or divided daily doses. Methods of determining the most effective means and dosage of administration are well known to those of skill in the art and will vary with the composition used for therapy, the purpose of the therapy, the target cell being treated and the subject being treated. Single or multiple administrations can be carried out with the dose level and pattern being selected by the treating physician.
  • compositions containing the squalamine can be administered by any suitable route, including oral, rectal, intranasal, topical (including transdermal, aerosol, buccal an sublingual), parenteral (including subcutaneous, intramuscular, intravenous), intraperitoneal and pulmonary.
  • suitable route including oral, rectal, intranasal, topical (including transdermal, aerosol, buccal an sublingual), parenteral (including subcutaneous, intramuscular, intravenous), intraperitoneal and pulmonary.
  • the preferred route will vary with the condition and age of the recipient, and the disease being treated.
  • the preferred routes of administration are oral, topical, subcutaneous, intramuscular and/or intravenous.
  • the squalamine can be formulated readily by combining it with pharmaceutically acceptable carriers well known in the art.
  • Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a patient to be treated.
  • compositions for oral use can be obtained by combining the active compound with a solid excipient, optionally grinding a resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores.
  • suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl- cellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone (PVP).
  • disintegrating agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.
  • compositions for topical administration of the squalamine maybe formulated in conventional ophthalmologically compatible vehicles, such as, for example, an ointment, cream, suspension, lotion, powder, solution, paste, gel, spray, aerosol or oil.
  • ophthalmologically compatible vehicles such as, for example, an ointment, cream, suspension, lotion, powder, solution, paste, gel, spray, aerosol or oil.
  • These vehicles may contain compatible preservatives such as benzalkonium chloride, surfactants such as polysorbate 80, liposomes or polymers such as methylcellulose, polyvinyl alcohol, polyvinyl pyrrolidone and hyaluronic acid, which may be used for increasing viscosity.
  • preferred formulations are ointments, gels, creams or eye drops containing squalamine.
  • the squalamine for use according to the present invention is conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebuliser, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
  • a suitable propellant e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
  • the dosage unit may be determined by providing a valve to deliver a metered amount.
  • Capsules and cartridges of e.g. gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch.
  • the squalamine can be formulated for parenteral administration by injection, e.g. bolus injection or continuous infusion.
  • Formulations for injection may be presented in unit dosage form, e.g. in ampoules or in multi-dose containers, with an added preservative.
  • the compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as buffers, bacteriostats, suspending agents, stabilizing agents, thickening agents, dispersing agents or mixtures thereof.
  • compositions for parenteral administration include aqueous solutions of the active compounds in water-soluble form. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents that increase the solubility of the compounds to allow for the preparation of highly concentrated solutions. In a preferred embodiment, the squalamine is dissolved in a 5% sugar solution, such as dextrose, before being administered parenterally.
  • a 5% sugar solution such as dextrose
  • the agents of the invention may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks's solution, Ringer's solution, or physiological saline buffer.
  • physiologically compatible buffers such as Hanks's solution, Ringer's solution, or physiological saline buffer.
  • penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
  • the squalamine may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter or other glycerides.
  • VEGF Vascular endothelial growth factor
  • VPF vascular permeability factor
  • Papapetropoulos A. et ai "Nitric oxide production contributes to the angiogenic properties of vascular endothelial growth factor in human endothelial cells," J. Clin Invest. 1997; 100:3131-3139. 5. Gelinas DS et ai, "Immediate and delayed VEGF-mediated NO synthesis in endothelial cells: role of PDK, PKC and PLC pathways.” Br. J, Pharmacol. 2002:137: 1021-1030.

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Abstract

La présente invention concerne l'utilisation d'un composé d'aminostérol, la squalamine, capable d'inhiber la néoformation de vaisseaux sanguins dans un procédé amélioré de traitement de la dégénérescence maculaire, du cancer et d'autres maladies associées à une néoformation inappropriée de vaisseaux sanguins chez un individu présentant de l'hypertension.
PCT/US2007/078080 2006-09-08 2007-09-10 Procédé amélioré d'inhibition de la néoformation de vaisseaux sanguins Ceased WO2008031113A2 (fr)

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US84294106P 2006-09-08 2006-09-08
US60/842,941 2006-09-08

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WO2008031113A2 true WO2008031113A2 (fr) 2008-03-13
WO2008031113A8 WO2008031113A8 (fr) 2008-05-22
WO2008031113A3 WO2008031113A3 (fr) 2008-07-10

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PCT/US2007/078080 Ceased WO2008031113A2 (fr) 2006-09-08 2007-09-10 Procédé amélioré d'inhibition de la néoformation de vaisseaux sanguins

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2013537551A (ja) * 2010-08-17 2013-10-03 オーエイチアール・ファーマシューティカル・インコーポレイテッド スクアラミンの眼用製剤
EP2506856B1 (fr) * 2009-12-02 2019-01-02 Assistance Publique Hôpitaux de Marseille Composés aminostéroïdiens pour une application topique locale pour la décolonisation cutanéo-muqueuse de staphylococcus aureus

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6544761B2 (en) * 1994-12-13 2003-04-08 Human Genome Sciences, Inc. Human tissue inhibitor of metalloproteinase-4

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2506856B1 (fr) * 2009-12-02 2019-01-02 Assistance Publique Hôpitaux de Marseille Composés aminostéroïdiens pour une application topique locale pour la décolonisation cutanéo-muqueuse de staphylococcus aureus
JP2013537551A (ja) * 2010-08-17 2013-10-03 オーエイチアール・ファーマシューティカル・インコーポレイテッド スクアラミンの眼用製剤
JP2016166250A (ja) * 2010-08-17 2016-09-15 オーエイチアール・ファーマシューティカル・インコーポレイテッドＯｈｒ Ｐｈａｒｍａｃｅｕｔｉｃａｌ，Ｉｎｃ． スクアラミンの眼用製剤

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WO2008031113A3 (fr) 2008-07-10
WO2008031113A8 (fr) 2008-05-22

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