WO2014028059A1 - Inhibiteurs de rac1 pour le traitement de la maladie glomérulaire d'alport - Google Patents

Inhibiteurs de rac1 pour le traitement de la maladie glomérulaire d'alport Download PDF

Info

Publication number
WO2014028059A1
WO2014028059A1 PCT/US2013/032432 US2013032432W WO2014028059A1 WO 2014028059 A1 WO2014028059 A1 WO 2014028059A1 US 2013032432 W US2013032432 W US 2013032432W WO 2014028059 A1 WO2014028059 A1 WO 2014028059A1
Authority
WO
WIPO (PCT)
Prior art keywords
inhibitor
alport
mesangial
cdc42
racl
Prior art date
Application number
PCT/US2013/032432
Other languages
English (en)
Inventor
Dominic Cosgrove
Original Assignee
Father Flanagan's Boys Home Doing Business As Boys Town National Research Hospital
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Father Flanagan's Boys Home Doing Business As Boys Town National Research Hospital filed Critical Father Flanagan's Boys Home Doing Business As Boys Town National Research Hospital
Priority to EP13829878.1A priority Critical patent/EP2884988A4/fr
Publication of WO2014028059A1 publication Critical patent/WO2014028059A1/fr
Priority to US14/580,680 priority patent/US9719981B2/en
Priority to US15/631,454 priority patent/US10545134B2/en

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys

Definitions

  • Alport syndrome (also referred to as hereditary nephritis) is a genetic disorder characterized by abnormalities in the basement membranes of the glomerulus (leading to hematuria, glomerulosclerosis, and end-stage kidney disease (ESRD)), cochlea (causing deafness), and eye (resulting in lenticonus and perimacular flecks).
  • Alport syndrome is a primary basement membrane disorder caused by mutations in the collagen type IV COL4A3, COL4A4, or COL4A5 genes. Mutations in any of these genes prevent the proper production or assembly of the type IV collagen network, which is an important structural component of basement membranes in the kidney, inner ear, and eye.
  • Basement membranes are thin, sheet-like structures that separate and support cells in many tissues.
  • the abnormalities of type IV collagen in kidney basement membranes leads to irregular thickening and thinning and splitting of basement membranes, causing gradual scarring of the kidneys.
  • Alport Syndrome causes progressive kidney damage.
  • the glomeruli and other normal kidney structures such as tubules are gradually replaced by scar tissue, leading to kidney failure.
  • Deafness and an abnormality in the shape of the lens called anterior lenticonus are other important features of Alport Syndrome. People with anterior lenticonus may have problems with their vision and may develop cataracts.
  • the prevalence of Alport syndrome is estimated at approximately 1 in 5,000 births and it is estimated that the syndrome accounts for approximately 2.1 percent of pediatric patients with ESRD.
  • treatments are
  • kidney failure patients are advised on how to manage the complications of kidney failure and the proteinuria that develops is often treated with ACE inhibitors. Once kidney failure has developed, patients are given dialysis or can benefit from a kidney transplant, although this can cause problems. The body may reject the new kidney as it contains normal type IV collagen, which may be recognized as foreign by the immune system. Thus there is a need for improved therapeutic approaches for the treatment of Alport syndrome.
  • the present invention includes a method of treating Alport syndrome in a subject, the method including administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes a method of preventing glomerular disease progression in a subject diagnosed with Alport syndrome, the method including administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes a method of treating glomerulonephritis in a subject, the method including administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes a method of treating kidney injury due to biomechanical strain in Alport syndrome, the method including administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes a method of inhibiting deposition of laminin 211 in the glomerular basement membrane (GBM) in a subject, the method including administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes a method of inhibiting mesangial cell process invasion of the glomerular capillary loop in a kidney of a subject, the method including administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes a method of inhibiting Alport glomerular pathogenesis in a subject; the method including: determining that the subject is at risk for developing Alport glomerular disease; and administering an effective amount of a RACl inhibitor and/or a CDC42 inhibitor to the subject.
  • the determination that the subject is at risk for developing Alport glomerular disease is determined by family medical history, genetic testing, immunodiagnostic skin biopsy testing, and/or molecular diagnostic marker testing.
  • the determination that the subject is at risk for developing Alport glomerular disease is made prior to the onset of proteinuria in the subject.
  • the administration of an effective amount of a RACl inhibitor and/or a CDC42 inhibitor is initiated prior to the onset of proteinuria in the subject.
  • the RACl inhibitor and/or a CDC42 inhibitor blocks CDC42 activation of the endothelin type I receptor and/or the endothelin type II receptor.
  • the RACl inhibitor and/or a CDC42 inhibitor is an endothelin (ET) receptor antagonist.
  • the endothelin (ET) receptor antagonist is a dual antagonist of both the ETA receptor and ET B receptor.
  • the RACl inhibitor and/or a CDC42 inhibitor is bosentan or a derivative thereof
  • the RAC 1 inhibitor is N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N
  • NSC23766 or a derivative thereof.
  • FIGS. 1A to IF Laminin 211 localizes to the glomerular basement membrane (GBM) in Alport glomeruli. Dual immunofluorescence immunostaining was performed on wild type (Figs. 1A-1C) and Alport (Figs. ID- IF) glomeruli from 7 week 129 Sv mice. Glomerular basement membranes were labeled with labeled with anti-laminin a5 antibodies (Fig. 1 A and Fig. ID). Anti-laminin a2 immunostaining is shown in Fig. IB and Fig. IE. Note the irregular deposits of laminin 211 in the Alport GBM, especially in the thickened regions of the GBM (overlapping staining in Fig. ID and Fig. IE). Anti-laminin a2 immunostaining is not observed in the GBM of wild type mice (note the absence of overlapping immunostaining in Fig. 1 A and Fig. IB).
  • FIGS. 2A to 2L Mesangial processes invade the capillary loops of Alport glomeruli where they co-localize with laminin 211 deposits. Dual immunofluorescence immunostaining was performed on wild type or Alport kidney sections from 7 week old 129 Sv mice.
  • Figs. 2A- 2F show localization of laminin a2 and integrin a8 (a mesangial cell marker), and
  • Figs. 2G-2L show localization of laminin a5 (a GBM marker) and integrin a8. Note circumferential co- localization of laminin a2 and integrin a8 in the Alport glomerulus in Figs. 2D-2F, and the co- localization of integrin a8 and laminin a5 in Figs. 2J-2L indicating invasion of the glomerular capillary tufts with mesangial processes.
  • FIGs 3 A to 3C Mesangial processes invade the capillary loops of human Alport glomeruli where they co-localize with laminin 511. Cryosections from human Alport kidneys were stained with antibodies specific for laminin a5 (Fig. 3A) and integrin a8 (Fig. 3B). The integrin a8-specific mesangial processes localize adjacent to the laminin 15-positive GBM, consistent with mesangial process invasion.
  • FIGS 4 A to 4F Hypertension exacerbates mesangial invasion of the glomerular capillary tufts in Alport mice.
  • the X- linked Alport mouse model (on the C57 Bl/6 background) was made hypertensive by providing L-NAME salts in the drinking water from 5 weeks to 10 weeks of age. Control Alport mice were given normal drinking water. Glomeruli were analyzed by dual immunofluorescence immunostaining using antibodies against either laminin a2 or integrin a8. Extensive mesangial process invasion of the capillary tuft is observed in the glomeruli from the salt-treated mice relative to the mice given normal drinking water.
  • FIG. 1 Extensive mesangial process invasion of the glomerular capillary tufts is observed in CD 151 knockout mice.
  • Kidney cryosections from 8 week old wild type and CDC 151 KO mice (on the FVB background) were analyzed by dual immunofluorescence immunostaining using antibodies against either laminin a2 or integrin a8.
  • Extensive mesangial process invasion of the capillary tuft is observed in the glomeruli from CD151 knockout mice relative to wild type mice. Note that the extent of mesangial process invasion in CD 151 knockout mice is much greater than that observed in Alport mice.
  • FIG. 6 Biomechanical stretching of cultured primary mesangial cells induces expression of pro-migratory cytokines, CTGF and TGF- ⁇ mRNA.
  • FIG. 7 l integrin deletion in Alport mice results in markedly reduced mesangial process invasion of the glomerular capillary tufts.
  • the degree of mesangial process invasion of the glomerular capillary tufts was greatly reduced in the integrin ⁇ -null Alport mice relative to age/strain-matched Alport mice.
  • FIG. 8A shows migration of primary cultured mesangial cells is significantly reduced under conditions of integrin l deletion, Integrin linked kinase inhibition, Racl inhibition, and CDC42 inhibition, but not AKT inhibition. In contrast, the migratory potential of cultured integrin al- null mesangial cells is unaffected by inhibition of either Racl or CDC42.
  • FIG. 8B shows treatment of cultured mesangial cells with LPS induced cytoskeletal rearrangement with numerous actin spikes (untreated cells, A; LPS treated cells, B), and these morphological changes are blocked by treatment of cells with either Racl inhibitors (C), or CDC42 inhibitors (D).
  • Fig. 8C shows treatment of cultured mesangial cells with LPS results in polarized localization of CDC42 and associated with filopodia (Figure 8C(b), insert, compared to Golgi and cytosolic localization of CCD42 in wild type cells (8C(a)).
  • FIG. 8D shows a GTP-Racl pull-pull down assay which confirms LPS-mediated activation of Racl in cultured mesangial cells, that was blocked by pre-treatment with Racl inhibitors, but not CDC42 inhibitor.
  • Figures 9A to 9F Treatment of Alport mice with Racl inhibitors partially ameliorates mesangial cell process invasion of the glomerular capillary tufts.
  • Alport mice on the 129 Sv background were injected once daily with either saline or the Racl inhibitor NSC 23766 from 2 weeks to 6 weeks of age.
  • Kidney cryosections were analyzed by dual immunofluorescence immunostaining using antibodies against either laminin a2 or integrin a8. The degree of mesangial process invasion of the glomerular capillary tufts was ameliorated in the Racl inhibitor-treated mice relative to mice injected with saline.
  • FIG. 10A Figures 10A to IOC.
  • Laminin 211 potentiates mesangial process invasion of the glomerular capillary loops in Alport mice, and promotes mesangial cell migration in vitro.
  • laminin a2-deficient Alport mice show reduced mesangial process invasion of the glomerular capillary tufts.
  • Cryosections of kidney tissue from 8 week old laminin a2-deficient Alport mice were analyzed by dual immunofluorescence immunostaining using antibodies against either laminin a5 or integrin a8.
  • Fig. 10B shows wild type mesangial cells migrate more robustly on laminin 211 compared to laminin 521 (GBM laminin).
  • Wound scratch assays were performed using wild type mesangial cells cultured on wither recombinant purified laminins or commercially available laminins extracted from either placenta (primarily laminin 511) or muscle (primarily laminin 211). Images shown are representative of multiple replicates. In Fig.
  • IOC primary mesangial cells from laminin a2-deficient mice show impaired migratory potential relative to wild type mesangial cells. Boyden chamber assays were performed. Blinded cell counts from multiple replicates were analyzed. Asterisk denotes statistically significant differences (p ⁇ 0.05).
  • Alport syndrome (also referred to as hereditary nephritis) is a genetic disorder characterized by abnormalities in the basement membranes of the glomerulus (leading to hematuria, glomerulosclerosis, and end-stage kidney disease (ESRD)), cochlea (causing deafness), and eye (resulting in lenticonus and perimacular flecks).
  • Alport syndrome is a primary basement membrane disorder caused by mutations in the collagen type IV COL4A3, COL4A4, or COL4A5 genes. Mutations in any of these genes prevent the proper production or assembly of the type IV collagen network, which is an important structural component of basement membranes in the kidney, inner ear, and eye.
  • Basement membranes are thin, sheet-like structures that separate and support cells in many tissues.
  • the abnormalities of type IV collagen in kidney basement membranes leads to irregular thickening and thinning and splitting of basement membranes, causing gradual scarring of the kidneys.
  • Alport Syndrome causes progressive kidney damage.
  • the glomeruli and other normal kidney structures such as tubules are gradually replaced by scar tissue, leading to kidney failure.
  • Deafness and an abnormality in the shape of the lens called anterior lenticonus are other important features of Alport Syndrome. People with anterior lenticonus may have problems with their vision and may develop cataracts.
  • the prevalence of Alport syndrome is estimated at approximately 1 in 5,000 births and it is estimated that the syndrome accounts for approximately 2.1 percent of pediatric patients with ESRD.
  • treatments are
  • kidney failure patients are advised on how to manage the complications of kidney failure and the proteinuria that develops is often treated with ACE inhibitors. Once kidney failure has developed, patients are given dialysis or can benefit from a kidney transplant, although this can cause problems. The body may reject the new kidney as it contains normal type IV collagen, which may be recognized as foreign by the immune system. Thus there is a need for improved therapeutic agents for the treatment of individuals with Alport syndrome, especially for the treatment of presymptomatic individuals, before the onset of proteinuria.
  • Alport syndrome is characterized by delayed onset progressive glomerulonephritis associated with sensorineural hearing loss and retinal flecks (Kashtan and Michael, 1996, Kidney Int; 50(5): 1445-1463).
  • the most common form (80%) is X-linked and caused by mutations in the type IV collagen COL4A5 gene (Barker et al, 1990, Science; 8; 248(4960): 1224-7).
  • the two autosomal forms of the disease account for the remaining 20% of Alport patients, and result from mutations in the COL4A3 and COL4A4 genes (Mochizuki et al, 1994, Nat Genet; 8(1):77- 81).
  • the a3(IV), a4(IV) and a5(IV) proteins form a heterotrimer and is assembled into a subepithelial network in the glomerular basement membrane that is physically and biochemically distinct from a subendothelial type IV collagen network comprised of l(IV) and a2(IV) heterotrimers (Kleppel et al., 1992, J Biol Chem; 267(6):4137-4142). Mutations in any one of the three type IV collagen genes that cause Alport syndrome results in the absence of all three proteins in the GBM due to an obligatory association to form functional heterotrimers (Kalluri and Cosgrove, 2000, J Biol Chem; 275(17): 12719-12724).
  • the net result for all genetic forms of Alport syndrome is the absence of the a3(IV) a4(IV) a5(IV) subepithelial collagen network, resulting in a GBM type IV collagen network comprised only of l(IV) and a2(IV) heterotrimers.
  • Alport syndrome results from mutations in type IV collagen COL4A3, COL4A4, or COL4A5 genes. These mutations may be either autosomal recessive (mutations in either COL4A3 or COL4A4 genes (Mochizuki et al, 1994, Nat Genet; 8(1):77-81)) or X-linked (mutations in COL4A5 (Barker et al, 1990, Science; 8;248(4960): 1224-7)).
  • Alport syndrome is also known as congenital hereditary hematuria, hematuria- nephropathy-deafness syndrome, hematuric hereditary nephritis, hemorrhagic familial nephritis, hemorrhagic hereditary nephritis, hereditary familial congenital hemorrhagic nephritis, hereditary hematuria syndrome, hereditary interstitial pyelonephritis, and hereditary nephritis.
  • the present invention includes methods of treating Alport syndrome in a subject by the administration of a RACl inhibitor and/or a CDC42 inhibitor.
  • the administration of a RACl inhibitor and/or a CDC42 inhibitor may result in one or more of the following: inhibiting migration of mesangial cells, inhibiting irregular deposition of mesangial laminin 211 in the GBM, inhibiting accumulation of mesangial integrin ⁇ 8 ⁇ 1 in the capillary loops, inhibiting invasion of the capillary loops by mesangial cell processes, inhibiting mesangial filopodial invasion of the glomerular capillary tuft, and/or preventing, or slowing the onset of proteinuria.
  • the present invention includes methods of preventing, slowing, and/or managing glomerular disease progression in a subject diagnosed with Alport syndrome by the
  • a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes methods of treating glomerulonephritis associated with Alport syndrome in a subject by administering a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes methods of treating kidney injury due to biomechanical strain in Alport syndrome by administering a RACl inhibitor and/or a CDC42 inhibitor.
  • the present invention includes methods of inhibiting deposition of laminin 211 in the glomerular basement membrane (GBM) by administering a RACl inhibitor and/or a CDC42 inhibitor.
  • the laminins are major proteins in the basal lamina, a layer of the basement membrane, a protein network foundation for most cells and organs.
  • Laminins are trimeric proteins that contain an a-chain, a ⁇ -chain, and a ⁇ -chain, found in five, four, and three genetic variants, respectively.
  • the laminin molecules are named according to their chain composition. Thus, laminin-511 contains a5, ⁇ , and ⁇ chains (Aumailley et al., 2005, Matrix Biol;
  • Laminin-211 (composed of ⁇ 2, ⁇ and ⁇ chains (Ehrig et al, 1991, PNAS; 87:3264-3268) is the main laminin isoform in skeletal muscle (Leivo and Engvall, 1988, PNAS; 85: 1544-1588; and Patton, 1997, J Cell Biol; 139: 1507-1521) and identification of laminin a2 chain mutations in a severe form of congenital muscular dystrophy (merosin-deficient congenital muscular dystrophy; MDC1A) established the importance of laminin-211 for normal muscle function (Helbling-Leclerc et al., 1995, Nat Genet; 11 :216-218).
  • MDC1A merosin-deficient congenital muscular dystrophy
  • the present invention includes methods of inhibiting mesangial cell process invasion of the glomerular capillary loop of the kidney by administering a RACl inhibitor and/or a CDC42 inhibitor.
  • RACl also referred to herein as Racl
  • Rho Rho family of GTPases.
  • Members of this superfamily appear to regulate a diverse array of cellular events, including the control of cell growth, cytoskeletal reorganization, and the activation of protein kinases.
  • Rho regulatory of stress fibers
  • Cdc42 regulatory of filopodia
  • the methods of the present invention may be used for the presymptomatic treatment of individuals, with the administration of a RACl inhibitor and/or a CDC42 inhibitor beginning after the determination or diagnosis of Alport syndrome, by prior to the onset of symptoms, such as for, example, proteinuria.
  • the diagnosis of Alport syndrome in an individual may be made, for example, by family medical history, genetic testing, immunodiagnostic skin biopsy testing, and/or molecular diagnostic marker testing. Such methods may be combined with a step of obtaining a diagnosis of Alport syndrome by the use of one or more such diagnostic means.
  • a RACl inhibitor or a CDC42 inhibitor can block the activation of RAC1/CDC42 members of the rho family of small GTPases.
  • Any of a wide variety of RACl inhibitors may be used with the methods described herein, including, but not limited to, NSC23766 and derivatives thereof (Gao et al, 2004, PNAS; 101 :7618-7623), EHT 1864 and derivatives thereof (Shutes et al, 2007, J Biol Chem; 282:35666-35678), W56 (Gao et al, 2001, J Biol Chem; 276:47530), F56 (Gao et al, 2001, J Biol Chem; 276:47530), and any of the RACl inhibitors described by Ferri et al.
  • a RACl inhibitor may be NSC23766 or a derivative thereof.
  • Human CDC42 is a small GTPase of the Rho-subfamily, which regulates signaling pathways that control diverse cellular functions including cell morphology, migration, endocytosis and cell cycle progression.
  • Any of a wide variety of CDC42 inhibitors may be used with the methods described herein, including, but not limited to, secramine (Pelish et al, 2006, Nat Chem Biol; 2(l):39-46), ML141 (Surviladze et al, "A Potent and Selective Inhibitor of Cdc42 GTPase," Probe Reports from the NIH Molecular Libraries Program [Internet], Bethesda (MD): National Center for Biotechnology Information (US); 2010), or an endothelin receptor antagonist, such as, for example, bosentan, ambrisentan, or derivatives thereof.
  • Bosentan an endothelin receptor antagonist, is indicated mainly for the treatment of pulmonary arterial hypertension (PAH) (see Rubin et al, 2002, N Engl J Med; 346(12): 896- 903). In 2007, bosentan was also approved in the European Union for reducing the number of new digital ulcers in patients with systemic sclerosis and ongoing digital ulcer disease.
  • PAH pulmonary arterial hypertension
  • TRACLEER is designated chemically as 4-tert-butyl-N-[6-(2- hydroxy-ethoxy)-5-(2-methoxy-phenoxy)[2,2]-bipyrimidin-4-yl]- benzenesulfonamide monohydrate, has the chemical formula C27H 3 iN 5 0vS,and the CAS Registry number 157212-55- 0.
  • a CDC42 inhibitor blocks CDC42 activation of endothelin type I and/or endothelin type II receptor.
  • a CDC42 inhibitor may be an endothelin (ET) receptor antagonist (ERA) and may blocks endothelin receptors.
  • ET endothelin
  • ERA endothelin receptor antagonist
  • Three main kinds of ERAs are know: selective ETA receptor antagonists (sitaxentan, ambrisentan
  • a CDC42 inhibitor is bosentan or a derivative thereof.
  • One or more additional therapeutic modalities may be administered along with one or more agents of the present disclosure.
  • the present invention the present invention
  • agents of the present disclosure may allow for the effectiveness of a lower dosage of other therapeutic modalities when compared to the administration of the other therapeutic modalities alone, providing relief from the toxicity observed with the administration of higher doses of the other modalities.
  • One or more additional therapeutic agents may be administered before, after, and/or coincident to the administration of agents of the present disclosure.
  • Agents of the present disclosure and additional therapeutic agents may be administered separately or as part of a mixture of cocktail.
  • an additional therapeutic agent may include, for example, an agent whose use for the treatment of Alport syndrome, kidney disease, kidney failure, and/or proteinuria is known to the skilled artisan.
  • an angiotensin-converting enzyme (ACE) inhibitor such as ramipril or anapril, may be administered.
  • ACE angiotensin-converting enzyme
  • treating can include therapeutic and/or prophylactic treatments. Desirable effects of treatment include preventing occurrence or recurrence of disease, alleviation of symptoms, diminishment of any direct or indirect pathological
  • agents of the present disclosure can be administered by any suitable means including, but not limited to, for example, oral, rectal, nasal, topical (including transdermal, aerosol, buccal and sublingual), vaginal, parenteral (including subcutaneous, intramuscular, intravenous and intradermal), intravesical, or injection into or around the tumor.
  • parenteral administration in an aqueous solution for example, the solution should be suitably buffered if necessary and the liquid diluent first rendered isotonic with sufficient saline or glucose.
  • aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous, intraperitoneal, and intratumoral administration.
  • sterile aqueous media that can be employed will be known to those of skill in the art. Some variation in dosage will necessarily occur depending on the condition of the subject being treated. The person responsible for
  • administration will, in any event, determine the appropriate dose for the individual subject.
  • preparations should meet sterility, pyrogenicity, and general safety and purity standards as required by the FDA. Such preparation may be pyrogen- free.
  • the inhibitor may be administered in a tablet or capsule, which may be enteric coated, or in a formulation for controlled or sustained release.
  • a formulation for controlled or sustained release Many suitable formulations are known, including polymeric or protein microparticles encapsulating drug to be released, ointments, gels, or solutions which can be used topically or locally to administer drug, and even patches, which provide controlled release over a prolonged period of time. These can also take the form of implants.
  • compositions of one or more of the inhibitors described herein may also include, for example, buffering agents to help to maintain the pH in an acceptable range or preservatives to retard microbial growth.
  • Such compositions may also include a pharmaceutically acceptable carrier.
  • pharmaceutically acceptable carrier refers to one or more compatible solid or liquid filler, diluents or
  • compositions of the present disclosure are formulated in pharmaceutical
  • Therapeutically effective concentrations and amounts may be determined for each application herein empirically by testing the compounds in known in vitro and in vivo systems, such as those described herein, dosages for humans or other animals may then be extrapolated therefrom. With the methods of the present disclosure, the efficacy of the administration of one or more agents may be assessed by any of a variety of parameters known in the art.
  • An agent of the present disclosure may be administered at once, or may be divided into a number of smaller doses to be administered at intervals of time.
  • an agent of the present disclosure may be administered twice a day, three times a day, four times a day, or more.
  • an agent of the present disclosure may be administered every other day, every third day, once a week, every two weeks, or once a month.at once, or may be divided into a number of smaller doses to be administered at intervals of time.
  • an agent of the present disclosure may be administered continuously, for example by a controlled release formulation or a pump.
  • dosage and duration of treatment is a function of the disease being treated and may be determined empirically using known testing protocols or by extrapolation from in vivo or in vitro test data. It is to be noted that concentrations and dosage values may also vary with the severity of the condition to be alleviated. It is to be further understood that for any particular subject, specific dosage regimens should be adjusted over time according to the individual need and the professional judgment of the person administering or supervising the administration of the
  • compositions and that the concentration ranges set forth herein are exemplary only and are not intended to limit the scope or practice of the claimed compositions and methods.
  • an "effective amount" of an agent is an amount that results in a reduction of at least one pathological parameter.
  • an effective amount is an amount that is effective to achieve a reduction of at least about 10%, at least about 15%, at least about 20%, or at least about 25%, at least about 30%>, at least about 35%, at least about 40%>, at least about 45%, at least about 50%>, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%), at least about 80%>, at least about 85%, at least about 90%>, or at least about 95%, compared to the expected reduction in the parameter in an individual not treated with the agent.
  • a subject includes, but is not limited to, humans and non-human vertebrates.
  • a subject is a mammal, particularly a human.
  • a subject may be an individual.
  • a subject may be an "individual,” “patient,” or “host.
  • a subject is an individual diagnosed with Alport syndrome. Diagnosis may be by any of a variety of means, including, but not limited to, family history, clinical presentation, pathological determination, and/or genetic testing.
  • Such as subject may be a male or a female.
  • Non-human vertebrates include livestock animals, companion animals, and laboratory animals.
  • Non-human subjects also include non-human primates as well as rodents, such as, but not limited to, a rat or a mouse.
  • Non-human subjects also include, without limitation, chickens, horses, cows, pigs, goats, dogs, cats, guinea pigs, hamsters, mink, and rabbits.
  • in vitro is in cell culture and “in vivo” is within the body of a subject.
  • pharmaceutically acceptable carrier refers to one or more compatible solid or liquid filler, diluents or encapsulating substances which are suitable for administration to a human or other vertebrate animal.
  • isolated refers to material that has been either removed from its natural environment (e.g., the natural environment if it is naturally occurring), produced using recombinant techniques, or chemically or enzymatically synthesized, and thus is altered “by the hand of man” from its natural state.
  • Alport glomerular disease is delayed onset and progressive, with onset generally occurring in the first decade of life.
  • Abnormal laminins (laminin 211 and laminin 111) progressively accumulate in the glomerular basement membranes (GBM) of mice dogs and people with Alport syndrome.
  • the present invention shows that laminin 211 activates focal adhesion kinase leading to downstream signaling through the NFkappaB transcription factor which results in maladaptive dysregulation of genes that drive the progression of Alport glomerular pathology.
  • the deposition of laminin 211 in the GBM is an important driver of glomerular pathogenesis in Alport syndrome.
  • the present invention also shows that laminin 211 is being deposited by mesangial cell filopodia that are invading the interface between the glomerular endothelial cells and the GBM. This important observation indicates that the activation of filopodial invasion of the GBM is an early event in glomerular disease initiation in Alport syndrome. Agents that can attenuate or arrest the activation of mesangial filopodial invasion will have important therapeutic application.
  • Cell culture studies show that filopodia formation involves dynamic actin microfilament remodeling that is regulated by the rho family of small GTPases. These structures emerge from lamellipodia, found at the leading edge of migrating cells.
  • Lamellipodia formation is regulated by activation of the Ras related C3 botulinum toxin substrate 1 (RACl) small GTPase, and filopodia formation is regulated by the activation of cell division control 42 homologue (CDC42) small GTPase.
  • Cross-talk between RAC 1 and CDC42 has been demonstrated.
  • This example determined that the mesangial process invasion into the capillary loops in Alport syndrome involves the activation of RACl and/or CDC42.
  • the autosomal recessive Alport mouse model (a COL4A3 gene knockout mouse on the 129 Sv/J background) was treated with the small molecule inhibitor for RACl, NSC 23766 (commercial available) at a
  • Kidneys were harvested at 6 weeks of age (at a stage when glomerular disease is well advanced and proteinuria is >300 mg/dl). One kidney was used for histology and the other for glomerular RNA isolation. The kidney cryosections were analyzed by dual immunofluorescence immunostaining for integrin a8 (a mesangial cell surface marker) and laminin a5 (a glomerular basement membrane marker).
  • this example identifies novel therapeutic targets for the treatment of Alport syndrome, namely agents that can block the activation of RAC1/CDC42 members of the rho family of small GTPases and thus prevent invasion of the glomerular capillary tufts by mesangial lamellipodial/filopodial processes.
  • mesangial process invasion By blocking mesangial process invasion, the deposition of laminin 211 in the GBM is abrogated, thus preventing the activation of maladaptive expression of proteins known to contribute to glomerular disease progression.
  • Alport syndrome results from mutations in type IV collagen COL4A3, COL4A4, or COL4A5 genes. Mutations in any of these genes results in the absence of all 3 in the GBM type IV collagen network due to an obligatory association to form heterotrimers. The result is a thinner and less crosslinked GBM collagen network resulting in delayed onset progressive glomerulonephritis. The molecular trigger for disease onset is unknown.
  • a comparative analysis of glomerular disease progression in Alport mice and CD151 knockout mice revealed a progressive irregular deposition of mesangial laminin 211 in the GBM.
  • biomechanical insult results in the induction of mesangial filopodial invasion of the glomerular capillary tuft leading to the irregular deposition of mesangial laminin 211 and the initiation mechanism of Alport glomerular pathology.
  • a comparative analysis of glomerular disease progression in Alport mice and CD151 knockout mice revealed a progressive irregular deposition of mesangial laminin 211 in the GBM.
  • Co-localization studies showed that the mesangial integrin ⁇ 8 ⁇ 1 also progressively accumulates in the capillary loops of both models as well as in human Alport glomeruli, indicating an invasion of the capillary loops by mesangial cell processes.
  • L-NAME salt-induced hypertension accelerated mesangial cell process invasion and laminin 211 accumulation, suggesting biomechanical strain plays a role in this mechanism.
  • Cultured mesangial cells showed reduced migratory potential when treated with either integrin linked kinase inhibitor, Racl inhibitors, CDC42 inhibitors, or by deletion of integrin al .
  • Alport syndrome is characterized by delayed onset progressive glomerulonephritis associated with sensorineural hearing loss and retinal flecks (Kashtan and Michael, 1996, Kidney Int; 50(5): 1445-1463).
  • the most common form (80%) is X-linked and caused by mutations in the type IV collagen COL4A5 gene (Barker et al, 1990, Science; 8; 248(4960): 1224-7).
  • the two autosomal forms of the disease account for the remaining 20% of Alport patients, and result from mutations in the COL4A3 and COL4A4 genes (Mochizuki et al, 1994, Nat Genet; 8(1):77- 81).
  • the a3(IV), a4(IV) and a5(IV) proteins form a heterotrimer and is assembled into a subepithelial network in the glomerular basement membrane that is physically and biochemically distinct from a subendothelial type IV collagen network comprised of l(IV) and a2(IV) heterotrimers (Kleppel et al., 1992, J Biol Chem; 267(6):4137-4142). Mutations in any one of the three type IV collagen genes that cause Alport syndrome results in the absence of all three proteins in the GBM due to an obligatory association to form functional heterotrimers (Kalluri and Cosgrove, 2000, J Biol Chem; 275(17): 12719-12724).
  • the net result for all genetic forms of Alport syndrome is the absence of the a3(IV) a4(IV) a5(IV) subepithelial collagen network, resulting in a GBM type IV collagen network comprised only of l(IV) and a2(IV) heterotrimers.
  • the podocytes are exposed to GBM that has an embryonic type IV collagen composition (Kalluri et al, 1997, J Clin Invest; 99(10):2470-2478; and Harvey et al, 1998, Kidney Int; 54(6): 1857-1866). This could result in altered cell signaling that may trigger the onset of the disease. It has been proposed this type of mechanism may account for the reactivation of laminin 111 expression in podocytes (Abrahamson et al., 2003, Kidney Int;
  • glomeruli from Alport mice have been shown to have elevated deformability relative to wild type glomeruli (Wyss et al, 2011, Am J Physiol Cell Physiol; 300:C397-C405), and salt-induced hypertension has been shown to accelerate glomerular disease progression in Alport mice (Meehan et al., 2009, Kidney Int; 76:968-976).
  • This example shows that deletion of laminin 211 in Alport mice ameliorates the mesangial process invasion of the glomerular capillary loops in Alport mice, demonstrating for the first time a functional role for GBM laminin 211 in Alport glomerular pathogenesis.
  • the cellular origin of GBM laminin 211 has not been previously determined.
  • This example shows that the source of GBM laminin 211 in Alport GBM is mesangial cell processes, which are invading the capillary tufts. Salt-mediated hypertension exacerbates this mesangial process invasion.
  • a knockout mouse for the integrin ⁇ 3 ⁇ 1 co-receptor CD 151 which results in reduced adhesion of podocytes pedicles to GBM laminin 521, also develops mesangial process invasion of the capillary loops with GBM deposition of laminin 211, demonstrating the same phenotype for a completely unrelated component of the capillary structural barrier.
  • the CD 151 knockout mouse model also shows accelerated glomerular disease progression in response to hypertension (Sachs et al, 2012, J Clin Invest; 122(l):348-58).
  • GBM laminin 211 in Alport mice is of mesangial origin. In the glomerulus, laminin 211 is normally found only in the mesangial matrix.
  • Figures 1A to 1C demonstrate mesangial distribution of laminin 211 in wild type mice, which is distinct from the glomerular basement membrane (collagen a3(IV)).
  • Figures ID to IF demonstrate the irregular distribution of laminin 211 in the GBM which appears to accumulate preferentially in irregularly thickened regions of the GBM (here the GBM is marked by immunostaining with antibodies specific for laminin a5).
  • the cellular source of the GBM laminin 211 has never been
  • Fig. 4 shows that salt-induced hypertension clearly accelerates the inundation of the glomerular capillary loops by mesangial processes as evidenced by the presence of integrin a8 immunopositivitiy in the GBM (Fig. 4D-4F).
  • al(IV)/a2(IV) and a3(IV)/a4(IV)/a5(IV) collagen to one comprised only of al(IV)/a2(IV) collagen.
  • the latter is thinner and known to contain fewer interchain disulfide crosslinks (Gunwar et al, 1998, J Biol Chem; 273(15):8767-75) which would intuitively be expected to result in increasing the elasticity of the glomerular filtration barrier.
  • a completely different model was examined that would also be expected to affect the elastic integrity of the glomerular filtration barrier, the CD151 knockout mouse.
  • CD151 is a tetraspanin co-receptor for integrin ⁇ 3 ⁇ 1 which functions to increase the affinity of integrin ⁇ 3 ⁇ 1 for its GBM ligand, laminin a5 (Nishiuchi et al, 2005, Proc Natl Acad Sci USA; 102(6): 1939-44). Deletion of CD151 results in glomerular disease with morphological changes in the GBM strikingly similar to Alport syndrome (Baleato et al., 2008, Am J Pathol;
  • pro-migratory responses will be activated in vitro by mechanically stretching cultured primary mesangial cells.
  • Primary cultured mesangial cells derived from 129 Sv/J mice, were subjected to cyclic cell stretching using the Flexcell system for 24 hours. Expression of several pro-migratory cytokines was quantified by real time RT-PCR. The results in Fig. 6 demonstrate that expression of both TGF- ⁇ and CTGF are significantly elevated in cells subjected to biomechanical stretching relative to cells cultured under identical conditions, but not subjected to stretch.
  • Fig. 7 shows that deletion of al integrin markedly reduces the dynamics of mesangial process invasion of the capillary tufts in Alport mice, consistent with the reduction in GBM laminin 211 deposition shown here and previously
  • Fig. 8B After 30 minutes, treated cells undergo a stark morphological change about half of the cells sprouting numerous filopodia (Fig. 8B(b), denoted by asterisks), that are easily discernable, blinded, in numerous replicate experiments.
  • Cells treated with LPS in combination with either the Racl inhibitor NSC 23766 or the CDC42 inhibitor ML 141 could not be distinguished in blinded experiments form untreated wild type mesangial cells (Figs. 8B(c) and 8B(d), respectively.
  • Fig. 7, Fig. 8, and Fig. 9 confirm that mesangial process invasion of the glomerular capillaries is a Racl -dependent process, and that Racl activation is attenuated by integrin al deletion both in vitro and in vivo. Furthermore, LPS activation of filopodia in wild type mesangial cells (but not in a 1 -null mesangial cells) involves both Racl and CDC42 activation, suggesting ⁇ integrin-dependent cross talk between the two small GTPases in the signaling complex.
  • Laminin 211 enhances mesangial cell migration and mesangial process invasion of the capillary loops.
  • a laminin a2-deficient mouse was crossed with the Alport mouse to produce a double knockout.
  • One effect of laminin o2 deficiency was a marked reduction of mesangial process invasion of the capillary loops (Fig. 10A). This indicates that laminin 211 might facilitate mesangial process invasion of the capillary loops.
  • cell migration assays were performed on either laminin 211 or laminin 521 (GBM laminin). Two different laminin preparations were used.
  • Integrin a8 was used as a specific mesangial cell surface marker to demonstrate that mesangial processes invade the capillary tufts and co-localize with laminin 211, a mesangial laminin.
  • Integrin a8 is expressed in mesangial cells, but not in other glomerular cell types (Hartner et al, 1999, Kidney Int; 56(4): 1468-80), and its expression is generally restricted to smooth muscle cells and neuronal cell types (Bossy et al, 1991, EMBO J; 10(9):2375-2385; and Schnapp et al, 1995, J Cell Sci; 108:537-544).
  • Alport mutations which can be either autosomal recessive (mutations in either COL4A3 or COL4A4 genes (Mochizuki et al, 1994, Nat Genet; 8(1):77-81)) or X-linked (mutations in COL4A5 (Barker et al, 1990, Science; 8;248(4960): 1224-7)) result in the absence of the collagen a3(IV)/a4(IV)/a5(IV) network from the GBM.
  • Lipopolysaccharide which activates both Racl and CDC42 in wild type mesangial cells, failed to activate Racl or CDC42 (Fig. 8B), and failed to activate actin cytoskeletal
  • Rho GTPases Classically, Racl activation is associated with lamellipodia formation and CDC42 activation is associated with filopodia formation (Nobes and Hall, 1995, Cell; 81(l):53-62). Recently, evidence for crosstalk between the two Rho GTPases has emerged (Zamudio-Meza et al, 2009, J Gen Virol; 90(Pt 12):2902-11). This phenomenon is likely regulated through the guanine nucleotide exchange factor ⁇ , which contains binding sites for both CDC42 and Racl (Chahdi et al, 2004, Biochem Biophys Res Commun; 317(2):384-9; and Chahdi et al, 2005, J Biol Chem; 280(l):578-84).
  • This example provides evidence for cross-talk between Racl and CDC42 in cultured mesangial cells regulating actin cytoskeletal rearrangement including: showing that treatment of mesangial cells with LPS, known to activate rapid actin cytoskeletal rearrangement (Bursten et al., 1991, Am J Pathol; 139(2):371-82), activates Racl in wild type mesangial cells (Fig. 8C(d)); showing that membrane localization of CDC42, a known prerequisite for its activation, is blocked by addition of RAC1 inhibitors coincident with LPS stimulation (Fig. 8C(a-c)); and showing that inclusion of either Racl inhibitors or CDC42 inhibitors upon stimulation of mesangial cell cultures with LPS blocks actin cytoskeletal rearrangements (Fig. 8B).
  • the example shows that wild type mesangial cells migrate more robustly when cultured on laminin 211 compared to laminin 521, and that primary mesangial cells from laminin a2-deficient mice show impaired migration relative primary wild type mesangial cells from age/strain matched mice (Figs. 9B and 9C).
  • mesangial matrix molecules are likely deposited in the GBM, and local action of mesangial cytokines (TGF- ⁇ and CTGF, for example) and MMPs might also contribute to the structural and functional properties of the Alport GBM (irregular thickening, splitting, permeability, etc). In addition, all of these events are very likely to influence podocyte cell health.
  • mesangial process invasion of the GBM is an important early event that precipitates glomerulosclerosis in Alport syndrome.
  • the observation of mesangial process invasion of glomerular capillary loops in human Alport glomeruli provides relevance for these observations to the human disease.
  • a better understanding of the activation process might reveal novel targets capable of preventing this event and arresting the Alport glomerular pathogenesis in its pre-initiated state.
  • mice All mice used in these studies were on pure genetic backgrounds. Autosomal recessive Alport mice were on the 129 Sv background. X-linked Alport mice were on the C57 Bl/6 background, laminin a2-deficient mice were on the 129 Sv background, integrin a 1 -null mice were on the 129 Sv background (Gardner et al, 1996, Dev Biol; 175(2):301-13), and CD151 knockout mice were on the FVB background (Takeda et al, 2007, Blood; 109(4): 1524- 32). All experiments were performed using strain/age-matched control mice. All animal studies were conducted in accordance to USDA approved standards and under the approval of the institutional IACUC. Every effort was made to minimize pain and discomfort.
  • Affinity purified rabbit anti-collagen a3(IV) antibodies were as previously described. Slides were rinsed with IX PBS and incubated with the appropriate Alexa Fluor donkey secondary antibodies at 1 :300 for 1 hour at room temperature. They were then rinsed again with IX PBS and mounted with Vectashield Mounting Medium with Dapi (Vector).
  • Transwell cell migration assays were performed as described by Daniel et al. ⁇ Lab Invest. 2012 Jun;92(6):812-26) with some modifications. 8 micron, 24-well plate control inserts (BD Bioscience, Bedford, MA) were coated overnight at 4°C with 100 ⁇ of 0.1% gelatin/PBS then washed IX with PBS. MES cultures were incubated in 1% FCS overnight, then 0.05% BSA-containing media for at least 8 hours, washed IX with PBS and carefully tryspinized to ensure a single cell suspension and limited "clumping" of cells.
  • Lipopolysaccharides (Sigma- Aldrich) was added to cells, incubated 1 hour, fixed in ice cold acetone for 5 minutes and allowed to air dry ⁇ 2 hours. Cells were stained with a 1 : 100 dilution of antibodies specific for CDC42 (10155-1-AP, ProteintechTM), and phalloidin (Molecular Probes) imaged. Untreated, LPS alone and LPS plus inhibitors treatments were repeated on two different derivations of primary mesangial cells with qualitatively consistent results.
  • Pull down assay Pull down experiments for Racl in mesangial cells were done using the Racl Activation Assay Bicochem Kit (BK035, Cytoskeleton Inc, CO) and according to manufacturer instructions with minor modifications. Briefly, 500-800 ⁇ g of protein lysates were incubated with 20 ⁇ of PAK-PBD beads for 1 hour at 4°C. Pull down samples and total protein lysates (30-50 ⁇ g of protein) were run in a 12% SDS-PAGE gel, transferred to PVDF
  • 129 Sv mice as described in the previous examples, five animals per group, are being dosed daily with bosentan at 10 mg/kg by oral gavage using a carboxymethylcellulose vehicle from 2 to 7 weeks of age.
  • the groups are: WT vehicle; Alport vehicle; WT Bosentan; and Alport Bosentan.
  • Proteinuria and blood urea nitrogen (BUN) will be measured.
  • Dual immunofluorescence immunostaining will be performed with antibodies for integrin alpha 8 and laminin alpha5, as well as integrin laminin alpha 2 and pFAK397. Glomerulosllerosis and fibrosis scores will be determined using histochemical stain (Masson's trichrome).
  • mice will show reduced proteinuria and BUN measures and reduced fibrosis and glomerulosclerosis scores. It is expected that integrin alpha 8 immunostaining in the glomerular capillary loops will be markedly reduced if not absent, and that laminin alpha 2 in the GBM will be markedly reduced with concomitant reduction of PFAK397 immunostaining in the podocytes (which may be absent).
  • a reduction in mesangial process invasion which will be ascertained by the dual immunostaining experiments, will indicate that the endothelin blockade directly effects the activation of this process, and link this effect to improved renal health.
  • the numbers will allow statistical analysis of the data.
  • Urine will be normalized to urinary creatinine levels by colorimetric microplate assay, and normalized urine samples will be assayed for albumin using a microplate ELISA assay. Blood urea nitrogen levels will be measured in plasma using a colorimetric microplate assay.
  • Cohort two These animals will be harvested at 20 weeks of age. This is an age where advanced renal disease is uniformly evident in untreated mice, but not end stage (which is typically 23-26 weeks). Animals will be anesthetized and then transcardially perfused first with PBS, then one kidney clamped off at the renal artery and the other transcardially perfused with Dynabeads for magnetic recovery of glomeruli and isolation of glomerular RNA.

Landscapes

  • Health & Medical Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Epidemiology (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Urology & Nephrology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

La présente invention concerne des procédés permettant de traiter le syndrome d'Alport chez un sujet par l'administration d'un agent qui peut bloquer l'activation des membres RAC1/CDC42 de la famille rho de petites GTPases. Lesdits agents comprennent, sans toutefois s'y limiter, le bosentan antagoniste des récepteurs de l'endothéline. Ladite administration prévient l'invasion des floculus glomérulaires par des processus lamellipodiaux/filopodiaux mésangiaux, bloque l'invasion des processus mésangiaux, abroge le dépôt de laminine 211 dans le GBM, et prévient l'activation de l'expression mal adaptée de protéines connues pour contribuer à la progression de la maladie glomérulaire.
PCT/US2013/032432 2012-08-17 2013-03-15 Inhibiteurs de rac1 pour le traitement de la maladie glomérulaire d'alport WO2014028059A1 (fr)

Priority Applications (3)

Application Number Priority Date Filing Date Title
EP13829878.1A EP2884988A4 (fr) 2012-08-17 2013-03-15 Inhibiteurs de rac1 pour le traitement de la maladie glomérulaire d'alport
US14/580,680 US9719981B2 (en) 2012-08-17 2014-12-23 RAC1 inhibitors for the treatment of alport glomerular disease
US15/631,454 US10545134B2 (en) 2012-08-17 2017-06-23 RAC1 inhibitors for the treatment of Alport glomerular disease

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US201261684566P 2012-08-17 2012-08-17
US61/684,566 2012-08-17
US201361764389P 2013-02-13 2013-02-13
US61/764,389 2013-02-13

Related Child Applications (1)

Application Number Title Priority Date Filing Date
US14/580,680 Continuation-In-Part US9719981B2 (en) 2012-08-17 2014-12-23 RAC1 inhibitors for the treatment of alport glomerular disease

Publications (1)

Publication Number Publication Date
WO2014028059A1 true WO2014028059A1 (fr) 2014-02-20

Family

ID=50101384

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2013/032432 WO2014028059A1 (fr) 2012-08-17 2013-03-15 Inhibiteurs de rac1 pour le traitement de la maladie glomérulaire d'alport

Country Status (2)

Country Link
EP (1) EP2884988A4 (fr)
WO (1) WO2014028059A1 (fr)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9719981B2 (en) 2012-08-17 2017-08-01 Father Flanagan's Boys' Home RAC1 inhibitors for the treatment of alport glomerular disease
WO2020020896A1 (fr) * 2018-07-25 2020-01-30 Boehringer Ingelheim International Gmbh Empagliflozine à utiliser dans le traitement du syndrome d'alport
WO2020257636A1 (fr) 2019-06-21 2020-12-24 Father Flanagan's Boys' Home Doing Business As Boys Town National Research Hospital Anticorps neutralisants dirigés contre l'endothéline humaine

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040186083A1 (en) * 2003-03-18 2004-09-23 Pharmacia Corporation Combination of an aldosterone receptor antagonist and an endothelin receptor antagonist and/or endothelin converting enzyme inhibitor
US20110212083A1 (en) * 2008-11-06 2011-09-01 University Of Miami Office Of Technology Transfer Role of soluble upar in the pathogenesis of proteinuric kidney disease
US20110236397A1 (en) * 2008-11-06 2011-09-29 University Of Miami Limited proteolysis of cd2ap and progression of renal disease

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040186083A1 (en) * 2003-03-18 2004-09-23 Pharmacia Corporation Combination of an aldosterone receptor antagonist and an endothelin receptor antagonist and/or endothelin converting enzyme inhibitor
US20110212083A1 (en) * 2008-11-06 2011-09-01 University Of Miami Office Of Technology Transfer Role of soluble upar in the pathogenesis of proteinuric kidney disease
US20110236397A1 (en) * 2008-11-06 2011-09-29 University Of Miami Limited proteolysis of cd2ap and progression of renal disease

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See also references of EP2884988A4 *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9719981B2 (en) 2012-08-17 2017-08-01 Father Flanagan's Boys' Home RAC1 inhibitors for the treatment of alport glomerular disease
US10545134B2 (en) 2012-08-17 2020-01-28 Father Flanagan's Boys' Home RAC1 inhibitors for the treatment of Alport glomerular disease
WO2020020896A1 (fr) * 2018-07-25 2020-01-30 Boehringer Ingelheim International Gmbh Empagliflozine à utiliser dans le traitement du syndrome d'alport
WO2020257636A1 (fr) 2019-06-21 2020-12-24 Father Flanagan's Boys' Home Doing Business As Boys Town National Research Hospital Anticorps neutralisants dirigés contre l'endothéline humaine

Also Published As

Publication number Publication date
EP2884988A4 (fr) 2016-04-20
EP2884988A1 (fr) 2015-06-24

Similar Documents

Publication Publication Date Title
Rojansky et al. Elimination of paternal mitochondria in mouse embryos occurs through autophagic degradation dependent on PARKIN and MUL1
US10545134B2 (en) RAC1 inhibitors for the treatment of Alport glomerular disease
Axelrod et al. Axl as a mediator of cellular growth and survival
US6180597B1 (en) Upregulation of Type III endothelial cell nitric oxide synthase by rho GTPase function inhibitors
Bhattacharya et al. Prominin-1 is a novel regulator of autophagy in the human retinal pigment epithelium
Zallocchi et al. α1β1 integrin/Rac1-dependent mesangial invasion of glomerular capillaries in Alport syndrome
US6423751B1 (en) Upregulation of type III endothelial cell nitric oxide synthase by agents that disrupt actin cytoskeletal organization
Chen et al. Hypoxia promotes pulmonary vascular remodeling via HIF-1α to regulate mitochondrial dynamics
US20190350961A1 (en) Compositions and methods for the treatment of aberrant angiogenesis
Peng et al. Mechano-signaling via Piezo1 prevents activation and p53-mediated senescence of muscle stem cells
KR20220120711A (ko) 폐혈관 질환 치료용 조성물 및 방법
WO2017208174A2 (fr) Méthodes de traitement d'une maladie à l'aide d'inhibiteurs de pfkfb3
Zheng et al. MicroRNA‑126 suppresses the proliferation and migration of endothelial cells in experimental diabetic retinopathy by targeting polo‑like kinase 4
Wang et al. microRNA‐454‐mediated NEDD4‐2/TrkA/cAMP axis in heart failure: Mechanisms and cardioprotective implications
WO2014028059A1 (fr) Inhibiteurs de rac1 pour le traitement de la maladie glomérulaire d'alport
Mou et al. Macrophage‐targeted delivery of siRNA to silence Mecp2 gene expression attenuates pulmonary fibrosis
Ma et al. Super enhancer-associated circular RNA-CircKrt4 regulates hypoxic pulmonary artery endothelial cell dysfunction in mice
Sun et al. Histone deacetylase inhibitors reduce cysts by activating autophagy in polycystic kidney disease
Guan et al. Experimental diabetes exacerbates autophagic flux impairment during myocardial I/R injury through calpain‐mediated cleavage of Atg5/LAMP2
Jin et al. Depletion of CUL4B in macrophages ameliorates diabetic kidney disease via miR-194-5p/ITGA9 axis
WO2015054612A1 (fr) Traitement de maladies auto-immunes médiées par des lymphocytes t organo-spécifiques
Sekar et al. Impairing Gasdermin D-mediated pyroptosis is protective against retinal degeneration
Devi et al. CARD-only proteins regulate in vivo inflammasome responses and ameliorate gout
JP2023520002A (ja) 細胞内タンパク質ミスフォールディング関連疾患およびリソソーム蓄積症におけるｐｉ４ｋ阻害物質の応用
AU2020375441A1 (en) Treatment of renal cystic disease

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 13829878

Country of ref document: EP

Kind code of ref document: A1

REEP Request for entry into the european phase

Ref document number: 2013829878

Country of ref document: EP

WWE Wipo information: entry into national phase

Ref document number: 2013829878

Country of ref document: EP

NENP Non-entry into the national phase

Ref country code: DE